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The transport of mineral and organic matter into the soil profile by Lumbricus rubellus Hoffmeister Timmenga, Hubert J. 1987

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THE TRANSPORT OF M I N E R A L AND ORGANIC MATTER INTO THE S O I L P R O F I L E BY LUMBRICUS RUBELLUS H O F F M E I S T E R by HUBERT J . TIMMENGA L a n d b o u w k u n d i g I n g e n i e u r , W a g e n i n g e n , 1981 A T H E S I S SUBMITTED IN P A R T I A L F U L F I L M E N T OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY i n S o i l S c i e n c e THE F A C U L T Y OF GRADUATE S T U D I E S D e p a r t m e n t o f S o i l S c i e n c e We a c c e p t t h i s t h e s i s a s c o n f o r m i n g t o t h e r e q u i r e d s t a n d a r d THE U N I V E R S I T Y OF B R I T I S H COLUMBIA 13 S e p t e m b e r 1987 © HUBERT J . TIMMENGA, 1987 k 6 In presenting this thesis in partial fulfilment of the requirements for an advanced degree at the University of British Columbia, I agree that the Library shall make it freely available for reference and study. I further agree that permission for extensive copying of this thesis for scholarly purposes may be granted by the head of my department or by his or her representatives. It is understood that copying or publication of this thesis for financial gain shall not be allowed without my written permission. Department of The University of British Columbia 1956 Main Mall Vancouver, Canada V6T 1Y3 DE-6(3/81) ABSTRACT The biology and ecology of the earthworm Lumbricus rubellus Hoffmeister, 1843, and i t s effects on the turn-over of organic matter and s o i l are not well known. To gather t h i s information, the ingestion and egestion rates were measured using a l i t t e r b a g technique and the transport of organic matter was quantified with a newly developed method, using s o i l columns to which 1 4C l a b e l l e d plant material was added. The feeding habits of the worm were p o s i t i v e l y influenced by temperature in wet s o i l s (> -15m of water) and were negatively influenced in dry s o i l (< -15. m of water). The t o t a l egestion rate changed from 0.3 g.g-'.day"1 at 5 °C to 1.0 g.g~ 1.day~ 1 at 20° C in moist s o i l (- 5 m of water). The egestion rate at medium range temperatures, 10 and 15° C, was less affected by drought stress than at 5 and 20 °C. The egestion rate of carbon was a more stable parameter than the t o t a l egestion rate, and ranged from approximately 20 mg.g-1.day"1 at 5 °C, to 50 mg.g- 1.day _ 1 at 20 °C. The moisture and temperature effects were apparent in the Q 1 0 of the t o t a l egestion rate and of the egestion rate of carbon. The Q 1 0 ranged from 1.66 in wet s o i l s to 3.27 in dry s o i l s in the 5-15 °C interval and from 1.98 to 0.32 in the 10-20 °C range. For the egestion rate of carbon, the Q 1 0 i i ranged from 1.92 to 3.21 and from 1.28 to 0.47, respectively. The body water content of the worm varied considerably with the s o i l water p o t e n t i a l , and reached a maximum l e v e l of 5.5 kg.kg" 1 (dwt) between -15 metres of water and -30 metres of water. When under drought stress, worms stopped ingesting large quantities of s o i l , switched to a diet high in organic matter and lowered th e i r a c t i v i t y . In the 1"C column experiment, the t o t a l cast production was s i g n i f i c a n t l y related to depth. L. rubellus produced 15 % of the cast on the surface of the s o i l , 46 % in the 0-5 cm layer, 22 % in the 5-10 cm layer and 16 % in the 10-15 cm layer. Independent calculations from a) the uptake of 1 f tC labe l l e d carbon in earthworms, b) removal of l i t t e r from the surface and c) 1"C label recovered from cast, showed that the worms ingested 78-82 % of the offered organic matter as shoot l i t t e r and 18-22 % as root l i t t e r . 1"C orig i n a t i n g from shoot and root l i t t e r was recovered in casts throughout the p r o f i l e , indicating that the worms mixed food from a l l layers. i i i The t o t a l egestion rate found in the column experiment was 5.2 times higher than was found in the l i t t e r b a g technique under comparable conditions (2.34 vs 0.45 g.g" 1.day" 1). The egestion rate of carbon was similar in both techniques (37.1 vs. 46.1 mg.g~1.day~ 1 , 10 °C). In preliminary l i t t e r b a g t r i a l s , i t was found that L. rubellus egested 15.5 mg.g~1.day~1 of carbon (5 °C) for each of four food types offered. The 5 °C temperature t r i a l of the l i t t e r b a g technique, showed a similar amount of carbon egested. It was concluded that the worm needed a constant amount of carbon to provide nutrients and energy, of which a part or a l l may originate from ingested microorganisms. Based on the d i s t r i b u t i o n of cast in the p r o f i l e and the feeding strategies of L. rubellus, i t was concluded that t h i s earthworm cannot be c l a s s i f i e d as an epigeic worm. A new strategy class was proposed: eurygeic worms, earthworms l i v i n g in the l i t t e r - s o i l interface, mixing organic matter into the p r o f i l e and mineral s o i l into the l i t t e r layer. Based on the l i t e r a t u r e and results from the present study, a computer model was developed to simulate the longterm ef f e c t s of earthworms on an a g r i c u l t u r a l s o i l system. Simulations of the mixing of s o i l and organic matter in a l i m i t e d - t i l l a g r i c u l t u r a l system, showed that earthworms i v negatively affected the accumulation rate of surface l i t t e r and p o s i t i v e l y affected the organic matter content of the mineral s o i l . The model can be used to predict the trends in organic matter in s o i l s , important in s o i l conservation, mine reclamation and reforestation. v TABLE OF CONTENT ABSTRACT i i TABLE OF CONTENT V 1 LIST OF TABLES X LIST OF FIGURES x i i LIST OF ABBREVIATIONS x i i i LIST OF SPECIES NAMES xiv ACKNOWLEDGEMENTS x v I. INTRODUCTION 1 II. LITERATURE REVIEW 5 A. THE ROLE OF EARTHWORMS IN THE TURN-OVER OF ORGANIC MATTER AND SOIL 5 1. Ecological strategies and c l a s s i f i c a t o n of earthworms 5 2. Earthworm food 9 3. Impact of earthworms on the s o i l system 14 a. E f f e c t s of earthworms on s o i l structure and f e r t i l i t y 14 b. E f f e c t s of earthworms on i n f i l t r a b i l i t y 14 c. The mixing of organic matter into the p r o f i l e 15 d. Surface cast production 17 e. Parameters a f f e c t i n g the cast production of earthworms 18 B. LUMBRICUS RUBELLUS HOFFMEISTER, 1843 21 1 . D i s t r i b u t i o n 21 2. Reproduction 24 3. Food sources and eating habits 24 4. Respiration 25 5. Egestion and s o i l turn-over 25 6. Ecological c l a s s i f i c a t i o n and strategies 26 I I I . THE EFFECTS OF SOIL MOISTURE AND SOIL TEMPERATURE ON THE EGESTION AND INGESTION RATES OF LUMBRICUS RUBELLUS HOFFMEI STER 28 A. MATERIALS AND METHODS 28 1 . Introduction 28 2. Site description 28 3. F i e l d data c o l l e c t i o n 29 vi . 4. Earthworms used in the experiments 30 5. Sample c o l l e c t i o n 30 6. Litterbag technique 31 7. Water content of the worm 35 8. Chemical analysis 36 9. Calculations and S t a t i s t i c s 36 B. RESULTS AND DISCUSSION 36 1 . Litterbag technique 36 2. Calculations and s t a t i s t i c s 37 a. The grouping of the moisture and temperature levels 37 b. Calculation of the ingestion rates 37 c. S t a t i s t i c a l analysis 37 d. Curve f i t t i n g 39 3. Egestion rates of s o i l and organic matter 39 4. Egestion rate of carbon 43 5. Q 1 0 values of the a c t i v i t y of earthworms 45 6. Faecal organic matter 47 7. Faecal water content 49 8. Ingestion rates 50 9. Worm size 55 10. S o i l temperature and s o i l moisture in the f i e l d 55 11. Comparison of ingestion and egestion rates to l i t e r a t u r e data 56 12. Drought-survival strategies of Lumbricus rube 11 us 59 a. The water content of earthworms ... 59 b. Feeding behavior of the worm as related to the body water content 63 13. Testing of assumption 67 IV. THE TRANSPORT OF ORGANIC MATTER INTO THE SOIL PROFILE BY LUMBRICUS RUBELLUS 68 A. MATERIALS AND METHODS 68 1 . Introduction 68 2. Animals used in the column experiment 68 3. S o i l s used in the column experiment .... 69 4. Clover used in the column experiment ... 69 a. Production of clover 69 b. Radiolabelling of clover 70 5. Experimental Set-up 71 6. Experimental design and s t a t i s t i c s 73 a. Experimental design 73 b. S t a t i s t i c s 74 v i i 7. Sample preparation 74 8. Chemical Analysis 75 B. RESULTS AND DISCUSSION 76 1 . Production of clover 76 2. 1*C0 2 fumigation of the clover 76 3. Observations on materials and techniques used in the column experiment 77 a. Micro-arthropods in the s o i l 77 b. Plant material 78 c. Experimental temperature and s o i l moisture content 78 d. Diffusion method for carbon analysis 79 4. Airflow above the columns 79 5. S o i l water potentials in the columns... 80 6. Recovery of 1"C a c t i v i t y from the samples 80 a. Specific a c t i v i t y of the recovered materials 80 b. Total a c t i v i t y of recovered materials 83 c. 1 4C a c t i v i t y recovered from the bulk s o i l 83 7. Respiration and decomposition 85 a. Respiration 85 b. Weight loss of clover material .... 88 8. Recovery of casts from the s o i l columns 89 a. Description of the burrows 89 b. Description of casts 91 9. D i s t r i b u t i o n of casts 91 10. Egestion rates 93 1 1 . Organic matter 95 a. D i s t r i b u t i o n of organic carbon in the cast 95 b. 1 WC a c t i v i t y in casts as related to depth 97 c. Calculation of the use of added organic matter by the earthworms 98 12. Testing of assumption 100 EARTHWORM SIMULATION MODELS 101 A. SIMULATION MODELS DESCRIBING THE DYNAMICS OF SOIL MIXING BY EARTHWORMS 101 1 . Introduction 101 2. Published earthworm models 101 3. "MIXER", a new conceptual model describing earthworm a c t i v i t y in s o i l systems 104 a. Introduction 104 v i i i b. S o i l layers in the model 105 c. Population dynamics 105 d. Earthworm food 107 e. Cast production 109 f. Flow-diagram for MIXER 109 B. F-MIXER, A SIMULATION MODEL FOR SOIL MIXING BY EARTHWORMS 111 1. Introduction ., 111 2. A brief description of FORCYTE 113 3. Description of the FORCYTE version of MIXER 114 4. Comparison of F-MIXER to MIXER 116 5. Simulation 118 6. Improvements needed in F-MIXER 124 VI. GENERAL DISCUSSION 126 VII. SUMMARY AND CONCLUSIONS 135 BIBLIOGRAPHY 139 APPENDIX 1. Diagnosis of Lumbricus rubellus 153 APPENDIX 2-A. Diagram of the fumigation set-up 154 APPENDIX 2-B. Diagram of the s o i l column set-up 155 APPENDIX 2-C. S o i l moisture and s o i l temperature on Westham Island 156 APPENDIX 2-D. The population of L. rubellus on Westham Island 157 APPENDIX 2-E. Retention curve of Crescent series s o i l . . 158 APPENDIX 3-A. Length of l i t t e r b a g experiments 159 APPENDIX 3-B. Food choices of L. rubellus 160 APPENDIX 3-C. Size and age of L. rubellus, as related to the egestion rate 162 APPENDIX 4. The egestion rate of A. Chlorotica 165 APPENDIX 5. Turn-over of s o i l and organic matter calculated from the l i t t e r b a g technique 167 APPENDIX 6. Detailed description of F-MIXER 170 i x LIST OF TABLES Table Page Table 1. The range of conditions r e s t r i c t i n g the d i s t r i b u t i o n of L. rubellus 23 Table 2. F-values for interactions and contrasts; ingestion and egestion rates of L. rubellus 41 Table 3. Egestion rates for L. rubellus, p r o b a b i l i t y of data points being d i f f e r e n t 42 Table 4. The comparison of the t o t a l egestion rate with the egestion rate of carbon for four d i f f e r e n t temperatures and moisture contents 44 Table 5. The Q 1 0 of L. rubellus, calculated from the egestion rate, for di f f e r e n t moisture and temperature ranges 46 Table 6. The percentage of organic matter ingested by L. rubellus, calculated from IOM and ITOT and measured in the faeces 48 Table 7. Ingestion rate for organic matter, p r o b a b i l i t y of data points being d i f f e r e n t 52 Table 8. Ingestion rate for s o i l , p r o b a b i l i t i e s of data points being d i f f e r e n t 54 Table 9. The dry weights of the earthworms used in the experiments 55 Table 10. Some ingestion and egestion rates for temperate and t r o p i c a l earthworms 57 Table 11. Values of Mann-Whitney U-test, indicating differences in body water content 61 Table 12. The water content of s o i l and clover straw incubated at 20 °C 66 Table 13. 1"C A c t i v i t y in a i r samples measured during fumigation 77 Table 14. Specific 1"C a c t i v i t i e s of materials recovered from the s o i l columns 82 Table 15. Total 1 4C a c t i v i t y recovered from materials in the s o i l columns 84 x Table 16. F-values for contrasts of t o t a l cast, recovered from columns, after incubating earthworms with l a b e l l e d clover shoot or root material added. xi LIST OF FIGURES F i g u r e Page F i g u r e 1. The r e d i s t r i b u t i o n of s o i l and o r g a n i c m a t t e r i n the s o i l p r o f i l e by earthworms 7 F i g u r e 2. The e g e s t i o n r a t e ( g . g " 1 . d a y " 1 ) of L. rubellus r e l a t e d t o the s o i l temperature and t h e s o i l m o i s t u r e 40 F i g u r e 3. The i n g e s t i o n r a t e of o r g a n i c m a t t e r f o r L. rubel I us 51 F i g u r e 4. The i n g e s t i o n r a t e of s o i l f o r L. rubellus 53 F i g u r e 5. The body water c o n t e n t of L. rubellus, i n c u b a t e d a t d i f f e r e n t s o i l m o i s t u r e p o t e n t i a l s (19 °C) .60 F i g u r e 6. The body water c o n t e n t ( k g . k g " 1 , dwt) and the e g e s t i o n r a t e ( g . g ~ 1 . d a y " 1 ) of a d u l t L. rubellus a t 20 °C 65 F i g u r e 7. P r e s s u r e p o t e n t i a l s i n the s o i l column d u r i n g the i n c u b a t i o n 81 F i g u r e 8. Decrease i n 1"C a c t i v i t y (% f i r s t measurement) of C 0 2 r e l e a s e d t h rough r e s p i r a t i o n 86 F i g u r e 9. D i s t r i b u t i o n of c a s t ( % of t o t a l c a s t per column) a c c o r d i n g t o depth 94 F i g u r e 10. D i s t r i b u t i o n of Carbon (% Carbon) i n c a s t a c c o r d i n g t o depth 96 F i g u r e 11. Flow diagram f o r the earthworm model MIXER. ..110 F i g u r e 12-A. R e s u l t s of the s i m u l a t i o n of F-MIXER, the a c c u m u l a t i o n of s u r f a c e l i t t e r 121 F i g u r e 12-B. R e s u l t s of the s i m u l a t i o n of F-MIXER, the o r g a n i c m a t t e r c o n t e n t of l a y e r I I 121 F i g u r e 12-C. R e s u l t s of the s i m u l a t i o n of F-MIXER, the i n g e s t i o n of s o i l 122 F i g u r e 13. Earthworm s t r a t e g i e s as p e r c e i v e d from Bouche (1977) w i t h a new c l a s s added 132 x i i LIST OF ABBREVIATIONS Upper mineral s o i l horizon, intermixed with organic matter Subsoil horizons Body water content of earthworms (kg.kg- 1 dwt) Choice of food by earthworms, based on food quality disintegrations per minute Dry weight Egestion, Egestion rate (g.g- 1.day" 1 dwt) Faeces water content of earthworms (kg.kg' 1 dwt) Ingestion, Ingestion rate (g.g- 1 .day 1 dwt) Ingestion rate of organic matter (g.g - 1 .day 1 dwt) Ingestion rate of mineral s o i l (g.g- 1.day" 1 dwt) Total ingestion rate (g.q- 1.day" 1 dwt) L i t t e r layer on top of mineral s o i l Moisture content of the s o i l Organic Matter Site quality in r e l a t i o n to earthworm populations Temperature x i i i LIST OF SPECIES NAMES Al I ol obophora caliginosa Savigny A. chlorolica Savigny A. I onga Ude A. nod ur na Evans A. rosea Savigny A. luberculata Eisen Aporreel odea turgida Eisen Cryptodrilus fasti gat us Fletcher Dendrobaena depressa Rosa D. octraedra Savigny D. pi at yura Fitzinger Dichogaster agiIis Omodeo and Vai l l a u d Eisenia eiseni Lev in sen E. foetida Savigny E. nordenshi oldi Eisen Eiseni el I a letraedra Savigny Lumbricus castaneus Savigny L. festivus Savigny L. rubellus Hoffmeister L. terreslris Linnaeus Millsonia anomal a Omodeo M. I ami i oana Omodeo and Vai l l a u d Me gas col ex eel mi siae Jamison Microscolex dubius Fletcher Nicodrilus velox Bouche Octolasion I act eum Orley O. tytraeum Savigny Pheretima alexandri Beddard x i v ACKNOWLEDGEMENT S I thank my s u p e r v i s o r , Dr. Les L a v k u l i c h , and the members of my committee, Drs. Shannon Berch, A r t Bomke, Ken H a l l and e s p e c i a l l y V a l i n M a r s h a l l , f o r t h e i r e f f o r t and guidance. I g r a t e f u l l y acknowledge the a s s i s t a n c e of many f a c u l t y members at UBC. Dr. Tony Glas s (Department of Botany) k i n d l y accommodated my 1"C experiment i n h i s l a b and provided t e c h n i c a l a s s i s t a n c e . Without h i s h e l p and that of h i s s t a f f , the second part of my t h e s i s would have been g r e a t l y delayed. Dr. Jim S h e l f o r d and Mr. G i l l e s Galzy (Department of Animal Science) allowed me to use t h e i r r a d i o - i s o t o p e f a c i l i t i e s . Dr. George Eaton (Department of P l a n t Science) helped me with s t a t i s t i c a l a n a l y s e s . Dr. Hamish Kimmins and Mr. Kim S c o u l l a r ( F a c u l t y of F o r e s t r y ) helped me i n shaping the computer s i m u l a t i o n model to f i t FORCYTE and programming the model. Mr. Bernie Von S p i n d l e r and Mr. Peter Synadinos ( S o i l Science) helped me by d i s c u s s i n g and s o l v i n g equipment problems. Mr. Mike Curran (Department of S o i l Science) read and d i s c u s s e d the manuscript and made many h e l p f u l s u g g e s t i o n s . I am a l s o g r a t e f u l to Dr. Alan C a r t e r under whose guidance t h i s work was i n i t i a t e d and f o r s h a r i n g h i s experience with the l i t t e r b a g technique. He p r o v i d e d f i e l d data on the xv earthworm population in a f i e l d on Westham Island. Mr. Hugh Reynolds kindly allowed me to use a part of his farm as an experimental pl o t . I e s p e c i a l l y thank my wife, Yme, for her patience and understanding in dealing with my changing moods. Without her support, i t would have been very d i f f i c u l t to f i n i s h t h i s di ssertat ion. xvi I . INTRODUCTION S o i l organisms have many functions in the process of recycling nutrients and the d i r e c t i o n of energy flow in the s o i l . As decomposers and shredders of plant material, they release nutrients to the s o i l which become available to growing plants (Wallwork, 1970; Richards, 1972). The most v i s i b l e impacts of s o i l animals on the s o i l system result from the a c t i v i t i e s of the larger animals such as earthworms and millipedes. Earthworm a c t i v i t y in the f i e l d , as w i l l be discussed in the l i t e r a t u r e review, i s generally quantified by the c o l l e c t i o n of surface casts or by measuring the effects earthworms have on plant production. Because earthworms may cast below the surface of the s o i l , i t i s important to study the t o t a l cast production of a species and the d i s t r i b t i o n of i t s casts in the p r o f i l e , before predictions can be made about the role played in the s o i l system. A few species such as Lumbricus terrestris and Eisenia foetidai have been widely studied, but other species have not been investigated. The ecology of Lumbricus rubellus Hoffmeister, 1843, a pioneer species (Reynolds, 1976) i s reviewed herein. This t S c i e n t i f i c names, c i t e d in t h i s thesis are those reported by the authors. Some names w i l l not r e f l e c t modern concepts of earthworm nomenclature. 1 INTRODUCTION / 2 earthworm may play an important role in s o i l management and reclamation as a s o i l mixer and l i t t e r shredder and as a colonizer of young s o i l s . The objectives of t h i s thesis are: 1. to study the r e d i s t r i b u t i o n of organic matter in the s o i l by the earthworm L. rubellus: a. to quantify the moisture and temperature e f f e c t s on the egestion and ingestion rates of the worm; b. to quantify the cast production related to s o i l depth; c. to quantify the r a t i o of shoot l i t t e r to root l i t t e r in the diet of the earthworm; 2. to develop a simulation model to simulate the long-term change in organic matter content in the s o i l through earthworm a c t i v i t y . The main hypothesis is that L. rubellus i s an epigeic worm, as suggested by Bouche (1977), l i v i n g in the l i t t e r layer and feeding and casting in t h i s layer. It i s assumed that s o i l temperature and s o i l moisture content have no s i g n i f i c a n t influence on the worm a c t i v i t y (e.g. ingestion or egestion rate) and that L. rubellus uses only surface l i t t e r and casts only on the surface of the s o i l . INTRODUCTION / 3 In this thesis the role of L. rubellus in the r e d i s t r i b u t i o n of organic and mineral matter in the s o i l p r o f i l e w i l l be discussed based on the res u l t s of two experiments. In the f i r s t experiment, the influence of s o i l temperature and s o i l moisture on the a c t i v i t y (e.g. ingestion or egestion rate) of the earthworm was quantified with l i t t e r b a g experiments. In the second, the r e d i s t r i b u t i o n of organic matter in the s o i l p r o f i l e was studied in a newly developed column experiment, using 1 WC la b e l l e d clover. Several experiments were conducted before the main l i t t e r b a g experiment was executed. The results of these preliminary experiments support the arguments presented in this thesis, although they were primarily conducted to refine the l i t t e r b a g technique. Although the thesis i s concerned with short-term experiments describing egestion and ingestion rates and transport of organic matter, i t was f e l t that a simulation model would provide insight to the long-term effects of earthworms on the s o i l system. In the l a s t part of the thesis, a conceptual model, MIXER, is developed from l i t e r a t u r e data. It i s a non-specific, multi-species model, describing the parameters a f f e c t i n g the turn-over of mineral and organic, matter by earthworms. INTRODUCTION / 4 From this conceptual model and from the results found in the experiments, a computer simulation model was developed. This model simulates the movement of s o i l and organic matter in the s o i l p r o f i l e . Although free-standing, this simulation model, F-MIXER, was designed to be included in the ecosystem model FORCYTE (Kimmins, 1986). F-MIXER neither simulates the da i l y , weekly or monthly growth nor the s o i l turn-over of a population of earthworms, but describes long-term trends in organic matter in di f f e r e n t s o i l layers, resulting from earthworm a c t i v i t y . Results of a 45 year trend-simulation in an a g r i c u l t u r a l n o - t i l l cropping system are discussed. I I . LITERATURE REVIEW A. THE ROLE OF EARTHWORMS IN THE TURN-OVER OF ORGANIC MATTER AND SOIL 1. E c o l o g i c a l s t r a t e g i e s and c l a s s i f i c a t o n of earthworms Earthworm a c t i v i t y i s v i s i b l e in the f i e l d because of accumulations of cast on the surface. To quantify the role of earthworms in the s o i l system, i t i s necessary to categorize the b i o l o g i c a l and ecological c h a r a c t e r i s t i c s of the worms and then c l a s s i f y the animals with similar c h a r a c t e r i s t i c s into groups. Earthworms have been c l a s s i f i e d in many di f f e r e n t ways: by morphological c h a r a c t e r i s t i c s , d i s t r i b u t i o n , r e l a t i o n to human a c t i v i t i e s and ecological strategies. Bouche (1977) proposed the following c l a s s i f i c a t i o n , based on morphological c h a r a c t e r i s t i c s , for European lumbricids with d i f f e r e n t ecological strategies: Epigeic worms: pigmented, small worms, not good burrowers, slow eaters, moderately sensitive to l i g h t , have fast maturation rates and are found in environments r i c h in organic matter. These worms are highly reproductive and survive adverse conditions by producing large amounts of cocoons. They l i v e and feed in the l i t t e r layer. Examples of 5 LITERATURE REVIEW / 6 North American earthworms that are recognized as epigeics include Eisenia foetida and De ndr obaena oclaedra and an indigenous undescribed species in the genus Arct iost rol us (Spiers et a l . , 1986). Endogeic worms: nonpigmented burrowers of medium size , sensitive to l i g h t . These "worms are s o i l dwellers and s o i l eaters. Adverse conditions are survived in a quiescent state. These worms have a limited reproductive capacity, they l i v e and feed in the mineral s o i l . Aporreclodia turgida and Octolasion tyriaeum are included in the endogeic class (Shaw and Pawluk, 1986) Anecic worms: strong, muscular, deep-burrowing pigmented worms, moderately sensitive to l i g h t . They are large and slow growing. These worms have a low reproductive rate and may survive adverse conditions in a real diapause. They are deep burrowers and may feed on the surface. L. terrestris i s included in this c l a s s . Some species do not f a l l c l e a r l y into any one of the categories. The categories should therefore not be seen as r e s t r i c t i v e . Figure 1 shows the flow of materials in the s o i l p r o f i l e as related to the a c t i v i t i e s of worms of the three main categories. The influence of human a c t i v i t i e s on earthworm d i s t r i b u t i o n was described by J u l i n ' s system of c l a s s i f i c a t i o n , as LITERATURE REVIEW / 7 B C Figure 1. The r e d i s t r i b u t i o n of s o i l and organic matter in the s o i l p r o f i l e by earthworms. Figure drawn from l i t e r a t u r e data, Ah = s o i l layer with b i o l o g i c a l a c t i v i t y , BC = subsoil, L = L i t t e r , > turn-over of s o i l , - - -> organic matter used as food (after Bouche, 1977). LITERATURE REVIEW / 8 described and modified by Reynolds et a l . (1974). In order of increasing dependence on human influences, earthworms were c l a s s i f i e d as: hemerophobes, hemerodiaphores, hemerophiles and hemerobionts. The r-selection and K-selection concepts are generally accepted in ecological theory (Lee, 1985; Satc h e l l , 1980). The terms r- and K-selection originated from the Verhulst-Pear1 equation 6N/6t = r(1 - N/K)N which relates population, N, over time to the i n t r i n s i c rate of natural increase, r, and the carrying capacity, K, of the environment. Two selections were recognized: r - s e l e c t i o n , a selection for maximum population growth in uncrowded, unstable environments, and K-selection, a selection for competitive a b i l i t i e s in stable environments. Satchell (1980) used the strategy concept to describe earthworm survival strategies. The epigeics were c l a s s i f i e d as r-worms (high reproduction, small, fast growing) and the anecics as K-worms ( e f f i c i e n t s u r v i v a l , large worms, low reproduction). The mentioned strategies represent the l i m i t s of a continuum and many worm species such as L. rubellus, L. terrestris and AlIolobophora chlorotica show behavior that LITERATURE REVIEW / 9 includes elements of both strategies. D i f f i c u l t i e s may be encountered in placing earthworms in appropriate categories or strategies. To c l a s s i f y earthworm species according to the role they play in the ecosystem, an indepth study of each species must be done. 2 . Earthworm food As plant l i t t e r decomposes, i t i s invaded by a succession of microorganisms. Parasites permeate senescing tissue; bacteria, fungi, myxobacteria and protozoa, a l l l i v i n g on simple carbohydrates, follow. Cellulose and l i g n i n decomposers, mostly fungi, increase in population in lat e r stages of decomposition (Dickinson and Pugh, 1974). Different earthworm species ingest organic matter in di f f e r e n t stages of decomposition. L. rubellus for example, ingests r e l a t i v e l y undecomposed organic matter, while A. caliginosa feeds on well-decomposed material (Piearce, 1978). The decomposition of plant material, including the breakdown of calcium oxalate that plants contain, by bacteria and actinomycetes has been described by Cromack et a l . (1977). They noted high calcium levels in fungal hyphae and in fungal feeding o r i b a t i d mites (up to 18 %) and found calcium oxalate decomposing microorganisms, mostly LITERATURE REVIEW / 10 actinomycetes, in the gut of L. rubellus by plati n g out the gut content. It i s not certain from their data whether the microorganisms actually l i v e d in the gut or simply survived the digestive process. Relatively undecomposed organic matter may contain high leve l s of calcium oxalate. This substance may be broken down by fungi and act inomycetes and the calcium may. be taken up by these organisms and by earthworms feeding on them. This phenomenum was also reported by Spiers et a l . , (1986). They found decreasing amounts of calcium oxalate c r y s t a l s over the length of the gut of Arct i ost rot us spp, and suggested that the calcium oxalate was used by i n t e s t i n a l microorganisms for their energy needs. The calcium was then absorbed by the earthworm and excreted in the gut as calcium carbonate. Earthworms, such as L. rubellus, which feed on not well-decomposed organic matter, have active calcium secreting glands (Piearce, 1972). Earthworms ingest large quantities of s o i l and organic matter, but assimilate only a small amount of the carbon ingested (Uvarov, 1982; Bolton and P h i l l i p s o n , 1976). Baylis et a l . (1986) reported on a 3 2 P study in which feeding of several earthworm species, including L. rubellus, on l i v i n g clover roots was observed. It i s not clear from their paper LITERATURE REVIEW / 11 whether the worms actually fed on the roots, or ingested s o i l from the rhizosphere. The rhizosphere may contain large amounts of microorganisms, feeding on the root exudates. The label might have been transported to the microbes through root exudates. Satchell (1983) reported that fungi were s e l e c t i v e l y destroyed and ingested by L. i errest ri s and that algae and protozoa were digested by L. rubellus. Hartenstein et a l . (1981) found that when specimens of Eisenia foetida were fed with horse manure or sewage sludge, both containing large numbers of microorganisms, the worms ingested a smaller amount of food and gained weight s i g n i f i c a n t l y faster than control worms, fed with a s o i l mixture. Live bacteria and fungi from axenic cultures contributed to a greater weight gain of worms than did dead bacteria (Neuhauser et a l . , 1980 b). Flack and Hartenstein (1984) reported that E. foetida grew well on three species of protozoa, and 22 species of bacteria. Also, l y o p h i l i z e d microorganisms were successfully used as earthworm food. G r i t , however, including sand or ashed loam, was necessary for optimum growth. Heungens (1969) reported that when a conifer-needle mixture was treated with fungicides, less decrease in mixture depth was seen due to earthworm shredding (mainly by Dendrobaena spp.), than without d i s i n f e c t i o n . Either the fungicide was LITERATURE REVIEW / 12 extremely toxic to earthworms, or no fungal hyphae were available as earthworm food after the d i s i n f e c t i o n , causing a decline in earthworm populations. Lavelle et a l . (1983) found that s o i l with added hydrocarbons, extracted from leaf l i t t e r , made good worm food. The amount of extract added to the s o i l negatively correlated with the egestion rate of Millsonia anomala, indicating a possible d i r e c t n u t r i t i o n a l value of the hydrocarbons. The worms showed a sat i s f a c t o r y growth response to a l l l e v e l s of hydrocarbons provided. When s o i l s contained low amounts of hydrocarbons, a high l e v e l of microbial growth was observed in the gut content, while with high l e v e l s , microorganism growth was decreased. Proteins, o i l s and carbohydrates caused weight loss in E. foetida when presented as worm food. This may suggest that simple hydrocarbons are not suitable as worm food, while easily d i g e s t i b l e materials such as casein are toxic to earthworms due to the putrefying e f f e c t s in the gut (Neuhauser et a l . , 1980 b). Shaw and Pawluk ( 1 986) found that L. t err estris and the endogeic worms 0. tyraeum and A turgida equally increased the number of actinomycetes and bacteria in the faeces. In L. terrestris faeces, fungi p r o l i f e r a t e d , while the endogeic species enriched the s o i l with c e l l u l o s e decomposing bacteria such as Cytophaga. The increase in LITERATURE REVIEW / 13 microbe biomass in the faeces, as described by Parle (1963), may either be caused by the development of cysts and spores, ingested with the organic matter, and stimulated by a favourable environment in the gut, or by the growth and establishment in the gut of a population of microorganisms in a mutualistic r e l a t i o n s h i p with the earthworm as suggested by Shaw and Pawluk (1986). The evidence that earthworms u t i l i z e either dead or l i v i n g microorganisms as food or u t i l i z e hydrocarbons extracted from decaying leaf l i t t e r , may suggest that microorganisms, or their products, are an important food source for earthworms. Worms may ingest s o i l and organic matter inhabited by large populations of microorganisms, crush the soft-bodied organisms in the gizzard and then extract nutrients in the gut. The remaining materials, shredded l i t t e r , c e l l wall fragments etc., are a favourable environment for rapid increases in microorganism biomass in earthworm faeces. LITERATURE REVIEW / 14 3. Impact of earthworms on the s o i l system a. E f f e c t s of earthworms on s o i l structure and f e r t i l i t y Earthworms have been found to have a major impact on the s o i l system. Lee (1985), Springett and Syers (1984), Hayes (1983), Edwards (1981), Kirkham (1981), Edwards and Lofty (1977) and Satchell (1967) summarized the ef f e c t s earthworms have on s o i l structure. S o i l with worm casts contained more water-stable aggregates than non-cast s o i l s and had higher porosity. Crops grown on s o i l in which earthworms were active, showed higher y i e l d s through better root penetration and nutrient a v a i l a b i l i t y . Decomposition of organic matter may be enhanced through mixing and shredding by earthworms and the p r o l i f e r a t i o n of fungi and bacteria (Shaw and Pawluk, 1986) b. E f f e c t s of earthworms on i n f i l t r a b i l i t y The i n f i l t r a b i 1 i t y of water into the s o i l has been related to earthworm a c t i v i t y . Carter et a l . (1982) reported that i n f i l t r a t i o n rates in drained and undrained f i e l d s in the Fraser Valley, B r i t i s h Columbia, p o s i t i v e l y correlated with earthworm abundance. Ehlers (1975) reported on the i n f i l t r a t i o n of water in t i l l e d and u n t i l l e d p l o t s . He concluded that earthworm channels contribute to water drainage only when they reach the surface and that water LITERATURE REVIEW / 15 i n f i l t r a t i o n w i l l take place only at high rain i n t e n s i t i e s . Baker (1981) concluded from his l i t e r a t u r e review that earthworms increased the i n f i l t r a t i o n rate of water into the s o i l . His research also showed that when earthworms were removed from a turf grass putt and pitch, through application of pesticides, the burrows rapidly closed and the i n f i l t r a t i o n rate decreased. c. The mixing of organic matter into the p r o f i l e Mixing of organic matter into the s o i l p r o f i l e by earthworms, and the changes in the p r o f i l e were studied to describe the effects worms have on the s o i l p r o f i l e . Dietz and Bottner (1981) placed 1 UC labe l l e d l i t t e r on the surface of a grassland s o i l and used autoradiography to follow the movement of the 1"C into the p r o f i l e . Most of the decomposition products were transported by drainage, while a small portion was mixed into the s o i l by earthworms. Stout (1983) and Stout and Goh (1980) showed that "bomb carbon" ( 1 WC enrichment of the biosphere from nuclear bombs) was not mixed in the p r o f i l e by surface dwelling worms in a forested system in England, but was mixed into the p r o f i l e by sub-surface dwelling worms. Sub-surface or endogeic species do not mix surface l i t t e r into the p r o f i l e (Shaw and Pawluk, 1986), however, endogeics may mix "bomb carbon" into the p r o f i l e because 1"C0 2 i s taken up by shoots and the LITERATURE REVIEW / 16 radioactive material i s translocated to the roots. Partly decomposed roots are ingested by endogeics and the "bomb carbon" is cycled by these worms. The "bomb carbon" was also mixed into the p r o f i l e by introduced earthworms in a grassland in New Zealand. Radiolabelled Cs was followed in a f i e l d t r i a l in Oakridge, Tennessee by Crossley et a l . (1971). Earthworms (Oclolasion I act eum) mixed the label into the mineral s o i l . L. terrestris, introduced in coal s p o i l s , removed large quantities of l i t t e r from the surface (Vimmerstedt and Finney, 1970). Earthworms that invaded a New Brunswick mixed forest, Al I ol obophor a tuberculata, Dendrobaena oclaedra, L. festivus and L. terrestris, dramatically changed the s o i l p r o f i l e from a t y p i c a l Podzol to a p r o f i l e with an apparent Ah horizon. This transformation took place in approximately 4 years (Langmaid, 1964). Improved s o i l structure and a decrease in thatch were observed after earthworms, A. caliginosa and L. terrestris, were introduced in newly reclaimed polders in The Netherlands (Hoogerkamp et a l . , 1983). When earthworms were removed from an orchard s o i l in The Netherlands through over-use of copper-containing pesticides, the s o i l structure rapidly deteriorated and a thatch layer developed on top of the mineral s o i l (Van Rhee, 1963). LITERATURE REVIEW / 17 Thus, some earthworm species c l e a r l y play a role in the draining c h a r a c t e r i s t i c s of s o i l s and in the mixing of organic matter into the s o i l p r o f i l e . Worms can d r a s t i c a l l y change the p r o f i l e ; t h i s change may not be permanent once earthworms are removed from the s o i l system. d. Surface cast production Researchers f i r s t studied worm-cast production by c o l l e c t i n g only a few samples per f i e l d . These samples were scraped from the s o i l surface and would not represent the cast production in a larger f i e l d (Evans and Guild, 1947). Evans and Guild (1947) c o l l e c t e d casts on a much larger scale. They sampled from 1 m diameter plots and estimated an annual production of 31.4 t.ha" 1. Only two species, AlI ol obophora I onga and A. nocturna, both surface casting worms, were noted to contribute to the surface cast (Evans, 1948). Recently, wormcasts were co l l e c t e d from several plots per f i e l d to quantify the cast production of t r o p i c a l species. Watanabe and Ruaysoongnern (1984) c o l l e c t e d surface casts of the genus Pheretima in Thailand. Casting a c t i v i t y took place in the rainy season, June to November, and the production ranged between 132.6 and 224.9 t.ha~ 1.y~ 1. The cast production of Pheretima alexandri in India was described by Reddy (1982). This species produced between 23.4 and 140.9 LITERATURE REVIEW / 18 t . h a _ 1 . y _ 1 in a humid mixed woodland. Data on surface-cast production of dif f e r e n t earthworm communities were compiled by Lee (1985), Watanabe and Ruaysoongnern (1984), Reddy (1983), Edwards and Lofty (1977), Evans (1948) and Evans and Guild (1947). Annual surface cast production varied between 4.5 and 90.2 t.ha" 1 in temperate regions with lumbricids, and between 50.4 and 2600 t.ha" 1 in t r o p i c a l regions. No data were found in the l i t e r a t u r e specifying t o t a l cast production (surface plus sub-surface cast) in the f i e l d . e. Parameters aff e c t i n g the cast production of earthworms Parameters aff e c t i n g cast production of earthworms were studied for several species. Lavelle (1975) related food consumption and growth of Millsonia anomala, a t r o p i c a l earthworm, to s o i l water p o t e n t i a l . Maximum worm a c t i v i t y was occurred at pF 2.0 - 2.5 (1.0 - 3.1 m of water); worms became inactive when the pF rose to between 3 and 4.2 (10 and 160 m of water). Bouche (1983), s p e c i f i e d the pF ranges for earthworm species: the pF l i m i t s for anecics were 2.06 - 3.08 (1.1 - 12 m of water), for epigeics, pF 2.31 - 3.33 (2 - 21 m of water), while for endogeics no l i m i t s were established. The cast production was influenced by s o i l temperature: temperatures up to 30 °C increased cast production of M. LITERATURE REVIEW / 19 anomal a (Lavelle, 1975), and temperatures in the 5 - 1 0 °C range increased feeding and burrowing a c t i v i t y of AlI ol obophora rosea (Bolton and P h i l l i p s o n , 1976). Immature worms of the species M. anomala showed a r e l a t i v e intake of s o i l 3 times higher than that of adults (30 °C) (Lavelle, 1975). This phemomenon was also reported by Bolton and P h i l l i p s o n (1976) for A. rosea, although the difference between very young worms and adults was much smaller. Differences in energy budget (food intake) between small immature, large immature and adult worms, i l l u s t r a t e the changing metabolic demands of individuals through t h e i r l i f e cycle (Lee, 1985). Food qua l i t y influenced the t o t a l cast production as was described by Martin (1982). He mixed grass meal with s o i l and found that the egestion rate of L. r u b e l l u s decreased when more grass meal was added to the mixture. Lavelle et a l . (1983) added hydrocarbons, extracted from leaf l i t t e r , to s o i l on which M. anomala fed. U. anomal a showed a decreased cast production when increased amounts of hydrocarbons were added to the s o i l , while the worms continued to grow. Hydrocarbons may have increased the food value of the offered s o i l , because they include nutrients produced by or extracted from microorganisms in the l i t t e r LITERATURE REVIEW / 20 from which the hydrocarbons were extracted. Cast production of L. rubellus and A. caliginosa depended on the calcium concentration of the s o i l . Calcium added to a f i e l d in New Zealand, increased the surface cast production of those species (Springett and Syers, 1984). In summary, t o t a l cast production of earthworms may be p o s i t i v e l y affected by s o i l moisture, s o i l temperature and calcium concentration, while increased food quality decreased the amount of casts produced. The age of the worm i s a s i g n i f i c a n t factor in the cast production: young immature earthworms have a higher egestion rate than c l i t e l l a t e d adults. LITERATURE REVIEW / 21 B . LUMBRICUS RUBELLUS HOFFMEISTER, 1843. 1 . D i s t r i b u t i o n The earthworm Lumbricus rubellus Hoffmeister, 1843, (Annelida, Lumbricidae), inhabits humus r i c h moist s o i l s such as in pastures and riverbanks, under stones, boards or decaying leaves (Reynolds et a l . , 1974). drained a g r i c u l t u r a l f i e l d s (Carter et a l . , 1982; Carter, unpublished). It i s perhaps the most widespread species in the world. It has been reported in Europe, North America, Aus t r a l i a and New Zealand (Robotti, 1984). Reynolds et a l . (1974) described i t as one of the most widely d i s t r i b u t e d and abundant species in Eastern North America: the worm i s recorded in 25 states of the US and 6 Canadian provinces. It i s a common species in the Vancouver area in drained a g r i c u l t u r a l f i e l d s (Carter et a l . , 1982; Carter, unpublished). However, no records were found on L. rubellus in the p r a i r i e states and provinces. The dispersal of t h i s species, and of earthworms in general, was described by Lee (1985), Bouche (1983), Schwert (1980) and Gates (1976). Passive migration of earthworms may take place by water, land s l i d e s and animals including humans. Most North American species were introduced from Europe by immigrants bringing farm animals and plant stock. Further LITERATURE REVIEW / 22 d i s t r i b u t i o n of worms in Canada took place through plant stock and livestock shipments, spreading of manure and by fishermen discarding surplus b a i t . L. rubellus is usually found in the upper 8 cm of the s o i l (Edwards and Lofty, 1977) and according to Byzova (1965), i t moves down during dry surface conditions but i s found in the l i t t e r layer under very wet conditions. Persson and Lohm (1977) reported that L. rubellus was found in the 15-30 cm depth of a grassland s o i l in Sweden. The species i s capable of surviving a wide range of conditions (Curry and Cotton, 1983; Eijsackers, 1983). Conditions under which L. rubellus may be found, are l i s t e d in Table 1. Reynolds (1976) described the species as a pioneer species, and Lee (1985) noted that L. rubellus appeared early in the development of a pasture system, but later was found as a minor component. Bengston et a l . (1979) suggested that the survival of A. caliginosa was superior to that of L. rubellus in introductions using l i t t e r b a g s in an Iceland hayfi e l d . In Europe, L. rubellus is not a dominant species (Edwards and Lofty, 1977; Van Rhee, 1963); however, Eijsackers (1983) reported i t to be present in great abundance in abandoned LITERATURE REVIEW / 23 Table 1. The conditions r e s t r i c t i n g the d i s t r i b u t i o n of L. rubellus Parameter Value Reference pH 2.5-9.0 temp, optimum 15 - 18° C Lofs-Holmin (1983) Edwards and Lofty (1977) s o i l moisture moist s o i l s Reynolds (1974) texture clay - gravely sand Edwards and Lofty (1977) l i g h t loam Ma (1983) a l t i t u d e lowland to alpine Zajonc (1982) f i e l d s in The Netherlands. L. rubellus is commonly found in association with AlI ol obophora rosea, A. longa, A. caliginosa, A. chlorotica, Lumbricus castaneus, and L. terrestris (Zajonc, 1982; Edwards and Lofty, 1977). In Coastal B r i t i s h Columbia, L. rubellus i s a dominant species in drained s i l t y clay loam s o i l s , with maximum densities ranging from 13.1 to 25.0 g.m"2 (dwt) (Carter, unpublished; Carter and Bandoni, unpublished). LITERATURE REVIEW / 24 2 . Reproduction L. rubellus produces large numbers of cocoons (79 - 100 per worm per year). The cocoons incubate 10 weeks and the worms grow to maturity in 40 weeks (Evans and Guild, 1948; Edwards and Lofty, 1977). Each cocoon hatches one, sometimes two, hatchlings (Evans and Guild, 1948). Cocoon production depends on s o i l temperature, s o i l moisture, food a v a i l a b i l i t y and quality (Evans and Guild, 1948), and s o i l texture (Ma, 1983). Ma (1983) reported that the worm showed a higher a c t i v i t y and cocoon production in a sandy s o i l , compared to a s i l t loam. 3. Food sources and e a t i n g h a b i t s Food sources of L. rubellus include manure, l i t t e r or organic matter (Edwards and Lofty, 1977), and r e l a t i v e l y undecomposed plant remains (Piearce, 1972). Piearce (1978) found that the gut of L. rubellus contained an abundance of organic matter, (mostly f i b r e s and grass leaves) and a minor al g a l component. Selective feeding by L. rubellus on algae was also observed by Nekrasova et a l . (1976). Baylis et a l . (1986) reported that L. rubellus contained 3 2P, added to the leaves of clover plants and transported to the roots. LITERATURE REVIEW / 25 4 . Respiration Byzova (1965) reported an oxygen consumption rate for L. rubellus of 89 mm 3.g _ 1.h _ 1. The worm had a rate that was similar to that of midstrata and deep s o i l dwellers (Byzova, 1965). The sp e c i f i c haemoglobin content of the blood was 14.4 mg.g"1 (dwt), the lowest of the four species investigated (Byzova, 1973). 5. Egestion and s o i l turn-over Not much information i s available regarding ingestion and turn-over of s o i l and organic matter by L. rubellus. Martin (1982) found a maximum egestion rate of 3.01 g. g - 1 . d a y 1 ( l i v e weight) for s o i l mixed with 4.4 g.kg" 1 grass meal. The figure for s o i l only was 1.92 g . g - 1 . d a y 1 (wet weight). A removal of 0.15 g . g - 1 . d a y 1 (dwt) from the surface was reported for hazel leaves and between 0.32 and 0.40 g . g - 1 . d a y 1 (dwt) for manure (Edwards and Lofty, 1977). Carter et a l . (1983) reported an egestion rate for s o i l of 0.90 g.g" 1.day 1, using a chromium oxide ( C r 2 0 3 ) technique. Sharpley and Syers (1977), and Syers et a l . (1979) reported maximum surface cast production by L. rubellus in grassland s o i l s in the spring and f a l l , which they could relate to s o i l moisture and temperature. Heungens (1969) reported a decrease in l i t t e r depth through the shredding of pine l i t t e r by a community of Lumbricids, mainly Dendrobaena spp. LITERATURE REVIEW / 26 but including L. rubellus, L. castaneus, Eisenia eiseni and A. cI or ol i ca . When lime was applied to a f i e l d , L. rubellus produced s i g n i f i c a n t l y more surface cast than without lime. In the laboratory, the worm tended to mix lime and phosphate rock ho r i z o n t a l l y in the p r o f i l e , while A. caliginosa mixed v e r t i c a l l y (Springett, 1983). 6. E c o l o g i c a l c l a s s i f i c a t i o n and s t r a t e g i e s Bouche (1977) c l a s s i f i e d L. rubellus as an epigeic species (small worms, high reproduction, l i v i n g in the l i t t e r l a y e r ) , although the worm has also c h a r a c t e r i s t i c s of the anecics (burrowing). In Ju l i n ' s system (Reynolds, 1976), the worm was described as a hemerobiont, clo s e l y t i e d to human populations. Satchell (1980) described the survival strategy of L. rubellus as that of an r- s t r a t e g i s t (high reproduction, fast recovery), although the worm was reported to have some c h a r a c t e r i s t i c s of the K-strategist, for example i t s moving into the s o i l p r o f i l e under adverse conditions. The species could adapt i t s e l f to many conditions and has been regarded as a pioneer species. The worm ingests s o i l and r e l a t i v e l y undecomposed plant material and may select LITERATURE REVIEW / 27 microorganisms as food. Limited information i s available on s o i l turn-over, and no information was found on the d i s t r i b u t i o n of casts in the s o i l p r o f i l e . The fact that the species did not exactly f i t in either the su r v i v a l or the strategy c l a s s i f i c a t i o n might explain i t s ada p t a b i l i t y and widespread d i s t r i b u t i o n . III. THE EFFECTS OF SOIL MOISTURE AND SOIL TEMPERATURE ON THE EGESTION AND INGESTION RATES OF LUMBRICUS RUBELLUS HOFFMEISTER A. MATERIALS AND METHODS 1. Introduction The r e d i s t r i b u t i o n of organic matter in the s o i l by L. rubellus may be influenced by the s o i l temperature and s o i l moisture content. The effects of temperature and moisture on the cast production of the earthworm L. rubellus was quantified, using a l i t t e r b a g technique. In t h i s study i t was assumed that the temperature and moisture regimen of the s o i l do not affect the earthworm a c t i v i t y , measured as the amount of s o i l and organic matter ingested or egested. The ef f e c t s of drought stress on the earthworm were discussed using the results of the l i t t e r b a g experiment. 2. Site description The study s i t e was located on Westham Island (122°.30' W, 49°.36' N) , 55 kilometres south of Vancouver, B.C., situated in the delta of the Fraser River. The s o i l s were of f l u v i a l o r i g i n and were c l a s s i f i e d by Luttmerding (1981) as s i l t y clay loam-textured Orthic Gleysol (typic haplaquept in the US c l a s s i f i c a t i o n ) of the Crescent Series. To improve 28 MOISTURE AND TEMPERATURE / 29 a g r i c u l t u r a l production, the o r i g i n a l , poorly-drained s o i l had been drained with perforated p l a s t i c drains, spaced 20 m apart and situated at a depth of 1.2 m. The drains emptied into a ditch that was pumped year-round. During 1983, f i e l d data were c o l l e c t e d from the experimental pl o t . The plot (35 by 35 m) was in the clover phase of a barley-clover-potato rotation. During the second year of th i s study (1984) the part of the f i e l d that was used for the experiment was kept under clover, while the remainder of the f i e l d was under potatoes. 3. F i e l d data c o l l e c t i o n During the two years in which the experiments were conducted, s o i l temperature and gravimetric s o i l moisture content were measured. The s o i l temperature at 5, 10 and 15 cm depth was continuously recorded using a thermograph, and twice monthly, six s o i l samples were taken in a pre-designated sub-plot to determine the s o i l water content. These samples were taken from the 0-5 cm layer and the 5-10 cm layer. MOISTURE AND TEMPERATURE / 30 4 . Earthworms used in the experiments The earthworms used in the experiments, were Lumbricus rubellus Hoffmeister, 1843 (Annelida, Lumbricidae), coll e c t e d from the clover f i e l d on Westham Island by handsorting. Only c l i t e l l a t e d adults were c o l l e c t e d prior to each experimental run. 5. Sample c o l l e c t i o n S o i l was co l l e c t e d from the plough layer, a f t e r the top 3 cm was removed and discarded because t h i s layer contained large amounts of earthworm casts and had a root mat. Clover hay was c o l l e c t e d in the spring after i t had been in the f i e l d during the winter months. S o i l and clover hay were stored in the freezer and were l e f t to thaw several days prior to each t r i a l . The organic material was washed in tap water to remove old faecal material and s o i l , and cut into 1 cm pieces. The s o i l was broken into aggregates of about 1 cm in diameter. S o i l c o l l e c t e d on Westham Island was also used to create a stable environment with a known moisture content. This s o i l was broken into aggregates < 0.5 cm and brought to the desired water content by watering with a plant sprayer and mixing by hand. MOISTURE AND TEMPERATURE / 31 6. L i t t e r b a g t e c h n i q u e Ingestion rates and egestion rates of L. rubellus were estimated using a l i t t e r b a g technique (Carter, personal communication). Often, l i t t e r b a g s are used for decomposition studies. The exclusion of s o i l organisms and the easy recovery of plant materials is the main purpose for the use of the bags (McBrayer and Cromack, 1980; Uvarov, 1982). The mesh size i s related to the size of organism to be excluded. Bags of various size mesh have also been used to contain s o i l and organisms in experiments to measure growth and reproduction (Satchell, 1971) and survival (Bengston et a l . , 1979) of earthworms and of ingestion and egestion studies of millipedes (Carter, pers. communication). In using l i t t e r b a g s to study the food uptake and egestion rates of contained organisms, the mesh size should be s u f f i c i e n t l y small to contain the animals and their casts, but not too small to impede gas and water exchange. Uvarov (1982) reported d i f f i c u l t i e s with water exchange in li t t e r b a g s placed in the f i e l d , especially for those with a mesh size of < 0.01 mm. Litterbags were made from fine mesh polyester material (no-see-um netting, 0.3 mm mesh s i z e ) , measured 10 by 10 cm, and were assembled by stapling a s t r i p of p l a s t i c on the MOISTURE AND TEMPERATURE / 32 folded hems. The bags were closed by stapling the twice-folded hems. The 0.3-mm mesh size was assumed to be large enough not to impede gas and water exchange. P a r t i a l l y dried material was used for the lower water contents and water was added to the content of the bag for the higher moisture lev e l s , to prevent possible effects of a small mesh size. To indicate the moisture status of the s o i l , the s o i l water tension (metres of water) or the s o i l water potential (metres of water or Joules.kg" 1) was used. These units indicate the energy state of the water in the s o i l ( H i l l e l , 1980). S o i l s with similar potentials could then be compared as to the effects on earthworms or plants. The relat i o n s h i p between s o i l moisture and the s o i l water tension i s described as the retention curve ( H i l l e l , 1980). When the s o i l moisture content i s used, a complete description of the s o i l , including a retention curve, should be provided to relate the moisture content to s o i l water potentials. The retention curve for the Crescent s i l t y clay loam is included in Appendix 2-E. S o i l water potential has been used for indicating the moisture status of the s o i l in several earthworm studies. Evans and Guild (1948) related the cocoon production of MOISTURE AND TEMPERATURE / 33 earthworms to pF values, the log transformation of the s o i l water tension in centimetres of water. Lavelle (1975) related the ingestion of s o i l by t r o p i c a l earthworms to the pF values and Reynolds and Jordan (1975) advocated the use of s o i l water tensions in habitat descriptions. Bouche (pers. communication) advocated the use of s o i l water potential ("...II est ess e n t i e l que vos resultats soient exprimes en pF ... car le % H 20 ne veut rien dire ! . . . " ) . Earthworms were starved for 48 hours at incubation temperatures (5, 10, 15 and 20 °C), then put into the li t t e r b a g s containing a pre-weighed amount of food, consisting of a mixture of s o i l and clover hay. The bags were buried in large p l a s t i c washbasins, f i l l e d with moistened s o i l . The basins were covered in p l a s t i c to prevent evaporation and were kept in the incubator for seven days. The loose packing of s o i l aggregates in the basins and the large volume of a i r between the p l a s t i c and the s o i l , prevented anaerobic conditions. In preliminary tests i t was found that the s o i l showed v i r t u a l l y no difference in water content before and after a feeding t r i a l ; therefore the moisture content of the f i l l e r s o i l was measured at the end of the experiments. The following moisture levels were used in the experiments : 0.33, 0.30, 0.27 and 0.24 kg.kg - 1 (-5, -9, -17 and -25 m of water respectively). Higher s o i l MOISTURE AND TEMPERATURE / 34 moisture contents were not used because of smearing of s o i l and faecal material by the worms. Thirty worms, each in a separate l i t t e r b a g , were incubated in each moisture-temperature combination. No s i g n i f i c a n t difference in egestion rate (g.g" 1.day" 1) between 7, 10 and 14 days incubations was found in preliminary tests. Therefore subsequent experiments were run for 7 days (See Appendix 3-A). Fresh faeces may contain a high l e v e l of ammonia, which might be toxic to earthworms as described by Neuhauser et a l . (1980 a) for Eisenia foetida in experiments l a s t i n g for one month. Based on the short duration of the present experiments i t can be assumed that no cast material was consumed and no toxic build-up of ammonia took place. After incubation, the worms were removed from the l i t t e r bags. With a glass rod the anterior end of the worm was massaged u n t i l a small amount of cast was produced (Bolton and P h i l l i p s o n , 1976). The cast material was then immediately transferred onto a pre-weighed aluminum dish and weighed on a micro balance. After being oven-dried (4 days, 65 ° C ) , the cast was weighed again and the water content was calcu l a t e d . The worms were then starved for 48 hours to remove their gut content, and freeze-dried to determine MOISTURE AND TEMPERATURE / 35 t h e i r dry weights. F a e c a l p e l l e t s were s o r t e d out and the components of the l e f t o v e r food were separated, oven-dried (4 days, 65 °C) and weighed. 7. Water content of the worm S o i l of the Crescent s e r i e s was p a r t i a l l y d r i e d and crushed s u f f i c i e n t l y to pass a 2 mm s e i v e and was then e q u i l i b r a t e d i n a porous p l a t e e x t r a c t o r to reach predetermined s o i l water p o t e n t i a l s between -3 and -60 m of water. A f t e r e q u i l i b r a t i o n , the s o i l was t r a n s f e r r e d i n t o p l a s t i c c o n t a i n e r s and 5 earthworms were added to each c o n t a i n e r . Sub-samples were taken, d r i e d and weighed to c o n s t r u c t a r e t e n t i o n curve. The experiments were done i n d u p l i c a t e f o r both a d u l t and immature a d u l t earthworms. The l a t t e r were w e l l developed n o n - c l i t e l l a t e d specimens. A f t e r i n c u b a t i n g seven days at 19 CC i n the c l o s e d c o n t a i n e r s , the worms were weighed and then p l a c e d in P e t r i d i s h e s on moist f i l t e r paper t o have t h e i r gut emptied. The faeces were c o l l e c t e d , a i r - d r i e d and weighed. The worms were immediately k i l l e d by f r e e z i n g , dehydrated i n the f r e e z e r and weighed. The water content of the"worms was c a l c u l a t e d assuming that the gut content c o n t a i n e d 53 % water (dwt), the l i q u i d l i m i t of the Crescent s o i l (De V r i e s , p e r s o n a l communication). MOISTURE AND TEMPERATURE / 36 8. Chemical a n a l y s i s Total carbon of s o i l and cast materials was determined by dry oxidation, using a Leco carbon analyser. 9. C a l c u l a t i o n s and S t a t i s t i c s The experiment was designed as a 4X4 f a c t o r i a l , with equal spacing beween temperature and between moisture l e v e l s . Normality was tested using skewness and kurtosis of the standardized population, the homogenity of variances was tested and appropriate logarithmic transformations were applied. Functions were f i t t e d for l i n e a r i t y , quadratic or cubic relations and the p r o b a b i l i t i e s of the contrasts were calculated. The equality of the means was tested with a series of student t-tests (egestion experiment) or a series of Mann-Whitney U-tests (water content of the worms). B. RESULTS AND DISCUSSION 1. L i t t e r b a g technique Faecal material was e a s i l y separated from the left-over food because of the d i s t i n c t shape and structure of the p e l l e t s . Their rounded shapes were d i f f e r e n t from the angular s o i l aggregates and their colour was darker and greener than the bulk s o i l . MOISTURE AND TEMPERATURE / 37 2 . Calculations and s t a t i s t i c s a. The grouping of the moisture and temperature levels The experiments involving egestion rates, were conducted at several moisture l e v e l s , but incubated at constant temperatures. The drawback of t h i s approach was that the moisture regimens were not exactly the same in each temperature t r i a l . However, when grouped, the midpoints of the ranges (moisture content, kg.kg" 1) were equally spaced and therefore a f a c t o r i a l approach in the s t a t i s t i c a l analysis was used. b. Calculation of the ingestion rates The c a l c u l a t i o n of the ingestion rate was d i f f i c u l t , because individual input and output values had to be used while the correction for the water content was based on bulk measurements. The food material was not dried before i t was put in the l i t t e r b a g , because a change in physical, chemical and especially microbiological c h a r a c t e r i s t i c s , would have resulted from drying. c. S t a t i s t i c a l analysis Equal numbers of data points per stratum are required for ANOVA tes t s . As the number of data points varied between 24 and 30, due to worm mortality and escapes, the number of MOISTURE AND TEMPERATURE / 38 data points was reduced to 24 for each stratum. There are several methods to reduce the number of data points. The method used was that of 'reasoning - out' cases. Outliers were examined and i f there was reason to believe that the value could be explained as not a normal one, these points were deleted from the data set. Care was taken that both high and low o u t l i e r s received a similar treatment. Preliminary tests showed that about 50 % of the v a r i a b i l i t y was caused by only 10 % of the cases in a c e r t a i n set. The equalized data sets were normally d i s t r i b u t e d . They were tested for normality by describing the skewness and kurtosis of the standardized d i s t r i b u t i o n s . Also a L i l l i e f o r s test did not reject the hypothesis that the egestion rate was normally d i s t r i b u t e d (p > 0.10). To homogenize the variances, the data was exponentially transformed. Powers of transformation were calculated assuming V(ju) = hu . The calculated powers were: 0.07489 for the egestion rate (E), 0.0959 for the ingestion rate of organic matter (IOM), 0.0951 for the ingestion rate of s o i l (ISOL) and a ln transformation for the t o t a l ingestion rate (ITOT = IOM + ISOL). After transforming the data, the variances were s t i l l not s i g n i f i c a n t l y homogenous, but the large number of r e p l i c a t i o n s in each stratum (24) made the use of parametric s t a t i s t i c s possible (Siegel, 1980). As a precaution, the separation of means was tested with an F-test and a MOISTURE AND TEMPERATURE / 39 (non-parametric) Kruskal-Wallis K-sample test. The two tests gave similar s i g n i f i c a n t results (p < 0.01). d. Curve f i t t i n g The factors Temperature and Moisture as well as the T*M interaction were s i g n i f i c a n t for a l l variables (Table 2). Matching curves through testing orthogonal polynomials (curve f i t t i n g ) yielded many s i g n i f i c a n t interactions and i t was not possible to draw any conclusions regarding type of curve, i n f l e x t i o n point or threshold values. Therefore, the equality of means was tested with a series of t - t e s t s . The data points in Figure 2 were connected by hand. 3. Egestion rates of s o i l and organic matter Both temperature and moisture c l e a r l y influenced the egestion rate of L. rubellus. In wet s o i l a s i g n i f i c a n t difference was found between the egestion rates at various temperatures (Table 3). When the s o i l wetness decreased from 0.33 to 0.30 kg.kg 'M -5 to -10 m of water), the E did not change s i g n i f i c a n t l y . At a lower moisture l e v e l , (0.27 kg.kg" 1, - 17 m of water), the E increased s i g n i f i c a n t l y for a l l temperature t r i a l s except the highest temperature (20 °C), where i t decreased s i g n i f i c a n t l y . In this case, the egestion rate for dry s o i l (0.24 kg.kg" 1, -25 m of water) was s i g n i f i c a n t l y lower than for medium dry s o i l s . The MOISTURE AND TEMPERATURE / 40 Figure 2. The egestion rate (g.g" 1.day" 1) of L. rubellus related to the s o i l temperature and the s o i l moisture. Each point represents 24 worms. MOISTURE AND TEMPERATURE / 41 T a b l e 2 . F - v a l u e s f o r i n t e r a c t i o n s and c o n t r a s t s d e s c r i b i n g E g e s t i o n and I n g e s t i o n r a t e s o f L. r u b e l l u s , i n c u b a t e d a t d i f f e r e n t m o i s t u r e c o n t e n t s and s e t t e m p e r a t u r e s . (** p < 0 . 0 1 , * p < 0 . 0 5 , n = 24) E g e s t i o n I n g e s t i o n (OM) I n g e s t i o n (SOIL) TEMP 63 . 7** 5 . 76** 5 3 . 6 * * MOIST 1 1 3 . 8 * * 8 .6 6** 9 8 . 7 * * T*M 3 9 . 0 * * 3 . 9 1 * * 34 . 5** T L i n 1 3 6 . 2 * * 6 . 8 4 * * 9 9 . 3 * * TQuad 514 . 9 * * 6 . 7 1 * * 6 1 . 3 * * TDev 0 . 0 5 3 . 3 7 0 .052 M L i n 1 6 9 . 4 * * 0 . 2 1 1 6 3 . 4 4 * MQuad 1 2 7 . 2 * * 1 2 . 4 * * 9 6 . 9 * * MDe v 4 4 . 9 * * 1 3 . 3 * * 3 5 . 9 * * T L i n - M L i n 9 9 . 5** 0 .04 8 8 . 5 * * TLin-MQuad 16 5 . 6 * * 0 .46 1 5 1 . 3 * * T L i n * M D e v 1 8 . 5 * * 2 0 . 3** 4 . 2 3* TQuad-MLin 5 .8 8* 0. 54 4 . 7 0 * TQuad-MQuad 5 4 . 6 * * 0 . 2 8 5 2 . 3 * * TQuad*MDev 0 .26 0 . 3 7 0 .04 TDev-MLin 4 . 8 2 * 7 . 0 6 * 5. 82* TDev-MQuad 0 . 6 1 0 . 1 3 0 . 7 7 TDev-MDev 5 . 0 9 * 6 . 00* 2 .46 Table 3. Egest ion rates for L . rube H U H , incubated at d i f f e r e n t temperatures and moisture regimens. P r o b a b i l i t y of data po ints being d i f f e r e n t ( t - t e s t , * = p < 0.0S, ••••• = p < 0.01, n=2M). A. Temperature g r a d i e n t compared. M o i s t u r e c o n t e n t : 0.33 kg.kg •1 0. 30 0. 27 0. 2U Temp. Temp Temp Temp 5° C. 5° C. 5° 5° C 10 10 10 10 l b ;'; 15 1; 15 5*; & 15 20 &;» 20 20 :V 20 :'; sV Temp 5° 10 15 5 10 15 5 10 15 5 10 15 B. M o i s t u r e g r a d i e n t compared o I—I tn - 9 G > Z D -3 M 3 M > -3 C n Temperature: 5° C. 10 15 20 0.33 0.30 0.27 0. 2<4 0.33 0 . 30 0 . 27 0. 2M 0.33 0. 30 0.27 0. ?t 0.33 0. 30 0. 27 0. 21 M o i s t u r e 0.33 0.30 0.27 0.33 0.30 0.27 0.33 0. 30 0.27 0. 33 0. 30 0.27 MOISTURE AND TEMPERATURE / 43 general trend was that when the s o i l moisture content decreased, the egestion rate increased and then dramatically decreased. In wet s o i l s the temperature ef f e c t was noticable. In medium wet s o i l s ( - l O to -15 m of water), the moisture ef f e c t obscured the temperature e f f e c t , while in dry s o i l s (< -20 m of water), the moisture effect was important. The effects of the dry s o i l was more v i s i b l e at 5 °C and 20 °C, probably because these temperatures are near the l i m i t s of the temperature range of the worm. 4 . Egestion rate of carbon The egestion rate of carbon was calculated from the carbon content of the faecal material and the egestion rate. A s i g n i f i c a n t positive temperature ef f e c t (Kruskal-Wallis K-sample, p < 0.01) was found (Table 4). The earthworms processed less carbon at 5 °C than at a l l other temperatures. At 5 and 20 °C, the dryest s o i l showed a s i g n i f i c a n t l y lower egestion rate of carbon (Mann-Whitney U-test, p < 0.05, p < 0.01), while at 15 °C, the egestion rate of carbon in wet s o i l s was s i g n i f i c a n t l y lower than that of the 0.27 kg.kg" 1 moisture l e v e l (p < 0.01). No s i g n i f i c a n t differences were found between the treatments at 10 °C. A threshold l i m i t may affe c t the a c t i v i t y of the earthworm. The egestion of carbon seemed to reach a maximum at approximately 0.27 kg.kg"1 moisture ( - 17 m of water) MOISTURE AND TEMPERATURE / 44 Table 4. The comparison of the t o t a l egestion rate, E (total) (g.g~ 1.day~ 1, SD) with the egestion rate for carbon, E (carbon) (mg.~1.day~1, SD) for four temperatures, T ( °C) and four moisture contents in each set of temperatures (0.33, 0.30, 0.27 and 0.24 kg.kg" 1 respectively) T E (total) (SD, n=24) E (carbon) (SD) n 5 0.31 (0.061) 17.8 ( 6.40) 10 0.31 (0.078) 18.1 ( 5.28) 12 0.41 (0.195) 25.9 ( 5.95) 9 0.15 (0.063) 12.7 ( 4.51) 8 10 0.46 (0.117) 37. 1 (12.31) 14 0.44 (0.080) 35.6 (12.24) 12 0.53 (0.179) 39.8 ( 7.05) 8 0.40 (0.106) 40.8 ( 7.66) 1 1 1 5 0.51 (0.089) 34.3 (12.51) 10 0.48 (0.124) 34.0 ( 7.92) 14 0.73 (0.296) 51.1 (16.03) 14 0.49 (0.133) 40.8 ( 8.71) 9 20 0.91 (0.253) 47.7 (12.98) 1 1 0.97 (0.416) 50. 1 ( 8.30) 1 1 0.67 (0.388) 47.9 (10.85) 9 0.31 (0.769) 19.4 ( 8.76) 1 3 MOISTURE AND TEMPERATURE / 45 before i t decreased rapidly as the moisture content decreased further. M i t c h e l l (1983) reported on a similar threshold for the egestion rate of E. foetida, feeding on sewage sludge. The egestion rate of carbon is a more "stable" parameter than the t o t a l egestion rate, even when a lower number of data points per stratum due to incomplete data sets, were considered (n=24 vs n=9-14). 5. Q 1 0 v a l u e s of the a c t i v i t y of earthworms The egestion rate of earthworms may be seen as an index of earthworm a c t i v i t y because i t represents t h e i r n u t r i t i o n a l needs, and i n d i r e c t l y r e f l e c t s the res p i r a t i o n . P h i l l i p s o n and Bolton (1976) discussed the Q 1 0 values (the change in respiration rate over a 10 °C interval) for AlI ol obophora rosea and concluded that i t was essential to know the temperatures covering the i n t e r v a l , before any statements could be made. Howard (1971) showed a decrease in the o r e t i c a l Q 1 0 values with higher temperature intervals in case of a linear response to increasing temperatures. The Q 1 0 for L. rubellus, based on the egestion rates, varied with the temperature range and with the moisture content of the s o i l (Table 5). Because the egestion rate of worms MOISTURE AND TEMPERATURE / 46 Table 5. The Q 1 0 of L. rubellus, calculated from the egestion rate, for d i f f e r e n t moisture and temperature ranges. Worms were acclimatized at the incubation temperature. The incubation lasted for 7 days. Q 1 0 for the t o t a l egestion rate Temp. Moisture (kg.kg~ 1) 0.33 0.30 0.27 0.24 1.66 1.55 1.78 3.27 1.98 2.20 1.25 0.77 Q 1 0 for the egestion rate of carbon 5 - 1 5 ° 1.92 1.88 1.97 3.21 10 - 20° 1.28 1.40 1.20 0.47 incubated at 10 and 15 °C i s less influenced by drought stress than at 5 and 20 °C, the trend in Q 1 0 d i f f e r e d for each temperature range. In wet s o i l s , an increase in Q 1 0 was observed with increasing temperatures, contrary to what Howard (1971) expected. Large changes in Q 1 0 became apparent, especially in dry s o i l . 5 - 15° 10 - 20° MOISTURE AND TEMPERATURE / 47 The Q 1 0 for the egestion rate of carbon showed a similar trend with moisture content, while the temperature effect (5-15 vs 10-20 °C) was reversed. The Q 1 0 followed the t h e o r e t i c a l decrease with a higher temperature i n t e r v a l , as described by Howard (1971). The values close to 2.0, for the lower temperature i n t e r v a l , are similar to those reported as a general value for earthworms by P h i l l i p s o n and Bolton (1976). 6. F a e c a l o r g a n i c matter The amoiint of organic matter, measured in earthworm faeces indicates the feeding pattern of worms. Faeces may show a higher organic matter content than the s o i l earthworms feed on, indicating that worms s e l e c t i v e l y ingest organic matter. Table 6 shows both the measured organic matter content of the faeces and the ingested amount, calculated from organic matter recovered from the l i t t e r b a g s and the amount o r i g i n a l l y present. Although there i s inconsistency between measured and calculated values because of the d i f f i c u l t i e s in the recovery of organic matter from the bags, an increase in organic matter content with increasing s o i l dryness i s v i s i b l e in both the measured and calculated values. L. rubellus ingested r e l a t i v e l y more organic matter when incubated in s o i l with a low s o i l water content. This trend i s not apparent at 10 and 15 °C, because these temperatures MOISTURE AND TEMPERATURE / 48 Table 6. The percentage of organic matter ingested by L. rubellus at indicated s o i l moisture lev e l s and temperatures, and calculated from IOM and ITOT and measured in the faeces. % Organic Mattert Temp °C Moisture content 0.33 0.30 0.27 0.24 Calc. Meas. Calc. Meas. Calc. Meas. Calc. Meas. 5 14.5 9.7 16.9 10.4 14.8 9.4 31.5 14.8 10 14.6 13.9 19.8 14.1 11.7 14.3 7.2 18.9 15 5.7 12.7 7.2 11.4 15.3 12.1 13.5 13.9 20 10.5 9.2 10.8 11.9 19.2 14.1 36.5 21.3 t A conversion factor of 1.724 was used to calculate % OM from % C measured in the faeces. are close to the optimum temperature for the species. The earthworms are less affected by drought stress at . these temperatures. Also the egestion rates show this phenomenon. The worms might ingest (and thus egest) more organic matter because th i s material provided moisture to the worms, for i t MOISTURE AND TEMPERATURE / 49 contained more water than mineral s o i l . The l i b e r a t i o n of water through the assimilation of organic matter i s less l i k e l y , because worms assimilate only small quantities of organic matter and they are less active in dry s o i l s . The carbon content of the faeces was above the carbon content of the s o i l , i n dicating that a mix of organic matter and s o i l was taken as food. Lee (1985) compiled data on the carbon content in earthworm casts. For L. rubellus he reported 4.3 % carbon (7.4 % OM) for casts from pastures and 1.3 % carbon (2.2 % OM) for casts from pot experiments. The values for pastures are close to those found in the present research for worms not under drought stress. 7. F a e c a l water content It was noticed that faecal material with a high water content also contained a high amount of organic f i b r e s . The faeces water content for each worm recovered from the l i t t e r b a g s , was measured at the end of the incubations. The faeces water content varied between 0.53 kg.kg" 1 and 2.4 kg.kg" 1, and was always above the l i q u i d l i m i t of the Crescent s o i l , indicating that the faeces was produced as a sl u r r y . The production of s l u r r y - l i k e faeces was observed both in the laboratory and the f i e l d . In the f i e l d , water was rapidly absorbed into the s o i l on which the faeces was MOISTURE AND TEMPERATURE / 50 deposited. The s l u r r y - l i k e faecal material could only be observed immediately after the worms produced i t . 8. Ingestion rates The rates of ingestion of mineral s o i l and organic matter, both calculated from recovered materials, showed a similar trend as the egestion rate did (Figures 3 and 4, Tables 7 and 8). In wet s o i l s a po s i t i v e temperature effect was c l e a r l y v i s i b l e , both for the ingestion of organic matter and of s o i l . The ingestion of mineral s o i l peaked at a medium moisture content, except at 20 °C, where i t peaked at a higher s o i l moisture content. The ingestion rate of organic matter showed a less r e l i a b l e trend, probably because of errors in the recovery of the materials from the l i t t e r bag. Organic matter p a r t i c l e s were sorted out by hand. Some of the material was f i n e l y shredded and some of i t was embedded in cast. 9. Worm s i z e Worms were coll e c t e d from the f i e l d shortly before each experimental run. The size of the animals, a l l c l i t e l l a t e d adults, varied over the season. Animals c o l l e c t e d in the spring (5 °C. and 10 °C) were larger than those c o l l e c t e d in the f a l l (Table 9). In preliminary experiments, immature L. rubellus showed a higher egestion rate than did MOISTURE AND TEMPERATURE / 51 I 0 M 0 " IO - 2 0 - 3 0 S O I L W A T E R P O T E N T I A L (M OF WATER F i g u r e 3. The i n g e s t i o n r a t e of o r g a n i c m a t t e r f o r L. rubellus Table 7. Ingest ion rate for Organic Matter for I., r u b e l l u s , incubated at d i f f e r e n t temperatures and moisture regimens. P r o b a b i l i t y of data po ints being d i f f e r e n t ( t - t e s t , - p <0.OS, = p <0.01, n - 2 H) . Temp 5° 10 l b 20 A. Temperature g r a d i e n t compared. M o i s t u r e c o n t e n t : 0.33 Kg.kg" 1 0. 30 Temp 5° 6. 10 lb 20 Temp b° 10 15 20 0.27 Temp 5°d. 10 15 20 0.21 Temp 10 15 10 15 B. M o i s t u r e g r a d i e n t compared. 10 15 Temperatures : M o i s t u r e l e v e l s 0.33 0.30 0.27 0.21 0.33 0. 30 0.27 0.33 0. 30 0.27 0.21 10 15 0. 33 0. 30 0.27 0. 21 10 15 0.33 0. 30 0.27 0.33 0. 20 0. 27 0.33 0. 33 0.27 0. 21 20 0.33 0. 30 0. 27 2 O »—i in •-3 G » > O PI s > -3 G W a (SI to MOISTURE AND TEMPERATURE / 53 < ce 10 2 0 5 20" 15 I SOL 0 -10 -20 -30 S U I L W A T E R P O T E N T I A L (M OF WATER) F i g u r e 4. The i n g e s t i o n r a t e o f s o i l f o r L. rubellus Table 8 . Ingest ion ra te of s o i l f or L . r u b e l l u s , incubated at d i f f e r e n t temperatures and moisture regimens. P r o b a b i l i t y of data po int s beiTig d i f f e r e n t ( t - t e s t , * - p o . 0 S , = p < 0. 0 1 , n=24). A. Temperature g r a d i e n t compared. Temp S' 10 IS 20 M o i s t u r e c o n t e n t : 0.33 k g . k g " 1 Temp 5' 10 15 20 0. 30 0 .27 Temp 5' 10 15 20 C. 0. 21 Temp 5 C 10 15 20 H Z ] 5? 10 IS S 10 15 S 10 IS S 10 IS B. M o i s t u r e g r a d i e n t compared. Temperature: 5° C . 10 IS 20 M o i s t u r e c o n t e n t 0.33 0 .33 0 .33 0 .33 0 .30 0 .30 0. 30 0 . 30 0 .27 A A A A 0 .27 0 .27 A A A 0 .27 , ' A A A A 0.24 A A A A A A 0. 21 ft 0.21 A A A 0 . 21 A A A • A A 0 .33 0.30 0 .27 0 .33 0 .30 0 .27 0. 33 0.30 0. 27 0. 33 0. 30 0 .27 2 O to -3 G W > a PI 3: TJ PJ » > -3 G » PI MOISTURE AND TEMPERATURE / 55 Table 9. The dry weights of the earthworms used in the experiments (n=10). Incub. Dry weight (±SD) mg. Temp. (°C) 5 101.64 (23.19) 1 0 106.64 (30.47) 1 5 65.80 (15.43) 20 78.10 (26.23) c l i t e l l a t e d adults (Appendix 3-C), but weight of the adult worms was not s i g n i f i c a n t l y correlated with the egestion rate. 10. S o i l temperature and s o i l moisture in the f i e l d The average weekly s o i l temperatures, measured at 5 cm, in the clover f i e l d on Westham Island varied between 0 °C during a cold s p e l l in December and 18-20 °C during the summer months. The average yearly temperature at 5 cm depth, calculated from the monthly values, was 10.25 °C. The moisture content of the s o i l (0-5 cm) in the f i e l d on Westham Island was a steady 0.39 kg.kg" 1 (-0.3 m of water) during the rainy season, from November to March, and reached MOISTURE AND TEMPERATURE / 56 a low of 0.23 kg.kg"1 (-30 m of water) during the summer. During the winter months, the water content was the highest in the 0-5 cm layer, but in the summer (dry season) i t was the highest in the 5-10 cm layer (Appendix 2-C). 1 1 . Comparison of ingestion and egestion rates to l i t e r a t u r e data The ingestion and egestion rates of L. rubellus weregenerally within the ranges of those found in the l i t e r a t u r e (see Table 10). However, inconsistencies in the reporting of temperatures and moisture contents in the l i t e r a t u r e , made i t d i f f i c u l t to compare the actual values. The ingestion rate for organics and hazel l i t t e r (Satchell, 1967) were similar to the IOM for clover hay, and the ingestion of cow dung i s close to that of a mixture of clover leaves and s o i l . Martin (1982) reported egestion rates on a l i v e weight basis for L. rubellus. When those rates were recalculated on a dry weight basis, assuming worms contained 85 % water, his egestion rates were 10 to 20 times higher than those found by other authors and those found in the present study. Even i f the figures were not recalculated, they were 2 to 3 times higher. The earthworm Allobophora chlorotica , found on Westham Island along with L. rubellus, had an egestion rate three Table 10. Some Ingestion (I) and egestion rates (E) for temperate and tropical earthworms species food Allotobophora caliglnosa A. chIorotIca A. rosea adults . immatures Dendrobaena piatyura D. depressa LumbrIcus rube I I us sol 1 so 1 1 + sol 1 leaves 1 eaves sol 1 sol 1 sol 1 sol 1 + mea 1 cI over grass temp. C O 5, 10 5-15 f i e l d f i e l d 5 15 20 20 mo 1sture (kg.kg ') 0.33 f i e l d f i e l d 0.39 I or E rate (g.g-' day- 1 ) Reference 0.26 -1.7. 7.1-O. 52' 2.9' 9.7' 7.5 - 12.8' 0.09 -0.14 -0. 33' 0. 32 ' 0.65' 0.S2' 12.8' 20.0' P tearce ( 1972 ) Tlmtnenga (unpublished) P h i l l i p s o n and Bolton (1976) Z l c s l (1978) Z 1 C S 1 (1978) present study Carter et a l . (1983) Martin (1982) Martin ( 1982) sol 1 + clover 5-20 range 0 1 . 13 - 1 .0' present study hay woodland 1 11 ter - - 0. 16 - O. 32' Plearce ( 1972) clover leaves 5 0.39 0. 10' present study hazel 11tter - - 0. 14 ' Satchell (1967) cow dung - - 0. 46 - 0. .58' Satchell (1967) L. t errest r is leaves 5 - 0. 12 - O. , 36 ' Raw ( 1962) 1 eaves 9 - 0. 16 - 0. .21 1 Knollenberg et a l . (1985) leaves 23 - 0. 40 - 1 . 20' Satchell (1967) 1 eaves - - 0. 15 - 0 99' Van Rhee ( 1963) Mi croscolex dubfus sol 1 so 11 and mu 1 ch _ _ 20 10 .6' .4' Abbot and Parker (1981) Abbot and Parker (1981) Mi II son!a anomala adult sol 1 25 0.12 33 .0' Lavelle ( 1975) Immatures sol 1 25 0.12 150' Octolaslum lacteum leaf l i t t e r 16 - 1 . 9' Crossley et a l . ( 1971) 2 O n CO -3 a * > a n 3: RJ > a ) Ingestion or removal from surface, ') egestion. Ul MOISTURE AND TEMPERATURE / 58 times higher than L. rubellus when tested in the l i t t e r b a g technique (See Appendix 4). A. chlorotica, an endogeic species, consumed large amounts of s o i l and v i r t u a l l y no organic matter. The t r o p i c a l worm Millsonia anomal a, an endogeic species, had an egestion rate that was 10 to 50 times higher than that of worms from temperate regions (recalculated from Lavelle, 1975). Mi cr os col ex dubi us , another t r o p i c a l species, also showed a high egestion rate (recalculated from Abbot and Parker, 1981). If these values are representative for a l l t r o p i c a l earthworms, i t could be said that t r o p i c a l worms "work harder" than worms from temperate regions. MOISTURE AND TEMPERATURE / 59 12. Drought-survival strategies of Lumbricus rubellus a. The water content of earthworms The water content of adult L. rubellus varied with the s o i l water potential (Figure 5). No s i g n i f i c a n t difference (Kruskal-Wallis K-sample) was found between the body water content (BWC) for immatures subject to varying water potentials, but the BWC of the adults was s i g n i f i c a n t l y influenced by the s o i l water potentials (Mann-Whitney U-test, p < 0.05, Table 11). Immatures may have d i f f e r e n t ways to hold or extract water than adults do; they have di f f e r e n t metabolic demands (Lee, 1985), and a higher egestion rate than c l i t e l l a t e d adults, a l l of which may explain the difference between the body water content of immatures and adults in this experiment. The r e l a t i o n of body water to s o i l water potentials was described by Kuarjaseva (1982) for Eisenia nordenski ol di when she compared the weight of worms from s o i l s with d i f f e r e n t moisture contents with the weight of a "standard worm", a worm weighed afte r incubation on moist f i l t e r paper for 2 days. When her data were related to BWC and s o i l moisture content, a similar r e l a t i o n s h i p was found as i s shown in Figure 5 for adult L. rubellus. The maximum BWC was at a s o i l moisture content of 0.50 kg.kg" 1, but a r e l a t i v e MOISTURE AND TEMPERATURE / 60 SOIL WATER POTENTIAL (M OF WATER) Figure 5. The body water content of L. rubellus, incubated at d i f f e r e n t s o i l moisture potentials (19 °C). T a b l e 11. V a l u e s o f t h e Mann-Whitney U - t e s t i n d i c a t i n g d i f f e r e n c e s between body w a t e r c o n t e n t s o f L . r u b e l l u s , i n c u b a t e d f o r 10 days a t 1 9 ° C . i n s o i l w i t h t h e i n d i c a t e d s o i l w a t e r p o t e n t i a l s (* = p < 0.05, * * = p < 0.01). •p m 3 O Ui •p E -3 -9 -15 •20 •25 -30 •HO •60 m e t e r s o f w a t e r -3 n p e r t r e a t m e n t 8 23.0 14.7* 7.0** 17 .0* 10.0** 48.0 15 .0* 10 .0** 49.5 47.0 13.5* 10 .0** 49.0 45.0 48.0 29 .0 20.0* 22.0* 29.5 22.0* 20.0* 12.5* 27.0 4.0* * 3.5** 1.0** 0.0** 8 .0** -9 -15 -20 -25 -30 -40 8 10 10 10 10 10 -60 10 O cn 50 > z D •-3 W S T3 M JO > 3^ a w P3 CTi MOISTURE AND TEMPERATURE / 62 minimum in BWC in very wet s o i l s was not included. However, the BWC of the "standard worm" was 82 % of the maximum value at 0.50 kg.kg" 1. Stephenson (1945) kept L. terrestris on f i l t e r paper moistened with d i f f e r e n t saline solutions. He found that the worm weights f i r s t decreased with increasing concentrations, then increased and peaked at an external medium concentration of "60 equivalent mMol NaCl" and then decreased. The concentration of "60 equivalent mMol NaCl" was calculated to be equivalent to an osmotic potential of -28.8 metres of- water, while the minimum was found at -14 metres of water. The minima and maxima in worm weight were independent of the type of s a l t s used. The shape of the curve presented in his paper i s similar to the one presented in Figure 5 for adults. It was assumed that L. rubellus has a coelomic f l u i d with an osmotic potential of about -40 metres of water, similar to that of L. terrestris (calculated from Dietz and Alvarado, 1970, and Stephenson, 1945). L. rubellus could therefore ea s i l y absorb water from moist s o i l . In wet s o i l s , the steep gradient would generate an excess of water in the worm i f the water was not ac t i v e l y excreted (Dietz and Alvarado, 1970). The excretion of excess water was also reported by Ramsey (1949) and Wolf (1940), who showed that L. terrestris produced large amounts of urine under saturated or semi-saturated conditions. MOISTURE AND TEMPERATURE / 63 The brain of the worm regulates the salt content of the coelomic f l u i d through neuro-secretory control of dermal permeability and water loss through the nephridia (Carley, 1978; Zimmermann, 1973; Kamemoto et a l . , 1966). This mechanism of excretion i s triggered by changes in the nerve cord induced by the s a l t concentration of the coelomic f l u i d (Laverack, 1963). These strong active processes of water removal from the body could explain the lower body water content of the adult worms in wet s o i l s with water potentials close to -5 metres of water. When the s o i l water potentials were close to zero, an increasing water content may lower the osmotic potential of the coelomic f l u i d of the worm and the worm may reach a steady state where incoming water i s in equilibrium with the water a c t i v e l y excreted. This equlibrium probably i s at the body water content as described by Kuarjaseva (1982) for the "standard worm." b. Feeding behavior of the worm as related to the body water  content The feeding behavior of L. rubellus depended on the water poten t i a l of the s o i l , as was described previously, and the body water content was related to the s o i l water p o t e n t i a l . It i s therefore suggested that the body water content MOISTURE AND TEMPERATURE / 64 influenced the feeding habits of adult worms. In s o i l s with high water potentials, between 0 and -10 m of water (easily available water), the worms showed a s i g n i f i c a n t l y lower body water content and a high egestion rate. Medium potentials (-15 to -20 m of water) created a high but constant body water content but the egestion rate dropped s i g n i f i c a n t l y , and in s o i l s with a low s o i l water potential (< -35 m of water), the worms lost body water (Figure 6). Organic material that was used as food had a higher moisture content than the s o i l in which i t was incubated (Table 12). The worms switched to a diet high in organic matter to overcome moisture stress by eating materials that contained more water. This switch was observed both at 5 and 20 °C. The egestion figures for 10 and 15 °C suggested that under these temperatures the worms were more tolerant to moisture stress, probably because these temperatures were not near the l i m i t s of the temperature range of the species. In dry s o i l , not only was the need for water to mix into the faeces increased, but also as dry s o i l had a more negative water p o t e n t i a l , the absorpton of water by the worm from i t s environment through the skin was reduced. To stop eating can be seen as a strategy to decrease the water loss, as was the regulation of the permeability of the epithelium and MOISTURE AND TEMPERATURE / 65 Figure 6. The body water content (kg.kg" 1 (—o—) , dwt) and the egestion rate (g.g" 1 .day" 1 ) (• >• ••) of adult L. rubellus at 20 °C. MOISTURE AND TEMPERATURE / 66 Table 12. The water content of s o i l and clover straw incubated at 20 °C So i l moisture Clover straw (kg.kg- 1) (kg.kg" 1) 0.34 0.28 0.27 0.24 0.22 3.12 2.63 1.11 0.82 0.79 decreased a c t i v i t y of the nephridia. Also an "escape" reaction down the s o i l p r o f i l e , as was noticed in the f i e l d , may be part of the drought survival strategy as i s the switch to more organic matter in the d i e t . Bouche (1984) c l a s s i f i e d L. rubellus as an earthworm surviving drought stress in the cocoon stage. This may be the case when s o i l water potentials approach the permanent wilt i n g point ( -150 m of water, pF 4.2), but with s o i l water potentials > -60 m of water (pF 3.8), L. rubellus might survive in cracks and deep burrows covered in cast and mucus. Laverack (1963) reported that earthworms may lose up MOISTURE AND TEMPERATURE / 67 to 60 % of their body water and recover. L. rubellus may be able to survive periods of drought by "keeping quiet." 13. Testing of assumption The assumption that the egestion rate of L. rubellus i s not influenced by moisture and temperature i s rejected. The egestion rate i s temperature-dependent in wet s o i l s and negatively influenced by low s o i l water potentials. Low s o i l water potentials may have influenced the water balance of L. rubellus and forced the worm to change i t s behavior (e.g. stop feeding, ingest r e l a t i v e l y more organic matter, burrow down and create a "moist environment." IV. THE TRANSPORT OF ORGANIC MATTER INTO THE SOIL PROFILE BY LUMBRICUS RUBELLUS A. MATERIALS AND METHODS 1. Introduction The t r a n s p o r t of m i n e r a l and or g a n i c matter i n t o the s o i l p r o f i l e a s , w e l l as the d i s t r i b u t i o n of c a s t s i n the p r o f i l e was q u a n t i f i e d by using a column experiment. In t h i s column experiment, earthworms were fed with 1 4 C l a b e l l e d c l o v e r shoot and root l i t t e r and the c a s t was s o r t e d from the columns. The recovered 1 t tC a c t i v i t y was used to c a l c u l a t e the shoot to root r a t i o i n the d i e t of the worm. I t was assumed that L. rubellus, as an e p i g e i c s p e c i e s , i n g e s t s m a t e r i a l s only from the s u r f a c e l i t t e r and produces a l l c a s t s i n t h i s l a y e r ; t h i s assumption i s based on the hypothesis that L. rubellus i s an e p i g e i c s p e c i e s . 2. Animals used in the column experiment The earthworm, Lumbricus rubellus was c o l l e c t e d from a farmer's f i e l d near Abbotsford, B.C. T h i s f i e l d had been i n pasture f o r s i x years and had r e c e i v e d annual a p p l i c a t i o n s of d a i r y c a t t l e manure as s l u r r y . At the time of c o l l e c t i n g , the f i e l d was ploughed to be put i n t o s i l a g e - c o r n . The s o i l in t h i s f i e l d , c l a s s i f i e d by Luttmerding (1981) as O r t h i c 68 TRANSPORT OF SOIL / 69 Humic Gleysol of the Buckerfield series, was of a similar texture class as the Crescent series s o i l used in the column experiment described below. 3. S o i l s u s e d i n t h e c o l u m n e x p e r i m e n t S o i l s used for the column experiment were co l l e c t e d from a farmer's f i e l d on Westham Island. The f i e l d had been under potatoes the previous growing season. The s o i l s were of f l u v i a l o r i g i n and were c l a s s i f i e d by Luttmerding (1981) as s i l t y clay loam-textured Orthic Gleysol (typic haplaquept) of the Crescent seri e s . The top 3 cm of the s o i l was removed and discarded because of a surface crust and alga l growth, and s o i l was col l e c t e d from one spot to a depth of 15 cm. In the laboratory the s o i l was partly dried and then s u f f i c i e n t l y crushed to pass a 5-mm seive. 4 . C l o v e r u s e d i n t h e c o l u m n e x p e r i m e n t a. Production of clover Red clover (Trifolium prat ense L.) was grown from seed in the greenhouse. The s o i l medium used consisted of a s t e r i l i z e d greenhouse s o i l - t u r f a c e f mix (1:1); t h i s medium was inoculated with rhizobium but no nutrients were added. ft u r f a c e : baked montmorillonite clay. Trademark of IMC-IMCORE, Munde.lein, 111. TRANSPORT OF SOIL / 70 After 81 days the clover was harvested by washing s o i l and turface from the roots. Only plants that did not show flowering stems were co l l e c t e d to reduce v a r i a b i l i t y in p a l a t a b i l i t y . The plants were separated in 22 bundles and sub-samples were taken of shoots and roots for wet weight and dry weight determinations. Six bundles of fresh plant material were then put in a 1:4 d i l u t i o n of Long-Ashton nutrient solution and the roots were aerated overnight. These plants were used for the 1"C l a b e l l i n g . The remainder of the plant material was stored in the freezer u n t i l used in the experiment. b. Radiolabelling of clover In a sealed plexiglass box, the fresh plants were exposed to one pulse of 2000 jig . g - 1 C0 2 from Na 2C0 3 containing 250 nCi from NaH1 "C03 (110 /xCi.g" 1). The r a d i o l a b e l e d NaHC03 was dissolved in 5 mL of an al k a l i n e 0.5 M solution of Na 2C0 3. This solution was then injected through a membrane-covered port into a beaker suspended in the plexiglass box. The C0 2 was lib e r a t e d through inje c t i o n of 5 mL of a 4 N solution of HCl. To s t i r the solution, 5 mL of d i s t i l l e d water was injected with force. A small battery-powered fan c i r c u l a t e d the a i r in the box (See Appendix 2-A). TRANSPORT OF SOIL / 71 After 10 minutes an i n i t i a l a i r sample was taken by p u l l i n g 5 mL of a i r into a syringe f i l l e d with 5 mL of a 10 % KOH solution. After shaking for 20 seconds, the solution was put into a v i a l and a 1-mL aliquot was taken and added to 9 mL of ACSf s c i n t i l l a t i o n f l u i d . The sample was then counted in a s c i n t i l l a t i o n counter. After the sample was taken, l i g h t s (500 Watts) were switched on for 8 hours and further a i r samples were taken every 30 minutes. Lights were not applied during the night. The next morning, 10 mL of the Na 2C0 3 solution and 10 mL of the HCI solution were injected into the beaker and after four hours under l i g h t s , the box was aerated by p u l l i n g the a i r through a 10 % KOH solution to catch any surplus 1"C0 2. A l l plant material was then stored in the freezer before use. 5. Experimental Set-up The transport of organic matter into the s o i l p r o f i l e was studied using a column experiment. In a p l a s t i c tent (6 mil polyethelene) inside an incubator, columns were set-up, made from ABS p l a s t i c sewer pipe (10 cm diameter) and closed and sealed at the bottom with an ABS p l a s t i c cap. The cap had an outlet and was connected to an overflow unit with teflon tubing. With the overflow unit, the water table was fTrademark of Amersham Ltd., Chicago. TRANSPORT OF SOIL / 72 established at 1 cm below the surface of a 3 cm thick layer of sand (0.25 - 1 mm) that was used as an unconsolidated porous plate in the bottom of the column. The water table was kept constant by means of a p e r i s t a l t i c pump, c i r c u l a t i n g water to the overflow unit (Appendix 2-B). On top of the sand, 30 cm of s o i l of the Crescent series was placed. The columns were capped with 500 mL p l a s t i c beakers. Each of these beakers had an open a i r i n l e t and an outlet connected to a C0 2 trap and a i r was drawn through the system with an aspirator. After the s o i l in the columns was s u f f i c i e n t l y wetted to f i e l d capacity, 8 g (wet weight) of previously frozen root material was placed in each column, covered with 5 cm of s o i l and then previously frozen clover shoot material (12 g wet weight) and five earthworms were added to each column. The columns were recapped and sealed with petroleum j e l l y and the tent was sealed with tape. The temperature was kept at 10 °C and a 10 hour l i g h t regimen was imposed. At the end of the 30 days incubation period, a l l columns were stored in a freezer. Three treatments were used: six columns with l a b e l l e d shoot l i t t e r and non-labelled root l i t t e r , six columns with non-labelled shoot l i t t e r and la b e l l e d root l i t t e r and six TRANSPORT OF SOIL / 73 controls with neither material l a b e l l e d . The columns in a l l three treatments contained earthworms. To deal with the microbial decomposition of the clover materials, two columns were added to each of the two treatments with lab e l l e d materials. These "system controls" did not contain earthworms. One column containing non-labelled l i t t e r , was equipped with tensiometers at 4, 14 and 24 cm s o i l depth to measure s o i l water potentials in the s o i l column. 6. Experimental design and s t a t i s t i c s a. Experimental design The experimental design was a s p l i t - p l o t design, where the whole plots (columns) were completely randomized and two samples per plot were taken. The six sampling depths represented the sub-plot e f f e c t . Three treatments (la b e l l e d shoots, lab e l l e d roots, con t r o l ) , six r e p l i c a t e s (blocks per treatment) and two samples per block (halves of column) were chosen to maximize the sub-plot (depth) e f f e c t s and minimize the block e f f e c t s . A t o t a l of 36 samples of six depths each ( L, 0-5, 5-10, 10-15, 15-20 , 20+ cm) were taken. TRANSPORT OF SOIL / 74 b. S t a t i s t i c s One way analysis of variance was used to describe depth e f f e c t s . When i t was not possible to do an analysis of variance due to incomplete data sets (dpm in cast, percent carbon), the equality of means was tested with a Kruskal-Wallis K-sample test, while means were separated with a Mann-Whitney U-test (Siegel, 1980). 7. Sample preparation Before sorting through the columns, they were removed from the freezer (-25 °C) and stored overnight in a refigerator (+4 °C). The partly thawed s o i l columns were then pushed out of the p l a s t i c containers, s p l i t length-wise and the cast was sorted out from each of fi v e depth layers and the l i t t e r layer. A i r - d r i e d casts were resorted and only p a r t i c l e s that were p o s i t i v e l y recognized as casts, were included in the samples. Casts and the remains of the plants were then oven-dried (65 °C) and weighed. For the chemical analysis, casts and s o i l were crushed with mortar and pestle to pass a 1 mm seive. Plant materials were crumbled by hand. TRANSPORT OF SOIL / 75 8. Chemical Analysis The C0 2 traps, 250 ml erlemeyer flasks with a long and a short glass tube (4 mm inside diameter) inserted through holes d r i l l e d in a rubber stopper, were f i l l e d with 100 mL of a 0.1 N KOH solution and were changed every three to four days. A 1 mL aliquot was taken from each trap and added to 10 mL of s c i n t i l l a t i o n f l u i d and the a c t i v i t y was determined. In non-labelled controls, t o t a l C0 2 was determined by t i t r a t i o n of a 25 mL aliquot with 0.1015 M HCl with phenolphthalein as indicator. An excess of BaCl 2 was used to s t a b i l i z e the carbonates. The t o t a l carbon and the l a b e l l e d carbon were determined with a method derived from Snyder and Trofymo (1984) and Coughtrey et a l . (1986). In modified culture tubes (Snyder and Trofymo, 1984), s o i l , plant and worm samples were digested in a H 2SO a-H3PO t t-K 2Cr 20 7 mixture and carbon dioxide was trapped in 3 mL of a 2 M NaOH + 0.2 M Na 2C0 3 solution (Coughtrey et a l . , 1986). A 1 mL aliquot was taken from the trapping solution and was added to 1 mL NCSf s o l u b i l i z e r and aft e r vigorous shaking, mixed with 10 mL OCSf s c i n t i l l a t i o n c o c k t a i l . The a c t i v i t y was determined in a Packard LCS 4530 s c i n t i l l a t i o n counter, using internal standards and automatic e f f i c i e n c y c o n t r o l . Another 1 mL aliquot was transferred to a reagent tube and t i t r a t e d with 1.226 M HCl. •(•Trademark of Amersham Ltd. TRANSPORT OF SOIL / 76 For t h i s t i t r a t i o n , 3 mL of a 1 M BaCl 2 solution was added to s t a b i l i z e the carbonates and phenolphthalein was used as an indicator. B. RESULTS AND DISCUSSION 1. Production of clover At the time of harvest (after 81 days), clover plants were f u l l y grown and some had developed flowering stems. Only plants without flowering stems were c o l l e c t e d . The average shoot weight was 0.88 ± 0.450 g (dwt, SD, n = 13), with a dry matter content of 14.75 ± 1.65 %. The average root weight was 0.146 ± 0.066 g (dwt, n = 13) with 7.9 ± 1.12 % dry matter. The shoot to root r a t i o was 6.03. 2. 1 4 C 0 2 fumigation of the clover Approximately 360 g of fresh plants were placed in a sealed 52.6-L plexiglass box and 1"Carbon was added. After 210 minutes under 500-watt l i g h t s , v i r t u a l l y a l l labeled C0 2 was absorbed by the plants, No decrease in r a d i o a c t i v i t y (dpm) could be detected afterwards (Table 13). Overnight, an increase of a c t i v i t y was observed as plants respired C0 2 under low l i g h t conditions. TRANSPORT OF SOIL / 77 Table 13. 1"C A c t i v i t y in a i r samples measured during the fumigation of clover plants with l a b e l l e d C0 2. Time (min) dpm per sample 0 30 60 90 1 50 210 330 Overnight 61 20 1823 214 1 15 92 71 67 1141 3. Observations on materials and techniques used in the column experiment. a. Micro-arthropods in the s o i l Tullgren-funnels (10 cm screen, variable l i g h t 25 - 100 Watts; Edwards and Fletcher, 1971) were used to extract the s o i l before i t was placed in the columns. The extractions did not reveal any micro-arthropods. Micro-arthropods would have caused errors in the results by ingesting and transporting l a b e l l e d material. P a r t i a l l y drying, crushing TRANSPORT OF SOIL / 78 and sieving the s o i l eliminated a l l l i v i n g micro-arthropods. No extraction was done when the experiment was finished because the columns were immediately stored in the freezer. b. Plant material Discarding most of the stem material, only tops of plants and the complete roots were used as food in the columns. The dry matter content of the plant tops was 15.5 %, that of the roots 7.9 %. In the columns, 8 g of root material and 12 g (wet weight) of shoot material was used, equivalent to 0.8 and 2.4 t.ha" 1 (dwt) respectively. The calculated shoot to root r a t i o of the materials used in the columns was 3, about half of that of the young, fast growing plants in the greenhouse. The amount of shoot material i s close to the equivalent of one cut of clover hay in a four cut clover management system (Russell, 1977). c. Experimental temperature and s o i l moisture content The experimental temperature, the high moisture content and the short l i g h t regimen would r e f l e c t spring or f a l l conditions in the Lower Fraser Valley. In the f i e l d , earthworms were very active under these conditions. TRANSPORT OF SOIL / 79 d. Diffusion method for carbon analysis The determination of carbon with the "d i f f u s i o n method", gave higher results than those obtained with the Leco carbon method. During the digestion, the trapping solutions in the di f f u s i o n tubes tended to evaporate, as mentioned by Coughtrey et a l . , (1986). Snyder and Trofymo (1984) d i r e c t l y t i t r a t e d the trapping solution without taking sub-samples and did not encounter these problems. Using a correction factor based on results from a standard s o i l (as was done in the reported experiments) measuring the volume of the trapping f l u i d before taking sub-samples, as was suggested by Coughtrey et a l . (1986) or bringing the solution up to volume, w i l l correct t h i s difference. 4 . Airflow above the columns In order to remove C0 2 from the columns the t o t a l airflow was aimed at 600 mL.min"1. The actual flow ranged between 400 and 700 mL.min"1, because of fluctuating tap water pressures. An aspirator was used to create suction. The flow through the beakers closing the columns, was approximately 27 mL.minute - 1, creating an air-change every 37 minutes in the 500 mL above the columns. This flow was s u f f i c i e n t to prevent a build-up of C0 2. A l l a i r was drawn from inside the tent, creating an air-change in the tent every 14 hours. TRANSPORT OF SOIL / 80 5. S o i l water potentials in the columns Before the earthworms were introduced, the columns were equilibrated with water for 6 days. During t h i s period, the pressure potential increased to -0.8 m of water. The s o i l reached a steady state 9 days after introducing the earthworms, as can be seen in Figure 7. The fluctuations in s o i l water potential in the s o i l near the surface, might have been caused by changes in the flow of a i r through the beaker, that capped the column. The humidity inside the incubator was kept at 50 %, while the a i r above the s o i l may have been saturated. The a i r drawn through the system was not C0 2 free, therefore the variation in C0 2 trapped from columns may indicate a changing airflow. In column 15, with tensiometers, high amounts of C0 2 trapped, indicating a high airflow, coincided with low tensiometer readings, indicating a high evaporation rate during these periods. 6. Recovery of 1"C a c t i v i t y from the samples a. Spe c i f i c a c t i v i t y of the recovered materials Specific a c t i v i t i e s (AtCi.g" 1 of carbon) of a l l major components are l i s t e d in Table 14. Worms fed on labelled roots showed a lower s p e c i f i c a c t i v i t y than those fed on labe l l e d shoots. This difference in a c t i v i t y between roots and shoots was also noted in the recoverd cast. A large drop TRANSPORT OF SOIL / 81 cm of wottr Figure 7. Pressure potentials at d i f f e r e n t times, in the s o i l column during the incubation of earthworms (10 °C). TRANSPORT OF SOIL / 82 Table 14. Specific a c t i v i t i e s (yCi.g~ 1Carbon) of materials recovered from the s o i l columns. Labelled roots Labelled shoots Cast 0.0304 0.562 Worms 0.0264 2.68 L i t t e r before 3.23 3.67 L i t t e r after 0. 1 52 3.24 Av. Littert- 0.630 3.46 t-Average l i t t e r value is calculated from ' l i t t e r before' and ' l i t t e r a f t e r ' , using a weighted average, based on the evolution curves in Figure 8. in s p e c i f i c a c t i v i t y was found in root material over the course of the experiment. The high l i g n i n content of the root material and the leaching of carbohydrates from th i s material may have caused th i s drop in s p e c i f i c a c t i v i t y . In pul s e - l a b e l l i n g experiments with 1*C0 2, most of the carbon is incorporated in mobile carbohydrates and not in stru c t u r a l components of the plants. TRANSPORT OF SOIL / 83 b. Total a c t i v i t y of recovered materials The calculated t o t a l a c t i v i t y in the plant materials, used in the column experiment was 30.95 MCi. At the end of the experiment 20.77 uCi could be accounted for (Table 15). Losses may be caused by the contamination of the bulk s o i l , i n e f f i c i e n c y in sorting the casts from the columns and losses of 1"C0 2 during sorting and storage. Inefficiency in sorting may lead to an under-estimation of the egestion rate. Losses of 1"C0 2 during storage may aff e c t equally shoot or root materials, because the largest losses take place in the early stages of the experiment. No effect on the shoot to root r a t i o in the earthworm diet i s to be expected due to losses of a c t i v i t y from plant materials. c. 1"C a c t i v i t y recovered from the bulk s o i l Labelled shoots caused a s l i g h t increase of a c t i v i t y above background levels in the s o i l layers immediately below the surface. Above the lab e l l e d roots however, a very s i g n i f i c a n t increase in a c t i v i t y was noted (739 dpm.g"1 s o i l ) , most l i k e l y caused by the upward movement of moisture and gases in the column. In the f i e l d , normally a down flow of water takes place. Carbohydrates and other components, exuded by plants may move down into the p r o f i l e with the water. The downward movement of radio-labelled components exuded by plants that were treated with 1"C0 2 in the f i e l d , TRANSPORT OF SOIL / 84 Table 15. Total 1*C a c t i v i t y recovered from materials in the s o i l columns. Source //Ci In Shoots 23.58 Roots 7.37 Total in 30.95 Out Respiration 8.86 Cast roots 0.09 Cast shoots 1 .88 Worm roots 0.03 Worm shoots 2.65 Roots 0.14 Shoots 3.01 Roots blank 0.06 Shoots blank 2.00 S o i l roots 1 .07 S o i l shoots 0.98 Total out 20.77 TRANSPORT OF SOIL / 85 was described by Dietz and Bottner (1981). The components could be detected by radiography in the top 5 cm of the mineral s o i l . Casts col l e c t e d from the surface were not affected by this contamination (344 dpm.g"1 cast), but the cast recovered from the s o i l layers may have been affected, because large differences in a c t i v i t y between the cast from the surface and from the s o i l layers were found. A correction was made in the a c t i v i t y by subtracting 739 dpm, before the calculations were done to estimate the use of roots and shoots by earthworms. Further discussion of the difference in levels of 1ttC a c t i v i t y i s found in section 11 b. 7. Respiration and decomposition a. Respiration Column res p i r a t i o n , expressed as trapped 1 i ,C0 2, showed a high v a r i a b i l i t y within each treatment group, but the var i a t i o n was related to each column. This v a r i a b i l i t y may be explained by the position of the plants in the fumigation box. Plants at the side of the l i g h t source might f i x more C0 2 than those in the shade. Plants were not mixed before being used in the columns. Also s o i l packing might have caused v a r i a b i l i t y . When measured a c t i v i t y counts were related to those of the f i r s t sampling date, and expressed TRANSPORT OF SOIL / 86 IOO -% 5 0 -3 6 9 13 16 2 0 23 27 30 DAYS F i g u r e 8. Decrease i n 1 , C a c t i v i t y (% f i r s t measurement, SD) of C 0 2 r e l e a s e d through r e s p i r a t i o n . TRANSPORT OF SOIL / 87 as a percentage, the v a r i a b i l i t y was reduced (See Figure 8). The evolution rate of '"COj, expressed in average a c t i v i t y per day, varied from approximately 15000 dpm at day 3 to 1724 dpm at day 30 for columns with l a b e l l e d leaves and from 1600 to 210 dpm for columns with l a b e l l e d roots. The largest drop in evolved a c t i v i t y occurred during the f i r s t few days of the experiment. A t o t a l of 8.84 uCi was recovered. The background for thi s stage of the experiment was 35.8 ± 6.17 dpm (n=22) The trapped C0 2 and the recovered a c t i v i t y from columns used as system controls (no worms added) were not s i g n i f i c a n t l y d i f f e r e n t from the treatments. The earthworms did not have a noticeable e f f e c t on the rate of 1 U C 0 2 evolution, probably because the 1*C a c t i v i t y was confined to mobile carbohydrates. In a pu l s e - l a b e l l i n g experiment, only a limited amount of 1 t tC0 2 i s incorporated in the structural components of plants. These mobile carbohydrates, sugars etc., are ea s i l y assimilated by microorganisms inhabiting the plant material. The decomposition of str u c t u r a l components in later stages of the decomposition process, may be affected by shredding and mixing by earthworms and by the p r o l i f e r a t i o n of fungi and bacteria caused by earthworms. (Shaw and Pawluk, 1986). Earthworms may not have a TRANSPORT OF SOIL / 88 s i g n i f i c a n t effect on the decomposition of organic matter in the early stages of decomposition. b. Weight loss of clover material Shoot material lost 56 % of i t s o r i g i n a l weight in the columns without worms over the duration of the experiment, probably due to non-faunal decomposition and other processes. Root materials l o s t only 6.9 % of their weight. The constant temperature and high humidity in the columns, may have aided the decomposition processes. Weight loss during the decomposition of clover plants was described by Uvarov (1982). He found a weight loss of 60 % during the f i r s t year. Uvarov (1982) c i t e d several studies, reporting weight losses from clover stems and leaves as high as 61 %, over a period of 2 to 3.5 months. Thus the high weight losses during the 30 days of the experiment may not be unusual. The 1 *C a c t i v i t y in casts recovered from columns with la b e l l e d shoot material was substantially higher than that in cast from columns with lab e l l e d root material, while the s p e c i f i c a c t i v i t i e s of both shoots and roots were similar at the sta r t of the experiment. Differences in ingestion rate and a di f f e r e n t decomposition pattern of roots may explain t h i s difference. For shoots, the decomposition of soft TRANSPORT OF SOIL / 89 materials and a breakdown of simple carbohydrates takes place simultaneously; the s p e c i f i c a c t i v i t y stays constant. Roots, however, have a r i g i d structure and simple carbohydrates are leached from this material before a substantial weight loss is seen from the breakdown of ce l l u l o s e and l i g n i n . Malone and Reichle (1973) demonstrated thi s c l e a r l y when they followed l a b e l l e d cesium during the decomposition of root material. The cesium was lo s t from the roots much faster than the loss of biomass would indicate. 8. Recovery of casts from the s o i l columns a. Description of the burrows Cast p a r t i c l e s that could be recovered, were removed from the surface of each column. The partly frozen s o i l was then pressed out of the p l a s t i c columns. In the 0-5 cm layer, the burrow walls ( a l l cast material) were thick and easy to recognize and recover. Some of the burrows were blocked with cast aggregates. Along the sides of the columns, the worms had created "reinforced" burrows made of large amounts of casts. In the 5-10 cm layer, the burrow walls were not very thick but could e a s i l y be removed by peeling material from the surrounding s o i l . The 10-15 cm layer, and those below, contained burrows along the sides of the column. The s o i l at the sides of the column might have been less packed, TRANSPORT OF SOIL / 90 increasing aeration and pen e t r a b i l i t y and was thus more accessable to earthworms. This border ef f e c t and the optimum diameter of columns should be examined more c l o s e l y . These burrows along the sides of the column had very thin walls with v i r t u a l l y no recoverable casts and some burrows showed only s l i g h t l y smeared tracks. The small amounts of casts recovered from these burrows may indicate that the worms did not use them often. In several columns, worms were recovered from these deep burrows. Disturbance due to the discontinuation of the experiment and the transport of the columns might have caused an escape reaction forcing the worms down. The burrows with thin walls, might have been dug during the f i r s t few days of the experiment as part of a panic or escape reaction as discussed for A. rosea by Ph i l l i p s o n and Bolton (1976). No reports were found to confirm this behaviour of L. rubellus. In the f i e l d i t was noted that the worms p u l l into their burrows when they were disturbed. Burrows were found in the s o i l mass close to the water table. Due to the high s o i l water content, the burrow walls were smeared and no cast could be recovered. These deep burrows might indicate that a high s o i l water potential i s not a r e s t r i c t i n g factor in the burrowing depth of L. rubellus. The water table i t s e l f , anaerobic conditions or TRANSPORT OF SOIL / 91 toxic gas (H 2S) may be the regulating factors. b. Description of casts Casts were r e l a t i v e l y easy to recover because of their compact structure, irregular rounded and indented shapes, their colour (Munsell: 5Y, 3/2, dark o l i v e green or darker, when wet), the high amount of fibres in them and the lack of mottles inside broken-up aggregates. The bulk s o i l was of a loose structure, with blocky aggregates with sharp edges. The colour was l i g h t e r than that of the casts and less green (2.5Y 4/2, dark greyish brown, when wet) and not many fibres were v i s i b l e . 9. D i s t r i b u t i o n of casts The average amount of cast recovered from each column was 34.28 ± 3.85 g (n=l8). Columns from which less than 5 worms were recovered (10 % mortality was observed), showed a cast production that was s l i g h t l y less than the average (27.48 ± 4.66 g, n=8). Because the mortality occured systematically in a l l treatments no d i s t i n c t i o n was made regarding the recovery of worms. (Control = 30.76 ± 8.15; Leaves labelled = 32.63 ± 2.94; Roots l a b e l l e d = 30.38 ± 4.42 g, SD, n=6). Few casts were found in the 20+ cm layer, the bulk was found in 0-5 and 5-10 cm layers. The d i s t r i b u t i o n of t o t a l cast TRANSPORT OF SOIL / 92 T a b l e 16. F - v a l u e s for c o n t r a s t s of t o t a l c a s t , r e c o v e r e d from columns, a f t e r i n c u b a t i n g earthworms wi th l a b e l l e d c l o v e r shoot or root m a t e r i a l added . T o t a l c a s t r e c o v e r e d ; the l i n e a r t e s t was performed f i r s t (** p > . 99 , * p > . 9 5 ) ; SS = sum of squares , MS = mean s q u a r e ) . ANALYSIS OF VARIANCE - CAST Source o4 v a r i a t i o n cH F SS MS Treatment (T) 2 0. 35 1.B60 0. 940 C o n t r o l vs T k B ( T l ) 1 0.05 0. 13B 0. 138 Top vs . Bottom (T2) 1 0.64 1.742 1. 742 P o t / T - E r r o r (a) 15 0.54 40.B14 2. 721 C o r e / P / T - E r r o r (b) IB 1.60 90.270 5. 015 Depth (D) 5 B0. 84 ** 1264.835 . 252. 967 L i t t e r / o t h e r (DO) 1 1.27 3.960 3. 9B0 L i n e a r (DI) 1 363.01 ** 1135.937 1135. 937 G u a d r a t i c (D2) 1 39. B9 ** 124.641 124. 841 R e s i d u a l A. 0.01 0.076 0. 039 Treatment X Depth (TD) 10 1.64 51.461 5. 146 T l X DO 1 0. 10 0.316 0. 316 T l X DI 1 7.98 ft 24.971 24. 971 T l X D2 1 6.77 * 21.182 21. 182 T2 X DO 1 0.00 0.001 0. 001 T2 X DI 1 0.59 1.B34 1. 834 T2 X D2 1 0. 20 0. 632 0. 632 R e s i d u a l 4 0. 20 2.524 0. 631 D X P / T - E r r o r (c) 75 1.41 234.693 3. 129 D X C /P /T - E r r o r <d> 90 199.579 2. 216 T o t a l 215 1683.533 TRANSPORT OF SOIL / 93 was h i g h l y n e g a t i v e l y c o r r e l a t e d with depth, both l i n e a r l y and q u a d r a t i c a l l y (See Table 16). The percentage of t o t a l c a s t per column, recovered from each l a y e r from i n d i v i d u a l columns, gave a b e t t e r i n d i c a t i o n of the d i s t r i b u t i o n , because d i f f e r e n c e s i n c a s t p r o d u c t i o n among columns were e l i m i n a t e d (Figure 9). The order of t e s t i n g the polynomials appeared important: when the l i n e a r t e s t was performed f i r s t , 90 % of the v a r i a b i l i t y c o u l d be e x p l a i n e d with a l i n e a r equation, when the q u a d r a t i c t e s t was done f i r s t , 85 % of the v a r i a b i l i t y was a t t r i b u t e d to a q u a d r a t i c curve. Because the second t e s t i s done on the r e s i d u a l of the f i r s t one, the d e s c r i p t i o n of the curves depends on the sequence of t e s t i n g (Eaton, p e r s o n a l communication). 10. E g e s t i o n r a t e s The e g e s t i o n r a t e of L. rubellui was 2.34 g.g"May" 1 , c a l c u l a t e d f o r columns from which 5 worms were recovered. The average dry weight of the earthworms, measured before the experiment, was 0.094 ± 0.0238 g (n=13). The e g e s t i o n r a t e c a l c u l a t e d from the column experiment i s c l o s e to the r e s u l t s r e p o r t e d by M a r t i n (1982) f o r the e g e s t i o n of s o i l and grass meal (not converted to dry weight b a s i s ) . The eg e s t i o n r a t e i n the column experiment was higher than was TRANSPORT OF SOIL / 94 CM 0 10 I 5 20 25 20 30 40 50 % total cast F i g u r e 9 . D i s t r i b u t i o n of c a s t ( % of t o t a l c a s t per column, SD) a c c o r d i n g t o d e p t h . TRANSPORT OF SOIL / 95 found in the l i t t e r b a g technique (Chapter III) and was well above figures found in the l i t e r a t u r e for L. rubellus (see also Table 10). Different food or dif f e r e n t experimental conditions (temperature and moisture) may have caused the these differences in egestion rate. Further discussion may be found in section 11 c. Using a weighted average for the percent carbon in cast as related to depth, a t o t a l of 46.13 mg.g" 1.day 1 carbon was egested by the worms. No information was found in the l i t e r a t u r e on the egestion rate of carbon for L. rubellus or any other species. 1 1 . Organic matter a. D i s t r i b u t i o n of organic carbon in the cast Organic matter was redistributed in the s o i l p r o f i l e by the worms in two ways: through ingestion and casting and through physically p u l l i n g material into the burrows. The percentage of carbon in the cast indicates the amount of organic matter ingested by the worms. The carbon content of the cast was higher than that of the ingested s o i l (1.8 % vs. 1.24 % ) , but no s i g n i f i c a n t decreasing trend with depth was found (Kruskal-Wallis K-Sample test, Figure 10). TRANSPORT OF SOIL / 96 1 litter layer — 4 j ! • — , 1 "o i CO 1 1 1 1 1 0 2 0 % C F i g u r e 10. D i s t r i b u t i o n o f C a r b o n (% C a r b o n , SD) i n c a s t a c c o r d i n g t o d e p t h . TRANSPORT OF SOIL / 97 b. 1 I |C a c t i v i t y in casts as related to depth •For shoots no s i g n i f i c a n t decrease in 1*C a c t i v i t y was noted in the cast related to the depth. A large v a r i a b i l i t y (50 %) was found in the 1"C a c t i v i t y in these casts. This large v a r i a b i l i t y could be explained by the non-homogenized lab e l l e d plant materials used in the columns as earthworm food. Casts from columns with lab e l l e d roots, showed a d i f f e r e n t pattern. Casts found on the surface of the s o i l had a low a c t i v i t y (344 ± 29.5 dpm.g-1 cast, dwt), while cast in the s o i l layers was much higher (> 1000 dpm.g-1) with large v a r i a b i l i t y (80 % ) . A non-significant decrease in a c t i v i t y with depth was found. The contamination of the bulk s o i l above the r o o t - l i t t e r layer (approximately 700 dpm.g"1), may be proof of the rapid assimilation of mobile carbohydrates by microorganisms. The r e l a t i v e l y high contamination in the bulk soi] may also have influenced the le v e l of a c t i v i t y in the sub-surface cast. These carbohydrates were leached from the root material, as was discussed in 7 b. No contamination of the bulk s o i l was seen in columns with l a b e l l e d shoots and the a c t i v i t y of surface cast was not lower than that of sub-surface cast. The 1 < 4C a c t i v i t y in the cast in these columns was two orders of magnitude higher than in cast in columns with labelled roots. TRANSPORT OF SOIL / 98 c. Calculation of the use of added organic matter by the  earthworms Shoot material was almost completely removed from the surface of the s o i l , while most of the root material was not moved. In the zone where the roots were buried, horizontal burrows were found, but the root material i t s e l f was consumed only s l i g h t l y by the worms, indicating a low p a l a t a b i l i t y of the root material or a lack of a microorganism population on the material. Either cause may be supported by the fact that mobile carbohydrates leach from the roots in the early stages of the decomposition process. Invasion by l i g n i n decomposer populations may take longer than the 30 days the experiment lasted. After the decomposition weight-loss was subtracted from the o r i g i n a l amount of l i t t e r , approximately 15.4 % of the shoot material, mostly stems, was recovered from the columns with wovms. About 73 % of the roots were recovered, the loss by decomposition was minimal (6.9 % ) . Based on these ingestion figures, roots accounted for 17.9 % and shoots for 82.1 % of the consumption of added organic matter. The 1"C a c t i v i t y measured in the recovered earthworms was used to calculate the feeding by the worms on the shoot or root materials that were added to the columns. In t h i s TRANSPORT OF SOIL / 99 ca l c u l a t i o n , a weighted average of the evolvement of la b e l l e d C0 2 was used to correct for the decrease in s p e c i f i c a c t i v i t y of the root materials during the experiment. Roots contributed 17.8 % of the added organic matter that was ingested by the worms, while shoot material accounted for 82.2 %. Using a similar approach with the weighted average, the use of organic matter was calculated from the 1"C a c t i v i t y recovered from the cast materials. The calculated use of roots was s l i g h t l y higher (21.8 %) and the use of shoot material was s l i g h t l y lower (78.1 %) than was found in the previous c a l c u l a t i o n s . A l l three independent methods, the ca l c u l a t i o n from the ingestion of organic matter, from the a c t i v i t y in the worms and from the a c t i v i t y in the casts, gave similar r e s u l t s . In th i s type of column experiment, the use of added organic matter could have been estimated without the use of a radio-tracer. In experiments where the root materials are mixed into the s o i l , tracers are d e f i n i t e l y an asset to estimate the r e l a t i v e consumption of shoots and roots by earthworms. The shoot to root r a t i o in the diet i s v a l i d early in the decomposition of the plant materials. The p a l a t a b i l i t y of roots may increase over time, when a TRANSPORT OF SOIL / 100 decomposer population develops. On the other hand, L. rubellus is known to ingest f a i r l y undecomposed materials. The r a t i o may not change that much. 12. Testing of assumption L. rubellus burrowed to a depth of 35 cm, was found throughout the columns, deposited only 15 % of i t s cast on the surface of the s o i l and was active in the top 15 cm of the p r o f i l e where most of the cast was deposited. The worm ingested 20 % l i t t e r as root material. Based on thi s evidence, the assumption that L. rubellus i s an epigeic worm, only feeding and casting in the l i t t e r layer, must be rejected. V. EARTHWORM SIMULATION MODELS A. SIMULATION MODELS DESCRIBING THE DYNAMICS OF SOIL MIXING BY EARTHWORMS 1. Introduction Models have been used to increase the understanding of ecological systems, to predict the results of individual processes or a combination of processes or to predict the results of a single laboratory experiment. Several conceptual and simulation models have been developed for earthworms. The conceptual models describe the parameters that influence the d i s t r i b u t i o n of earthworms (Reynolds and Jordan, 1975) or the consequences of earthworm a c t i v i t y (Bouche and Kretzschmar, 1977). Simulation models estimate the population growth and/or s o i l turn-over for a single species over a r e l a t i v e l y short period. 2. Published earthworm models Reynolds and Jordan (1975) developed a conceptual model to describe the d i s t r i b u t i o n of earthworms. Many s o i l - r e l a t e d factors such as moisture, temperature, colour, aspect and texture were included. 101 EARTHWORM SIMULATION MODELS / 102 The consequences of earthworm a c t i v i t y in the s o i l p r o f i l e were described in a conceptual model by Bouche and Kretzschmar (1977). This model, "REAL," included population dynamics, mechanical a c t i v i t i e s of earthworms in the s o i l and the possible e f f e c t s of worm a c t i v i t y on the ecosystem. Growth and development of populations was described by the population dynamics model "MOTOMURA." This model was tested for earthworm populations across Europe by Lecordier and Lavelle (1982). The model could be used in 80 % of a l l cases, but the authors rejected i t because no relation could be established between the slope value in the model (the environmental constant) and any environmental factor in the f i e l d . Reichle (1971) described a mathematical model to simulate the carbon flow in a woodland system, containing earthworms. Carbon from five components i s followed: l i t t e r 0, and 0 2, mineral s o i l , earthworm biomass and extractable earthworm biomass. The ingestion rate was related to the Q 1 0f the re l a t i v e increase in assimilation over a 10 °C i n t e r v a l . Reichle's model u t i l i z e d data for Octolasium lacleum obtained by Crossley et a l . , (1971). A population of 14 g.irr 2 ingested 208 g.nr 2.y~ 2 of organic matter, and egested 167 g.m"2.y"2 with 40 g. i r r 2 . y ~ 2 being assimilated. EARTHWORM SIMULATION MODELS / 103 The model "WORM.FOR" was presented by Mit c h e l l (1983) to describe biomass production and food consumption by E. foetida in waste conversion systems. This model used functions based on empirical information. The growth-rate function was related to the growth conditions and the Q 1 0 was included in the description of temperature e f f e c t s . Four subroutines, mortality, growth, reproduction and ingestion were used in weekly steps to calculate biomass production and food consumption. Cast production of the t r o p i c a l earthworm M. anomala, was f i r s t calculated by Lavelle (1975), using a model based on population data and earthworm a c t i v i t y . This model was the basis for a simulation model "ALLEZ-LES-VERS" (Lavelle and Meyer, 1977, 1983). The population growth was the basis of the simulation. S o i l moisture and s o i l temperature data were used to calculate the turn-over of s o i l and organic matter, as they both influence earthworm a c t i v i t y . The simulation models presented above, describe the growth and a c t i v i t y of a single species in a s p e c i f i c environment during a short period. WORM.FOR dealt with E. foetida in waste conversion systems in a simulation over 20 weeks, ALLEZ-LES-VERS simulated the egestion of s o i l and the population dynamics of M. anomala in the Lamto savanna, EARTHWORM SIMULATION MODELS / 104 Ivory Coast, during a period of a year; and Reichle's model is s p e c i f i c for an Eastern North American hardwood forest, also for one year. None of these simulation models estimate the long-term effects of earthworms on the s o i l system. Information on long-term effects may be important for the reclamation of waste lands and in the study of "ecological agriculture" and in forest management. A s o i l mixing simulation model that i s widely usable, i s also needed to develop new hypotheses in the study of ecological processes. 3. "MIXER", a new conceptual model describing earthworm a c t i v i t y in s o i l systems a. Introduction Based on l i t e r a t u r e data, a new conceptual model, "MIXER," is developed, describing earthworm a c t i v i t y in s o i l systems. This new model includes the moisture and temperature relations for L. rubellus, that were developed in Chapter III . The model i s a multi-species, non-specific model that can be used in a wide range of conditions. It is based on the morphological - functional c l a s s i f i c a t i o n as described by Bouche (1977), as well as on components from the conceptual model by Reynolds and Jordan (1975). In th i s model, the movement of mineral and organic matter in the s o i l p r o f i l e i s ce n t r a l . The model can e a s i l y be adapted for EARTHWORM SIMULATION MODELS / 105 long-term computer simulations. b. S o i l layers in the model Earthworms react to their environment; their a c t i v i t y , reproduction, survival and growth are based on the environmental conditions of which moisture and temperature may be the most important (Bouche, 1984; Sa t c h e l l , 1980; Lavelle, 1975; Reynolds and Jordan, 1975). With regard to earthworm a c t i v i t y in the s o i l system, the s o i l p r o f i l e may be separated into three layers: the organic matter or l i t t e r layer on top of the s o i l (I), the mineral s o i l layer in which most of the b i o l o g i c a l a c t i v i t y takes place, ( I I ) , and the subsoil ( I I I ) . Because earthworms may change the pedogenic horizons in the s o i l p r o f i l e through s o i l mixing (Langmaid, 1964), terminology from the Canadian System of S o i l C l a s s i f i c a t i o n (Canada S o i l Survey Committee, 1978) i s not used. c. Population dynamics The carrying capacity of a s i t e w i l l determine the population density and must therefore be included in the model. Carrying capacity i s determined by the a v a i l a b i l i t y of food, environmental variables such as moisture, temperature, type of vegetation, drainage and s o i l management. Other factors are bird predation on earthworms EARTHWORM SIMULATION MODELS / 106 (Cuendet, 1983; Satchell, 1981), predation by s o i l vertebrates and other s o i l animals (Lee, 1985; Macdonald, 1983) and competition between species (Abbot, 1980; Bouche, 1977). A Site Quality Index, similar to one used in Forestry-management, would be a useful tool to estimate maximum earthworm populations in a given s i t e . Once the rel a t i v e carrying capacity for each worm species i s established, the recovery of the population can be predicted after a major (man-made) disturbance ( r o t o t i l l i n g , ploughing and seeding, harvesting, slashburning, pesticide spraying) or after earthworm introduction or s o i l improvement (drainage e t c . ) . The growth curve f i r s t may exhibit exponential growth, but once food or predation becomes l i m i t i n g , the growth curve f l a t t e n s out, and the carrying capacity i s reached. After the growth phase, the seasonal e f f e c t s , such as temperature and drought stress, w i l l become obvious in the biomass. In a g r i c u l t u r a l situations, where s o i l management takes place during each cropping season, or every two to three years in longer rotations, the maximum possible biomass may not be reached. In permanent pastures, forested systems and possibly in lands under a l i m i t e d - t i l l a g e cropping practice, carrying capacity may be reached. The t o t a l population of each worm species may be divided EARTHWORM SIMULATION MODELS / 107 into proportions of young (small) worms, immature adults and mature ( c l i t e l l a t e d ) adults. This is necessary because the ingestion rate of the worms depends on their age (Hartenstein et a l . , 1981; P h i l l i p s o n and Bolton, 1976; Lavelle, 1975). The r e l a t i v e abundance of each age group varies according to season (Reynolds, 1976). d. Earthworm food U n t i l recently, the food of earthworms was described as s o i l and organic matter; microorganisms are now regarded as an important food source of earthworms, as was discussed in Chapter I I . The quality of the food influences the amount ingested (Lavelle , 1975; Appendix 3-B), therefore a system should be developed to rate the quality of food based on p a l a t a b i l i t y , moisture content, carbon to nitrogen r a t i o , etc. This Food Quality Index could be a r e f l e c t i o n of the decomposition rate of organic matter (Reynolds and Jordan, 1975) or of microbiological a c t i v i t y . If the worms digest microorganisms that inhabit the food material, cast material may also be an important part of the diet as reported by Bouche et a l . (1983). The consumption of casts may be dependent on the aging of faecal material, following n i t r i f i c a t i o n and microorganism blooms, 6 - 7 weeks afte r egestion in temperate regions (Bouche et a l . , in press). Bouche et a l . EARTHWORM SIMULATION MODELS / 108 (1983) calculated that the anecic Nicodrilus velox ingested 50 % i t s carbon needs from recycled casts. The other carbon would come from the l i t t e r layer (Bouche et a l . in press). Therefore organic matter should be divided into age classes, each worm species w i l l ingest the age class of i t s choice. Ingestion of s o i l by some species can be seen as a method for "making up" a deficiency in food when organic matter cannot supply the amount of nutrients needed. S o i l might be a r e l i a b l e but "low q u a l i t y " source of food, and i t i s required for grinding processes in the gizzard. The ingestion of carbon, as discussed in Chapter I I I , may be an important parameter, describing the energy needs of earthworms and needs to be studied as a basis of food-intake behavior. The consumption patterns of earthworms with d i f f e r e n t strategies w i l l vary. The following feeding patterns are a r b i t r a r i l y established. Epigeic species: 100 % from layer I (L); endogeic species: 100 % from layer II (Ah); and anecic species: 70 % from layer 1,10 % from the II (Ah) and 20 % from layer III (BC). EARTHWORM SIMULATION MODELS / 109 e. Cast production A worm population that consists of several species with dif f e r e n t strategies, w i l l be the most e f f e c t i v e in mixing s o i l and incorporating organic matter. Cast production and the s o i l layer in which casts are produced are species s p e c i f i c . The following pattern of cast production i s a r b i t r a r i l y established: epigeic species: 100 % in layer I ; surface casting endogeic species: 90 % in layer I, 10 % in the II; sub-surface casting endogeic species, 10 % in layer I, 90 % in II; and anecic species: 100 % in layer I. f. Flow-diagram for MIXER The flow-diagram of the proposed conceptual model i s shown in Figure 11. The worm compartment contains the population growth parameters and the species d i f f e r e n t i a t i o n , as well as basic ingestion information. Site Quality (climate, n u t r i t i o n , plant community, s o i l type, succession) d i r e c t l y influences the abundance of species at the s i t e . The food intake of each species or group of species, depends on the temperature and moisture fluctuations in the s o i l and on the quality of food available. The amount of food ingested depends on the age of the worms, immature worms ingest r e l a t i v e l y more material than c l i t e l l a t e d adults. In the s o i l compartment, the sources of food and the EARTHWORM SIMULATION MODELS / 110 plant abundance by species 0 -biomass by age gut conte nt Figure 11 . Flow diagram for the earthworm model MIXER. I = L i t t e r layer, II = Upper s o i l layer with most of the b i o l o g i c a l a c t i v i t y , III = Subsoil, CH = Choice of food, G = Population growth, E = Egestion rate, In = Ingestion rate, M = Moisture e f f e c t , OM = Organic Matter, SQ = Site Quality, T = Temperature e f f e c t . The squares indicate d i f f e r e n t age of mater i a l . EARTHWORM SIMULATION MODELS / 111 d i s t r i b u t i o n of cast are described; included is ingestion of l i t t e r , cast and s o i l from the d i f f e r e n t s o i l layers. Worms ingest s o i l and organic matter from d i f f e r e n t layers in the p r o f i l e , according to their strategy. As earthworms ingest plant material that i s s u f f i c i e n t l y decomposed to their taste, a cohort system for l i t t e r i s included in the model. Each new batch of plant l i t t e r added to the s o i l , i s followed over time, t i l l the required state of decomposition is reached. This cohort system for plant l i t t e r also enables the researcher to model competition between species with d i f f e r e n t preferences for l i t t e r . Cast is deposited in the s o i l system at a depth, s p e c i f i e d for each worm species or strategy. By using d i f f e r e n t c o e f f i c i e n t s for the intake of certain food materials, several ingestion patterns can be simulated. Assimilation and excretion of mucus and urine i s not included in the model although Bouche et a l . (in press) described the importance of earthworm excretions to plants. B . F-MIXER, A SIMULATION MODEL FOR SOIL MIXING BY EARTHWORMS 1. I n t r o d u c t i o n The conceptual model MIXER, which includes the results of the moisture and temperature experiments, and the results of the 1"C column experiment were used to create a model for the simulation of movement of mineral and organic matter EARTHWORM SIMULATION MODELS / 112 into the s o i l p r o f i l e . In the discussion of MIXER, emphasis was given to the d i s t r i b u t i o n of cast in the p r o f i l e and the intake of food. The column experiment, described in Chapter IV, resulted in information on the d i s t r i b u t i o n of cast in the p r o f i l e by L. rubellus and i t s choice of organic matter. The column experiment i t s e l f i s used as an integral part of the computer simulation version of MIXER. The objectives for creating the simulation model were a) to test the understanding and knowledge of s o i l mixing by earthworms, b) to extrapolate the data from the column experiment and c) to provide the ecosystem modeling framework, "FORCYTE," with a s o i l mixing component. FORCYTE was choosen as the model to carry MIXER for three reasons. F i r s t t h i s ecosystem model lacked a s o i l mixing component; secondly because FORCYTE i s a long-term trend analysis model and t h i r d l y because MIXER could be included with a minimal amount of programming. Modification of FORCYTE to f i t the requirements of an a g r i c u l t u r a l ecosystem model, which would simulate crop production and the ef f e c t s of crop management, w i l l require substantially less time and e f f o r t than building and programming a separate model. The EARTHWORM SIMULATION MODELS / 113 simulation version of MIXER was therefore written as a sub-routine for a forestry model. This s o i l mixing model i s suited to simulate earthworm a c t i v i t y in both a g r i c u l t u r a l and forested systems.. 2. A brief description of FORCYTE The model FORCYTE (FORest nutrient Cycling and Y i e l d Trend Evaluator) i s a very large and complex model that evolved over several years to become an ecological modeling framework. In this respect, i t can be used, after suitable c a l i b r a t i o n s , to model plant production in any ecosystem. FORCYTE i s not sensitive to seasonal fluctuations in environmental conditions, but w i l l analyse trends in production levels over several decades. The simulation for forested systems i s done for up to three crop rotations (240 years). The most recent version of FORCYTE, FORCYTE-11, is a hybrid stand - individual tree growth model, based on h i s t o r i c a l y i e l d data and ecological processes of either managed or unmanaged forests. The model examines, on an ecosystem-type basis, the change over time in community composition, the production and biomass of mosses, herbs, shrubs and trees, the nutrient budget and nutrient c i r c u l a t i o n for up to fi v e nutrients, the inventory and dynamics of organic matter, and EARTHWORM SIMULATION MODELS / 114 the economics of production management (Kimmins, 1986; Kimmins et a l . , 1981). The s o i l part of the "SETUP" program of FORCYTE consists of "SOILDATA," the input f i l e , "FORSOIL," the simulation part and several output f i l e s . These output f i l e s are used in "FORCASTER" to simulate management regimens. The results of FORCASTER and FORSOIL are graphed and tabulated. The simulation of the SETUP program to aquire data to run FORCASTER, i s based on the simulation of short-term experiments, that can e a s i l y be done in the laboratory or as f i e l d experiments. Examples are sorption and desorption experiments, decomposition t r i a l s , etc. To include s o i l mixing by s o i l invertebrates as a sub-routine in FORCYTE, the column experiment from Chapter IV was used as a short-term experiment in SETUP to generate empirical data on the r e d i s t r i b u t i o n of s o i l and organic matter. For each earthworm species or group of species with similar strategies, a column experiment i s needed to c a l i b r a t e the model ( i . e . , epigics, e t c . ) . 3. Description of the FORCYTE version of MIXER The FORCYTE version of MIXER (F-MIXER) has been developed from the conceptual model to f i t the requirements of FORCYTE. To obtain information on s o i l mixing by earthworms, EARTHWORM SIMULATION MODELS / 115 a column experiment i s simulated, using s o i l and organic matter as well as the average s o i l moisture and s o i l temperature of a s i t e . In each time step (one year), a sp e c i f i c amount of root and shoot l i t t e r i s added to the columns and the worm population i s kept constant. The a v a i l a b i l i t y of carbon to the worms w i l l check their consumption of s o i l and organic matter. A maximum ingestion rate may be specified for each worm type. Worms in F-MIXER proportionally ingest organic matter from each s o i l layer, mix i t with s o i l to form cast and deposit the cast proportionally in the layers. Fluctuations in moisture and temperature status of the s o i l are averaged and are kept constant. The organic matter used in the simulation i s of palatable q u a l i t y and i s aged to f i t the worms' taste. Cast produced in a s o i l layer i s c l a s s i f i e d as humus after each year (one time step) and may be recycled by earthworms. As cast i s r e c l a s s i f i e d as humus, nutrients in excess of those spe c i f i e d for humus are released. These nutrients are used in FORCASTER to af f e c t plant growth and influence crop production predictions. Appendix 6 shows a detailed description of the processes and data requirements for F-MIXER. EARTHWORM SIMULATION MODELS / 116 4 . Comparison of F-MIXER to MIXER The results from a simulation of crop production or earthworm mixing using an ecosystem model, may not r e f l e c t the actual future state of the simulated ecosystem. F-MIXER was designed to evaluate trends based on the available knowledge of the processes shaping ecosystems, not as a model to predict exact quantities of mineral s o i l and organic matter cycled or to f i t the results of a short-term exper iment. The conceptual model MIXER as presented before, i s di f f e r e n t in several ways from the presented F-MIXER. MIXER requires detailed information on the s o i l temperature and moisture regimens at a certain s i t e for a short-term evaluation. Under Lower Mainland, B r i t i s h Columbia, conditions, the influence of drought on the earthworm a c t i v i t y i s minimal, as can be seen from the data presented in Appendix 5. The yearly temperature fluctuation, however, was seen in the calculated monthly cast production. Assuming a fluctuating temperature has the same effect on the egestion rate as the the average temperature, calculated from these fluctuations, no detailed information on s o i l temperature i s needed for long-term simulations. The eff e c t s of climate on crop production are covered in FORCYTE by using y i e l d curves and EARTHWORM SIMULATION MODELS / 117 the concept of biogeoclimatic zones. The y i e l d curve includes the effects of climate and other s i t e - s p e c i f i c parameters on the growth of plants. No similar information is available for earthworms, and no data were found regarding the effects of flu c t u a t i n g temperatures on the egestion of s o i l by earthworms. Reinecke and K r i e l (1981) reported a lower cocoon production for Eisenia foetida in earthworms incubated at a fluct u a t i n g temperature with an average of 20 °C, compared to a constant temperature of 20 °C. Differences in cocoon production were not found for 10, 15> and 25 °C. The topic of the effect of temperatures, fluctuating or constant, on earthworm a c t i v i t y , needs more study. To include detailed information on worm age-classes and moisture and temperature e f f e c t s , a d i f f e r e n t type of simulation model should be developed. This model may not simulate the long-term trends in the s o i l system, but w i l l describe the short-term e f f e c t s of an earthworm population. Such a model would be a population or production based model, similar to ALLEZ-LES-VERS (Lavelle and Meyer, 1978) or WORM.FOR (Mitchell, 1983). In long-term simulations, the age structure of the earthworm population may not be needed, although the egestion rate for EARTHWORM SIMULATION MODELS / 118 the t o t a l population should be calibrated to include the higher egestion rates of young specimens. Population density changes, as influenced by available food, should be incorporated into the model. 5. Simulation The b i o l o g i c a l functions of F-MIXER were tested with the simulation of the transport of mineral s o i l and organic matter in an a g r i c u l t u r a l system. In t h i s system, a n o - t i l l a g e management concept was assumed. A series of crops was followed over a 45 year period. After each growing season, crop residue was l e f t on the surface. The p a l a t a b i l i t y of t h i s residue for earthworms was assumed to be similar to that of clover material, decomposition rates for clover were used: 60 % weight loss during the f i r s t year for shoot material (Uvarov,1982) and 20 % for roots. Earthworm biomass at several l e v e l s (0, 50, 100, 200 and 590 kg.ha" 1, dwt) were used to simulate the organic matter and mineral s o i l transport in the p r o f i l e . Enrichment and depletion of organic matter was followed in the l i t t e r layer (I) , the upper mineral s o i l layer with earthworm a c t i v i t y (II) and a sub s o i l layer ( I I I ) . An earthworm biomass of 590 kg.ha" 1 (dwt) could not be supplied with enough surface l i t t e r (Figure 12-A). This high EARTHWORM SIMULATION MODELS / 119 l e v e l (590) chosen for the column experiment to obtain s i g n i f i c a n t casting in a short time, may not be r e a l i s t i c under f i e l d conditions. A l l surface l i t t e r was ingested and redist r i b u t e d into the p r o f i l e . The exhaustion of surface l i t t e r was expected because the worms removed nearly a l l the shoot l i t t e r during the 30 days incubation in the column experiment. At a l e v e l of 200 kg.ha" 1 (dwt), the worms just removed a l l the l i t t e r that was supplied each year, and mixed i t into layer I I . The curves in Figure 12-B, describing the organic matter content of that layer, show an increase in organic matter over time. The curves for the two highest populations indicate similar l e v e l s of organic matter and show that there i s no further increase in organic matter possible, once the l i t t e r layer i s consumed. Because in the model, the worms indiscriminately ingested a mixture of root material, mineral s o i l and humus, contrary to the results of the column experiments described in Chapter IV, their population should have been limited by the maximum intake of organic matter. Earthworm biomass between 200 and 250 kg.ha"1 as found in the Lower Mainland (see Appendix 2-D) and in a n o - t i l l s i tuation in Georgia EARTHWORM SIMULATION MODELS / 120 (Parmeleet, personal communication), are more r e a l i s t i c . Mechanisms in the model to actually link population densities with available organic matter and to model pr e f e r e n t i a l feeding on root materials, are currently being developed. The curve for the highest worm density (590) in Figure 12-B is lower than that for 200 because of recycling of organic matter and the decomposition of these recycled materials. The percent organic matter in layer II without worms, decreased s l i g h t l y over time due to decomposition. Large amounts of s o i l were ingested by the worms. The s o i l intake of the 200 kg.ha - 1 worm population, ranged from 48 to 111 t.ha" 1 (Figure 12-C), well within the range of published figures for surface cast in temperate regions (4.5 - 90.2 t.ha' 1). The worms need to process a certain amount of organic matter to survive (46 mg.g-1.day"1 of carbon at 10 °C in a s o i l at f i e l d capacity). In the model the worms get t h i s amount by ingesting surface l i t t e r and s o i l : 43 % of the carbon needs from layer I, 56 % from II and 1 % from II I . These values were calculated from the organic matter intake in the column experiment. When the earthworms add organic matter to the pool in layer II, by ingesting surface f R~. Parmelee, Ph.D. Candidate, Institute of Ecology, University of Georgia, Athens, Georgia, USA. EARTHWORM SIMULATION MODELS / 121 t / h o 10 0 50 100 - -2 0 0 59 0 5 / y / / / / / / V 5 10 20 45 YEARS F i g u r e 12-A. R e s u l t s of the s i m u l a t i o n of F-MIXER, the accumulation of s u r f a c e l i t t e r ( t . h a ' 1 (dwt), as a f f e c t e d by earthworm p o p u l a t i o n s (kg.ha" 1, dwt). %c 1 c 5 0 100 20 0 59 0 5 £ 10 20 45 YEARS F i g u r e 12-B. R e s u l t s of the s i m u l a t i o n of F-MIXER, the organic matter content (%) i n l a y e r I I , as a f f e c t e d by earthworm p o p u l a t i o n s ( k g . h a - 1 , dwt). EARTHWORM SIMULATION MODELS / 122 50 2 0 0 1 0 0 5 9 0 t/ha 400 v. 200 100 _ 5 10 20 Y EARS 4 5 Figure 12-C. Results of F-MIXER, the ingestion of s o i l ( t . h a - 1 , dwt) by di f f e r e n t populations of earthworms (kg.ha"'1, dwt). EARTHWORM SIMULATION MODELS / 123 l i t t e r and egest i t in the mineral s o i l , the concentration increases and thus less of a soil-organic matter mix is needed from this layer to s a t i s f y the carbon requirements. The egestion of s o i l w i l l therefore decrease over time. By t h i s mechanism i t i s assumed that cast material i s recycled by earthworms. The movement of mineral s o i l from one layer to the other, through earthworm a c t i v i t y , caused depletion of s o i l in mineral s o i l layers or an excess of s o i l in the l i t t e r layer. This problem was addressed by developing a method to simulate moving soil-layer-boundaries. Starting from the lowest layer, the d e f i c i t in mineral s o i l was f i l l e d by transferring s o i l from the layer above. Surface cast i s included in the upper mineral layer. The transferred s o i l contains an amount of humus similar to that of the s o i l in the receiving layer. The humus content of the l i t t e r layer depended on the decomposition rate of the clover materials (with the last decomposition class transferred to humus) and on the accumulation of cast. The cast that was deposited on the surface of the s o i l by the earthworms, was transferred in the model to the mineral s o i l . As the transferred materials contained the humus content of the receiving mineral layer, EARTHWORM SIMULATION MODELS / 124 some humus accumulated in the l i t t e r layer. This humus was re-used by the worms feeding on the l i t t e r layer. 6 . Improvements needed in F-MIXER The worms did not achieve their maximum organic matter intake, because in the model they did not switch to another organic matter source when one i s depleted. Indiscriminant ingestion of s o i l with roots and humus may not be a good representation of worm feeding. A correction factor or a "hunting routine" should be included in the model. For L. rubellus the ingestion r a t i o between shoot l i t t e r and root l i t t e r i s known (4 to 1, see Chapter IV), but no data are available on the switching of food sources by earthworms. Organic matter may have been a l i m i t i n g factor to the highest worm population. A population model describing the eff e c t s of food shortage on the population l e v e l of the worms would be needed to change the worm population according to the carrying capacity of the f i e l d . The layer-boundary adjustment should be fine-tuned. The l i t t e r layer contained humus from cast after the mineral matter i s transferred to the receiving layer. Inclusion of more s o i l layers could give a better simulation of the EARTHWORM SIMULATION MODELS / 125 changes in the s o i l p r o f i l e . An organic matter gradient in the s o i l could then be simulated. This version of F-MIXER includes only one earthworm strategy. Competition sub-routines are available and the model i s set up to include more strategies. However, no data could be found in the l i t e r a t u r e to c a l i b r a t e competition between earthworms. Data should be co l l e c t e d , using the column technique, on the r e d i s t r i b u t i o n of cast in the s o i l p r o f i l e by other species. The results of the simulations represented in Figure 12, should be validated in f i e l d plots for which h i s t o r i c a l data on worm populations and s o i l management are avai l a b l e . V I . GENERAL DISCUSSION The egestion rate calculated from the column experiment i s 5.2 times higher than that calculated from the l i t t e r b a g technique (2.34 vs 0.45 g.g" 1.day" 1). Direct comparison of the two techniques for measuring egestion rates i s d i f f i c u l t because the food source was not the same. In the l i t t e r b a g technique, the food (clover hay) was coll e c t e d from the f i e l d . This material had been in the f i e l d for several months and consisted of partly decomposed clover stems. The material used in the column experiment was grown in the greenhouse and consisted of young clover leaves. Young leaves are expected to decompose faster than the clover hay, and may contain a d i f f e r e n t microbial population. Large differences in egestion rates suggest that the young clover leaves are less palatable than the clover hay used in the l i t t e r b a g technique. The p a l a t a b i l i t y of the young clover leaves may be les s , either because they contained undesirable substances such as phenols, or they contained only limited amounts of microorganisms. Hence the worms supplemented the clover leaves with large quantities of s o i l . The development of a food q u a l i t y index based on the microbial a c t i v i t y and d i v e r s i t y should be helpful in future studies on earthworm feeding. When the egestion of organic carbon was compared for the two 126 GENERAL DISCUSSION / 127 techniques, both techniques showed similar r e s u l t s : 37.1 ± 12.31 rng.g"1.day"1 in the l i t t e r b a g technique , and 46.1 mg.g"1.day"1 in the transport experiment. In preliminary experiments, d i f f e r e n t kinds of foods were offered to L. rubellus (5 °C, moist s o i l ) . The egestion rate ranged from 0.09 g.g^.day" 1 for clover leaves, to 0.65 for s o i l . The egested amount of carbon was 15.5 rng.g"'.day"1 in a l l cases (Appendix 3-B). When the egested amount of carbon was calculated for the 5 °C l i t t e r b a g t r i a l , the following values were found: 17.8 ± 6.4, 18.1 ± 5.3, 25.9 ± 5.9 and 12.7 ± 4.5 rng.g"1.day"1 for wet to dry s o i l environments respectively. These values are not di f f e r e n t from the one found in the food selection t r i a l . The findings from these three independent experiments lead to the conclusion that the amount of carbon egested by earthworms, compared to the t o t a l egestion rate, i s a superior parameter to measure the carbon needs and a c t i v i t y of earthworms. The egestion rate of mineral s o i l plus organic matter can be seen as the flow rate of a " c a r r i e r " through the gut of an earthworm. This flow is needed to s a t i s f y the nutritonal needs of the earthworm. Organic matter can be seen as a superior food source, probably because of the large population of microorganisms i t contains. When the organic matter ( l i t t e r ) i s not supplying enough nutrients, the worms GENERAL DISCUSSION / 128 ingest large quantities of s o i l , including humus, to make up any d e f i c i e n c i e s . The t o t a l intake of organic matter i s a constant factor, only dependent on s o i l temperature and s o i l moisture which both a f f e c t the metabolic a c t i v i t y of the worms. Standards for temperature and moisture content in ingestion or egestion experiments are needed to compare the results for di f f e r e n t species. Only a small amount of organic matter i s assimilated by earthworms (e.g. Uvarov, 1982). It is well established that some earthworm species feed on microorganisms (see section II-A-2). The quality of food may well lay in the quantities of microorganisms inhabiting ( s o i l ) organic matter. Tests specifying the le v e l of a c t i v i t y or quantities of microorganisms may be used to specify the qual i t y of worm food. Both the column experiment and the l i t t e r b a g experiment were conducted at constant temperatures. Results from these experiments may not represent f i e l d s i t u a t i o n s . Diurnal and seasonal temperature fluctuations may cause changes in worm physiology, as was suggested by Reinecke and K r i e l (1981). At a fluctuating temperature of 20 °C, Eisenia foetida produced fewer cocoons, but more hatchlings per cocoon than at a constant temperature. More research i s needed to GENERAL DISCUSSION / 129 c l a r i f y the exact e f f e c t of a fluctuating temperature on earthworm a c t i v i t y and physiology. Although the egestion rates d i f f e r e d between both techniques, the l i t t e r b a g and the column experiment gave similar results for the egestion rate of carbon. This means that both techniques were equally suitable for measuring the egestion rate of carbon. The l i t t e r b a g technique does not require a large incubator and can be c a r r i e d out in a small space. The column experiment has many advantages. It y i e l d s more cast per column to analyze and also provides the d i s t r i b u t i o n of casts related to depth. The columns can be used both in the laboratory and in the f i e l d where excessive wetness i s detrimental for the l i t t e r b a g technique due to smearing of casts and s o i l by the earthworms. Workers in Europe regard L. rubellus as a species with l i t t l e influence on the s o i l structure (Bouche, personal communication, Edwards, personal communication). This may be based on the c l a s s i f i c a t i o n (epigeics), the low abundance in mature systems and on the assumption that the worm i s a surface casting species. Small amounts of cast found on the surface, actually only 15 % of the t o t a l cast production, might therefore be seen as an indication of a low egestion rate. GENERAL DISCUSSION / 130 In Europe, L. rubellus i s often found in association with other earthworms. Being a pioneer species, the worm may not be able to compete for food and space with other species once the habitat becomessuitable for K-strategists. Bouche (1977) suggested that epigeics and anecics compete for food. Abbot (1981) concluded from his competition experiments in the laboratory, that M. dubius did not do well in cultures with either E. foetida or AlI ol obophora trapezoi des . He suggested a toxic interaction between the species or that the a v a i l a b i l i t y of certain digestive enzymes in the superior species, would cause the decline. In associations with other worms, L. rubellus may be forced to occupy the very top of the s o i l , not preferred by other species. The published information about the role and a c t i v i t y of L. rubellus may also lead to the hypothesis that t h i s species developed d i f f e r e n t ecotypes: in Europe a surface dweller, and in North America a sub-surface dweller. However, both in North America (this study) and in Europe, the worm was found in the lower layers of the s o i l (Persson and Lohm, 1977; Byzova, 1965). Byzova's observation that the respiration rates of L. rubellus were similar to those of mid-strata worms, would indicate that i t is not an epigeic species. GENERAL DISCUSSION / 131 L. rubellus was active in the mineral s o i l - l i t t e r interface and created a burrow system. The worm also produced casts both in the l i t t e r layer (15 %) and in the mineral s o i l (85 % ) . Therefore the hypothesis that the earthworm species only l i v e s in the l i t t e r layer and only casts in this layer, is rejected. A new strategy class i s hereby proposed to include L. rubellus: eurygeic species {evpv = wide), worms that l i v e in the l i t t e r - s o i l interface and feed both on mineral s o i l and surface l i t t e r . Cast production i s both in the l i t t e r layer and in the mineral s o i l . These worms may have morphological c h a r a c t e r i s t i c s of both epigeic and endogeic worms, and are good s o i l mixers. This strategy i s included in Figure 13. Bouche's c l a s s i f i c a t i o n (Bouche, 1977) i s based on morphological c h a r a c t e r i s t i c s , while the functional and sp a t i a l emphasis i s implied. The eurygeic class i s based on functional and sp a t i a l c h a r a c t e r i s t i c s and may contain worms with d i f f e r e n t morphological c h a r a c t e r i s t i c s . Bouche (1977) showed that several European lumbricids were not covered by the three main classes. Wood (1974) distinguished two megascolecid worms from a mountain s i t e in Au s t r a l i a , as top s o i l species, with c h a r a c t e r i s t i c s to f i t the eurygeic c l a s s . These two species, Cryptodrilus fasligalus and Megascolex celmisiae fed on l i t t e r and on s o i l and occupied the top 10 cm of the mineral s o i l . Lavelle (1979) grouped Dichogaster agi I i s and Millsonia I ami ol ana GENERAL DISCUSSION / 132 BC SOIL AND ORGANIC MATTER TURN OVER ORGAN ICS USED AS FOOD F i g u r e 13. Earthworm s t r a t e g i e s as p e r c e i v e d from Bouche (1977) w i t h a new c l a s s added. GENERAL DISCUSSION / 133 from Ivory Coast in an intermediate class between epigeic and endogeic. These non-lumbricid species would also f i t in the eurygeic class, as would several t o p s o i l species from Lee's system (Lee, 1959, ci t e d by Lee, 1985). Non-native North American lumbricids that may be included in the eurygeic class are Aporrectodea caliginosa, Lumbricus festivus and L. castaneus. Adding the eurygeic class to Bouche's system, would make i t a strategy c l a s s i f i c a t i o n that is widely usable. The f i r s t approximation of a simulation model to test the understanding of s o i l mixing by earthworms revealed many gaps in our knowledge on earthworm ecology. From the simulation of s o i l mixing and the integration of MIXER in FORCYTE, i t became apparent that several topics require further research. Earthworm feeding, including a "hunting routine", and the characterization of the food need attention. The question of i n t e s t i n a l f l o r a versus the rapid development of microorganisms in the gut and faeces need to be resolved, and the question of the effects of fluc t u a t i n g temperatures on earthworm behavior should be explored. To run the simulation model for more than one species, basic data on the feeding and casting patterns are needed for a l l species (or groups of species) involved, as well as information on inter-species competition, establishment of GENERAL DISCUSSION / 134 qual i t y determinations for earthworm food and population dynamics as related to d i f f e r e n t s i t e s . V I I . SUMMARY AND CONCLUSIONS The objectives of the presented research were to describe the ecological strategy of Lumbricus rubellus Hoffmeister, in re l a t i o n to s o i l moisture, temperature and drought stress; to describe the transport of organic matter into the mineral s o i l ; and to develop a model to simulate t h i s transport. The following approaches were used: a l i t t e r b a g technique to study the eff e c t s of s o i l moisture and s o i l temperature on the egestion rate of L. rubellus and a column transport experiment in which 1 t t C l a b e l l e d clover material was offered as worm food, to study the food choice and the a b i l i t y of the worm to transport s o i l and organic matter. The l i t t e r b a g technique was adapted to be used with earthworms; a soil-moisture-buffer system was added by incubating the l i t t e r b a g s in a basin with s o i l of a predetermined moisture content. The column transport method was newly developed. S o i l columns were e q u i l l i b r a t e d on a porous plate and earthworms were fed with radio-labelled clover shoots or roots. The egestion rate was calculated and the r a t i o of shoots to roots in the earthworm diet was determined. This method appeared to be well suited to quantify 135 SUMMARY AND CONCLUSIONS / 136 egestion rates of earthworms. Results from the l i t t e r b a g experiment showed that both moisture and temperature affected the t o t a l egestion and ingestion rate of L. rubellus. The temperature effect was v i s i b l e under 'wet' conditions, the moisture effect was pronounced under 'dry' conditions. The earthworm reduced the intake of mineral s o i l when under drought stress to reduce water loss and ingested r e l a t i v e l y more organic matter. The faecal water content was always above the l i q u i d l i m i t of the s o i l and the faecal carbon content was above that of mineral s o i l . The body water content was related to the s o i l water po t e n t i a l . A maximum body water content was found at a s o i l water potential of -15 m of water. Results of the column experiment showed that L. rubellus produced 15 % of i t s cast on the surface, 46 % in the 0-5 cm layer, 22 % in the 5-10 cm layer and 16 % in the 10-15 cm layer of the s o i l . The worm p r e f e r e n t i a l l y ingested 78-82 % of the supplied organic matter as leaf l i t t e r and 18-22 % as root l i t t e r . The carbon content of the recovered cast was not s i g n i f i c a n t l y d i f f e r e n t in each of the s o i l layers; 1*C label originating from both surface l i t t e r and root SUMMARY AND CONCLUSIONS / 137 l i t t e r was recovered in cast throughout the p r o f i l e . These findings indicate that organic matter originating from the l i t t e r layer was mixed into the p r o f i l e by L. rubellus. 8. The two techniques for measuring earthworm a c t i v i t y , showed very d i f f e r e n t egestion rates for L. rubellus, 0 .45 g.g-'.day"1 for the l i t t e r b a g techinique and 2.34 g.g- 1.day 1 for the column experiment. The egestion rates of carbon, however, was similar (37.1 ± 12.31 vs 46.13 rng.g"1.day1, 10 °C). When the egestion of carbon (5 °C) was compared with that found in a preliminary food t r i a l , both tests showed a similar egestion of carbon (15.5 rng.g"1.day"1), similar to that found in the l i t t e r b a g technique (5 °C). 9. It was concluded that the ingestion of carbon r e f l e c t s the energy use of the earthworms and i s a more suitable parameter to measure earthworm a c t i v i t y than the t o t a l egestion rate. 10. Based on the presented evidence, the earthworm L. rubellus cannot be c l a s s i f i e d as an epigeic species. A new ecological strategy class was introduced: eurygeic worms, l i v i n g in the l i t t e r - s o i l interface. Several other non-native North American lumbricids and also some Australian and African megascolecid species can SUMMARY AND CONCLUSIONS / 138 currently be included in this c l a s s . 11. A conceptual model for mixing of s o i l and organic matter by earthworms was developed from the l i t e r a t u r e . This model is a multi-species, non s p e c i f i c model, contrary to the single-species, s p e c i f i c models presented in the l i t e r a t u r e . Data from the present column experiment were used to adapt the model as a sub-routine for FORCYTE, an existing model for ecosystem management simulations. Earthworms caused a decrease in the accumulation rate of surface l i t t e r and and increase in the organic matter content of the s o i l . Based on th i s simulation, several areas in earthworm ecology were i d e n t i f i e d that are in need of further research. BIBLIOGRAPHY Abbot , I . , 1980. Do earthworms compete for food? S o i l B i o l . Biochem. 1 2 : 523-530. 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E f f e c t s of f i v e s p e c i e s of earthworms on some s o i l p r o p e r t i e s . J . A p p l i e d . Ecology 20: 865-872. S p r i n g e t t , J.A. and J.K. Syers, 1984. E f f e c t of pH and cal c i u m content of s o i l on earthworm c a s t p r o d u c t i o n i n the l a b o r a t o r y . S o i l B i o l . Biochem. 16: 185-189. S t e e l , R.G.D., and J.H. T o r r i e , i960. P r i n c i p l e s and procedures of s t a t i s t i c s . McGraw-Hill, New York. 418 pp. Stephenson, W., 1945. C o n c e n t r a t i o n r e g u l a t i o n and volume c o n t r o l i n Lumbricus terrestris L. Nature 155: 635. Stout, J.D., 1983. Organic matter turn-over by earthworms. In: Earthworm Ecology. J.E. S a t c h e l l , e d i t o r . Chapman and H a l l , London. 35-48. Stout, J.D. and K.M. Goh, 1980. The use of radiocarbon to measure the e f f e c t s of earthworms on s o i l development. Radiocarbon 22: 892-896. Syers, J.K., A.N. Sharpley and D.R. Keeney, 1979. C y c l i n g of n i t r o g e n by s u r f a c e c a s t i n g earthworms i n a pasture ecosystem. S o i l B i o l . Biochem. 11: 181-185 Uvarov, A., 1982. Decomposition of c l o v e r green matter i n an a r a b l e s o i l i n the Moskow r e g i o n . P e d o b i o l o g i a 24: 9-21. Van Rhee, J.A.-, 1963. Earthworm a c t i v i t i e s and the breakdown of organic matter i n a g r i c u l t u r a l s o i l s . In: S o i l Organisms. J . Doeksen and J . van der D r i f t , e d i t o r s . North H o l l a n d P u b l . , Amsterdam. 55-59. Vimmerstedt J.P. and T.H. Finney, 1973. Impact of earthworm / 152 i n t r o d u c t i o n on l i t t e r b u r i a l and n u t r i e n t d i s t r i b u t i o n in Ohio s t r i p - m i n e s p o i l banks. S o i l S c i . Soc. Am. Proc. 37: 388-391. Wallwork, J.A., 1970. Ecology of s o i l animals. McGraw-Hill, New York. 282 pp. Watanabe, H., and S. Ruaysoongnern, 1984. Cast p r o d u c t i o n by megascolecid earthworm Phereiima spp i n Northeastern T h a i l a n d . P e d o b i o l o g i a 26: 37-44. Wolf, A.V., 1940. Paths of water exchange in the earthworm. P h y s i o l o g i c a l Zoology, 13: 294-308. Wood, M. A., 1974. The d i s t r i b u t i o n of earthworms (megascolicidae) i n r e l a t i o n to s o i l s , v e g e t a t i o n and a l t i t u d e on s l o p e s of Mt. Kosciusko, A u s t r a l i a . J . Anim. E c o l . 43: 87-106. Zajonc, I., 1982. Communities of earthworms (Lumbricidae: O l i g o c h a e t a ) i n meadows in the S l o v a k i a n C a r p a t h i a n s . P e d o b i o l o g i a 18: 341-349. Z i c s i , A., 1978. Feeding requirements of some l u m b r i c i d s p e c i e s and t h e i r s i g n i f i c a n c e i n the ecosystem i n v e s t i g a t i o n s i n Hungary. P e d o b i o l o g i a 18: 341-349. Zimmermann, P., 1973. Zur neuroanalen R e g u l a t i o n des Wasserhaushalts bei Lumbricus t e r r e s t r i s L. Z. Z e l l f o r s c h u n g 145: 103-118. APPENDIX 1. DIAGNOSIS OF LUMBRICUS RUBELLUS LUMBRICUS RUBELLUS HOFFMEISTER, 1843. (Red Worm, Red Marsh Worm) Diagnosi s length 25 - 150 mm diameter 4 - 6 mm adult weight 52 -160 mg (dwt) segments 9 5 - 1 2 0 tanylobic prostomium f i r s t dorsal pore 7/8 c l i t e l l u m xxvi, xxvii - xxxi, xxxii tuberculata pubertatis on x x x v i i i - xxxxi setae closely paired, aa:ab:bc:cd:dd = 5:1:5:5/6:19 dd=1/2 u and ab of x often on a pale genital tumescences male pores inconspicuous, without glandular p a p i l l a e on xv seminal v e s i c l e s , 3 pairs in 9, 11 and 12 + 13 spermathecae with short ducts opening in 9/10 and 10/11 colour: ruddy brown or red v i o l e t and iridescent d o r s a l l y , pale yellow v e n t r a l l y body c y l i n d r i c a l and sometimes dorso-ventrally flattened p o s t e r i o r l y (Reynolds et a l . , 1974) 1 53 APPENDIX 2-A. DIAGRAM OF THE FUMIGATION SET-UP Appendix 2-A. Diagram of fumigation set-up. 1 = clover plants; 2 = container with nutrient solution; 3 = plexiglass container; 4 = removable l i d ; 5 = seal; 6 = v e n t i l a t o r ; 7 = membrane injec t i o n port; 8 = mixing vessel; 9 = heat absorbing water bath; 10 = l i g h t source. 1 54 APPENDIX 2-B. DIAGRAM OF THE SOIL COLUMN SET-UP 13 Appendix 2-B. Diagram of the s o i l column set-up. 1 = l i d (500 ml c o n t a i n e r ) ; 2 = s e a l ; 3 = c l o v e r ( s h o o t s ) ; 4 = c l o v e r ( r o o t s ) ; 5 = s o i l ; 6 = s e a l ; 7 = porous p l a t e (sand); 8 = ABS column and l i d ; 9 overflow u n i t ; 10 = c i r c u l a t i o n pump; 11 = water r e c e r v o i r ; 12 = gas wash v e s s e l ; 13 = po l y p r o p y l e n e t e n t ; 14 = p l e x i g l a s s c o n t a i n e r ; 15 = mesh p l u g . 155 APPENDIX 2 - C . SOIL MOISTURE AND SOIL TEMPERATURE ON WESTHAM ISLAND 1 ID Appendix 2-C, Figure A. S o i l moisture and s o i l temperature on Westham Island, measured during 1983 and 1984. S o i l temperature at 5 cm depth, s o i l moisture 0-5 cm: •-• ; 5-10 cm: o-o . 156 APPENDIX 2 -D . THE POPULATION OF L. RUBELLUS ON WESTHAM ISLAND. Appendix 2-D, Figure A. The population of L. r u b e l l u s in two f i e l d s on Westham Island. • • = Barley-clover after potatoes; o o = Clover-peas-clover; 1981 - Carter (unpubl.), 1982 - Carter and Timmenga (unpubl.), 1983 - Carter and Bandoni (unpubl.). 157 APPENDIX 2-E. RETENTION CURVE OF CRESCENT SERIES SOIL. Appendix 2-E, F i g u r e A. The water r e t e n t i o n c u r v e f o r the C r e s c e n t S e r i e s s o i l from Westham I s l a n d , B r i t i s h C o l u m b i a . • = Supplementary d a t a from De V r i e s ( P e r s . Comm.) 158 APPENDIX 3 - A . LENGTH OF LITTERBAG EXPERIMENTS. The l i t t e r b a g technique was used in several preliminary experiments. The length of the incubation was derived from the following table. Appendix 3-A, Table A. The egestion rate of Lumbricus r u b e l l u s incubated at d i f f e r e n t periods (10° C., M = 0.39 kg.kg" 1, n = 6) Time (days) Egestion rate (±SD) g.g 1.day 3.5 0.67 (0.314) 7.0 0.50 (0.134) 10.0 0.45 (0.091) 14.0 0.51 (0.146) 159 APPENDIX 3-B. FOOD CHOICES OF L. RUBELLUS Using the l i t t e r b a g technique, L. r u b e l l u s was fed on d i f f e r e n t food materials: partly decomposed clover leaves, partly decomposed clover leaves plus s o i l , partly decomposed clover hay plus s o i l , partly decomposed barley straw plus s o i l and s o i l . The s o i l was co l l e c t e d from the Westham Island f i e l d , as described e a r l i e r . The worm showed remarkable differences in egestion rates (5 ° C ) , the lowest rate was found for clover leaves, while s o i l r e f l e c t e d the highest rate. The t o t a l amount of carbon egested remained constant in a l l cases and there was a s i g n i f i c a n t inverse linear c o r r e l a t i o n between the amount of carbon egested and the egestion rate (r = - 0.944, See Figure Appendix 3-B). The calculated egestion of carbon was 15.5 mg.g"1 .day""1 for a l l food sources. The carbon content was measured as Leco carbon. Barley straw was not ingested by the worms, the p a r t i c l e s were not shredded. However, i t was noted that fungal colonies were removed from the straw and ingested. 160 / 1 6 1 Appendix 3-B, Figure A. The c o r r e l a t i o n between the egestion rate (g.g~ 1.day" 1) of worms fed on d i f f e r e n t foods and the faeces carbon content (%) for L. r u b e l l u s (5 °C). APPENDIX 3 - C . SIZE AND AGE OF L. RUBELLUS, AS RELATED TO THE EGESTION RATE. Mature c l i t e l l a t e d adults and immature adults were incubated for 7 days at 10° C. and - 12.5 m of water. Immature worms showed a higher egestion rate than mature worms' (Figure A). There was a s i g n i f i c a n t negative c o r r e l a t i o n between weight and egestion rate when the population included both mature and immature worms (r = 0.737, Figure B). 162 / 163 o i i 1 0 - »0 - 20 W A T E R P O T E N T I A L (M OF WATER) Appendix 3-C, Figure A. The egestion rate (g.g" 1.day" 1) of mature, c l i t e l l a t e d and immature adults of L. r u b e l l u s , fed on clover leaves plus s o i l (10° C). / 164 o » • • 0 50 100 150 ZOO 25 0 WORM WEIGHT (MG) Appendix 3-C, Figure B. The egestion rate (g.g - 1.day" 1) of L. r u b e l l u s as related to the worm weight (mg), for worms fed on clover leaves and s o i l (10 °C,) APPENDIX 4. THE EGESTION RATE OF A. CHLOROTICA. The earthworm AlIolobophora c h l o r o t i c a Savigny, 1826,. was an abundant species in the Westham Island clover f i e l d . A t h i r d less abundant species, was Eiseni el I a t e t r a e d r a Savigny, 1826. A. c h l o r o t i c a was tested in the l i t t e r b a g technique on several occasions. In contrast to L. r u b e l l u s , t h i s species burrowed through the s o i l clods in the l i t t e r b a g and consumed only limited amounts of the offered organic matter. The egestion rate was three times as high as that of L. rubellus (1.09 (0.764) g.g-'.day"1 (SD), 6.04 % C in the cast, 15 °C) under similar conditions, and the carbon content of the faeces (2.33 % C in the cast, 15 °C) was s l i g h t l y higher than that of the offered s o i l . 165 / 166 Appendix 4, Table A. The egestion rate of A. c h l o r o t i c a under di f f e r e n t environmental conditions. Temp Season Moisture Specimens Egestion °C m of w n (SD) 5 f a l l - 3 10 spring - 3 10 f a l l - 3 15 spring - 10 9 1.74 (0.404) 10 1.36 (0.603) 11 2.93 (1 . 160) 5 3.03 (0.764) APPENDIX 5 . TURN-OVER OF SOIL AND ORGANIC MATTER CALCULATED FROM THE LITTERBAG TECHNIQUE The turnover of s o i l and organic matter by L. r u b e l l l u s was calculated using the method outlined by Lavelle (1975). For each monthly period, the average s o i l temperature (5 cm depth) and s o i l moisture content (0-5 cm) was calculated (using data from Appendix 2-C) and the ingestion rate for organic matter and s o i l were interpolated using data from Figures 3 and 4. The population biomass figures, co l l e c t e d by Carter (unpublished), and Carter and Timmenga (unpublished) were used to calculate the t o t a l amount of s o i l and organic matter ingested by earthworms (population figures from Appendix 2-D). A population of adult L. r u b e l l u s in an a g r i c u l t u r a l f i e l d and recovering from a major disturbance (harvesting of a potato crop followed by s o i l c u l t i v a t i o n ) , ingested 1.9 tonnes.ha - 1 organic matter and 14.6 tonnes.ha - 1 mineral s o i l during a year. This estimate does not account for the exhaustion of organic matter and i t s e f f e c t s on the worm population. The population figures used, r e f l e c t the recovery of a population after a major disturbance, the population may not have reached i t s l i m i t s . Total ingestion rate in each month was c l e a r l y influenced by s o i l temperatures, while drought e f f e c t s were hardly v i s i b l e . 167 / 168 Appendix 5, Figure A. The turn-over of s o i l ( x ) and organic matter (•) by a population of adult L. r u b e l l u s (August 1983 - September 1984; 1981-1982 population data were used,{o}.). / 169 Under Westham Island conditions the effect on ingestion rate caused by a drop in s o i l moisture content is not strong enough to s i g n i f i c a n t l y influence the effect of temperature. Earthworms were active throughout the year; in wet winter months, when temperatures were only a few degrees above freezing, worms were active in the l i t t e r layer. During summer when s o i l temperatures reached 20° C, s o i l water tension became the l i m i t i n g factor for earthworms and L . rubellus moved down into the p r o f i l e ; worms were found in cracks and old root channels at depth of 15 to 20 cm. Their burrows had accumulations of organic matter and were coated with mucus. In south coastal B r i t i s h Columbia, the s o i l temperature rarely exeeds the optimum temperature range (15 - 18 °C) of th i s species. APPENDIX 6. DETAILED DESCRIPTION OF F-MIXER. FORCYTE VERSION 11:11 SOILSDATA Input data not s i t e s p e c i f i c . Section 1.5: specify s o i l layers (n, layer 1 is L i t t e r l a y e r ) specify a v a i l a b i l i t y of earthworms (yes/no) specify humus type from cast ingestion pattern of worm (% total ingestion, weight basis per layer) egestion pattern of worm (% total egestion, weight basis per layer) specify youngest age of l i t t e r used by worms Input data s i t e s p e c i f i c . Section 2.10: mass of mineral s o i l in layers (no stones) mass of humus in layers maximum worm biomass on the s i t e (kg/ha) population recovery (time steps or population dynamics) ingestion rate of organic matter (kg/kg.time step) to sustain population maximum throughput.of cast (kg/kg.time step) a s s i m i l a t i o n rate for humus (% weight loss) a s s i m i l a t i o n rate for l i t t e r (% weight loss) Data input for simulation of column experiment: biomass of worms (kg/ha) time steps for duration of experiment (nn) population dynamics switch (yes/no) *to allow to hold population constant or to fluctuate population with a v a i l a b l e organic matter. added l i t t e r to surface (kg/ha) decomposition type of this l i t t e r added fi n e root biomass to s o i l (kg/ha) decomposition type of f i n e roots d i s t r i b u t i o n of root l i t t e r in s p e c i f i e d layers FORSOIL SIMULATION OF COLUMN EXPERIMENT Normalisation of d i s t r i b u t i o n s (ingestion, egestion, root l i t t e r d i s t r i b u t i o n f o l i a g e l i t t e r on top of s o i l (layer 1) No worms, (1.5) sub-program is deleted from program run I n i t i a l i z e various output parameters (cast per layer, decomp. per layer, humus per layer, organic concentration 170 per layer I n i t i a l i z e nutrient parameters (content in layer:, in humus, in l i t t e r ; from 2.5, 2.10) I n i t i a l i z e tabular output headings: time, wormmass, littermass ingested, humus mass Ingested, s o i l mass Ingested, cast mass egested for n layers (current) l i t t e r mass for n layers (total d i s t r i b u t i o n ) humus mass for n layers (total d i s t r i b u t i o n ) s o i l mass for n layers (current timestep only) % OM for n layers RUNNING OF F-MIXER (Loop over time) Zero values that describe the sums of added humus, added l i t t e r , net release nutrients Redistribute s o i l and organic matter to r e - a l i g n s o i l layers • I n i t i a l amount of mineral s o i l In each layer 1s saved (from 2.10). The mixing by worms 1n the previous time step has upset the layer content. To correct the layer content the following c a l c u l a t i o n Is done for each time step exept the f i r s t one. To start with bottom layer, compare avai l a b l e mineral s o i l with the I n i t i a l , a net difference 1s transported to/from the layer above m order to regain the i n i t i a l s o i l mass. Humus 1s transported with the mineral s o i l according to the concentration of the layer where the s o i l comes from. Decompose e x i s t i n g l i t t e r (rates from SOILDATA 2.8) •Method of decomposition: second last age class compared with last in array, nutrients are released according to change in mass and nutrient content ( l i t t e r 2.7; humus -2.5, pattern of change 1.4) net release of each nutrient is summed Add l i t t e r to each s o i l layer (2.10) Calculate expected amount OM Ingested •This amount is calculated from the worm biomass and the need of OM to sustain the population. A l l o c a t e ingestion per layer A l l o c a t e potential OM ingested per layer ( s o i l + humus + l i t t e r ) •The potential OM ingested is calculated from the humus and l i t t e r content of the s o i l layers. It 1s the summation of the proportion of OM for each layer. / 172 Soil 1s Ingested, rocks are exluded from the total mass of s o i l . Unused decomposables included in the s o i l , but separately s p e c i f i e d , are not ingested. Compare potential with expected amount of ingested OM Worms ingest the potential amount of mix Apply a s s i m i l a t i o n rates: ingested material is reduced Redistribute the cast into the s o i l layers according to given proportions (1.5) Soil humus decomposition: nutrient release Transfer last decomposition class of l i t t e r to humus Transfer cast to humus, humus and s o i l are d i s t r i b u t e d as s p e c i f i e d for each layer, a l l decomposition types go to s p e c i f i e d humus cl a s s , release of nutrients Calculate OM concentration for each s o i l layer Calculate cast for each layer Worm population switch •Population 1s constant, each time step, the same amount of biomass 1s added to the column, or, when switch is in use, the population w i l l Increase or decrease depending on the amount OM ava i l a b l e . The maximum population is taken into account. Create output 1ine Publications Timmenga, H.J., 1983 Effects of soil moisture, soi l temperature and food quality on the turn-over rate of organic and mineral matter by the earthworm Lumbricus  rube!lus in a clover system. AIC/CSSS Meeting, Truro, Nova Scotia, 1983. Abstract published in Proceedings. Timmenga, H.J., and D. Pederson, 1984. Earthworm population densities in a drained and an undrained f ie ld of the Boundary Bay Water Management Research Project. Internal Report for the Brit ish Columbia Ministry of Agriculture and Fisheries, Cloverdale, B.C. 7 pp. Timmenga, H.J., 1984. The possible effects of a r t i f i c i a l acid rain on so i l s , crops and soil biota. Year Report 1982-83, Faculty of Agriculture, UBC, Vancouver. 20-21. Timmenga, H.J., 1984. The water relations of the earthworm Lumbricus rubellus io a clay loam s o i l . CSSS Meeting, Banfff, Alberta, 1984. Abstract published in Proceedings. Carter, A., T.F. Guthrie, E.A. Kenney, H.J. Timmenga, 1984. Heavy metals in earthworms in non-contaminated and contaminated soils from ;near Vancouver. In: Earthworm Ecology, J .E . Satchell, editor. Chapman and Hal l , London. 267-274. fe Schreier, H. and H.J. Timmenga., 1986. Earthworm response to asbestos rich serpentinitic sediments. Soil B io l . Biochem. 18: 85-89. Timmenga, H.J., 1986. The f e r t i l i z e r and chemical products market of the Nursery and Greenhouse Industries in the south Coastal Region of Brit ish Columbia. Consultants Report for Coast Agri Fer t i l i ze r Ltd., Abbotsford, B.C. 14 pp. Timmenga, H.J., 198/. Soil degradation in Brit ish Columbia. Butter fat, Oairyland Foods, Burnaby. (in press). Timmenga, H.J., 1987. Soil conservation in Brit ish Columbia. Butter fat, Dairyland Foods, Burnaby, (in press). 

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