@prefix vivo: . @prefix edm: . @prefix ns0: . @prefix dcterms: . @prefix skos: . vivo:departmentOrSchool "Forestry, Faculty of"@en ; edm:dataProvider "DSpace"@en ; ns0:degreeCampus "UBCV"@en ; dcterms:creator "Francia, Faustino C."@en ; dcterms:issued "2011-10-28T19:05:31Z"@en, "1965"@en ; vivo:relatedDegree "Doctor of Philosophy - PhD"@en ; ns0:degreeGrantor "University of British Columbia"@en ; dcterms:description """Tim factors and mechanisms which consistently deliver newly overwintered adults of the ambrosia beetle Trypodendron lineatum (Olivier) to specific host trees under specific conditions have been considered. Studies and analyses of the behavioural patterns of the beetle in respect to light, host tree factors, and factors of the environment were made in recognition of the fact that preliminary research results demonstrated the predominant role that reactions to light played!, in the behaviour of the beetle. The results of the studies showed that the beetles, before flight, were strongly phototacstic at temperatures in the range of 5° to 56°C. Positive photic response was inhibited in an increasing percentage of individuals at temperatures above 56°. At 38°C, positive response to light by the beetles ceased to exists. The non-flown beetles reacted quickly to angular deviations of alignment in respect to a light source and turned, with almost equal readiness toward the light regardless of its angle. The beetles' responses to light may be classified as follows: (1) not inhibited, the beetles immediately, move toward the source of light; (2) initially inhibited positive response; (3) inhibited positive response, the beetles may or may not initially inhibited but the general movement toward She light source is not direct; (4) completely inhibited response, the beetles move with no apparent response to light. The photopositive response was found to mask: the other potential capabilities of the non-flown beetles. Exclusion of the photic stimulus from flight-inexperienced beetles resulted in response to host odour in an odour field. Flight experience was found to modify partially the simple photic reaction of some individuals in the absence of host odour, but the majority of the beetles' responses remained unchanged. The behaviour of Trypodendron is not strictly stereotyped in the sense that it follows a definite pattern. While flight may be normal as a conditioning mechanism preparatory to alighting and host finding, response to an "attractive” odour source and subsequent boring behaviour may be exhibited, in the absence of previous flight experience, under certain conditions."""@en ; edm:aggregatedCHO "https://circle.library.ubc.ca/rest/handle/2429/38385?expand=metadata"@en ; skos:note "The University of B r i t i s h Columbia FACULTY OF GRADUATE STUDIES PROGRAMME OF THE FINAL ORAL EXAMINATION FOR THE DEGREE OF DOCTOR OF PHILOSOPHY of FAUSTINO CAJUCOM FRANCIA B.SoF., Un i v e r s i t y of the Phi l i p p i n e s , 1952 M,S . j State U n i v e r s i t y of New York, College of Forestry at Syracuse University, 1957 FRIDAY, JULY 16, 1965, AT 10:30 A.M. IN ROOM 3332, BIOLOGICAL SCIENCE BUILDING COMMITTEE IN CHARGE Chairman: J . A, F. Gardner J„ J„ R. Campbell K. Graham W. S. Hoar 0. S z i k l a i J . W. Wilson D. J . Wort External Examiner: A. J . Thorsteinson Department of Entomology University of Winnipeg Winnipeg, Manitoba STUDIES OF SOME.ASPECTS OF BEHAVIOUR IN THE AMBROSIA BEETLE TRYPODENDRON LINEATUM (OLIVIER) ABSTRACT Behaviour patterns of the ambrosia beetle Tr^ E S ^ S S \" dron lineatum. (Olivier) were studied -in r e l a t i o n to various influences of i l l u m i n a t i o n , temperature, gravity, atmos-pheric pressure and host.factors. The beetles were i n the overwintered, reproductively mature state i n which they are prepared for t h e i r normal functions of emergence, f l i g h t d i s p e r s a l , host-discovery and attack. Experiments were confined to the study of the beetles i n pedestrian s i t u a t i o n s , Their responses were studied both before and a f t e r being given f l i g h t experience. Before f l i g h t the majority of i n d i v i d u a l s are strongly photopositive at temperatures i n the range of 5 to 36°C, the optimum being between 20° and 26°C. They become progressively photonegative as temperatures r i s e above 26°C, u n t i l a l l i n d i v i d u a l s shun the l i g h t at 40 C. The e f f e c t i s r e v e r s i b l e as temperatures return to optimum. A f t e r a period of f l i g h t the photopositive reaction i s weakened, n u l l i f i e d , or replaced by a nega-t i v e reaction i n some, but not a l l i n d i v i d u a l s , A change i n threshold of response to l i g h t becomes most c l e a r l y manifest i n the presence of a t t r a c t i v e host odour. When a photopositive reaction occurs, i t i s re-tained regardless of the d i r e c t i o n of the earth's g r a v i t a -t i o n a l f i e l d i n r e l a t i o n to the l i g h t source. The photo-p o s i t i v e response i s not a l t e r e d by two atmospheres pressure. A response to host odour becomes manifest under three d i f f e r e n t s i t u a t i o n s - (1) Female beetles bore in t o wood which i s i t s e l f a t t r a c t i v e or i s made so by a p p l i c a t i o n of a t t r a c t i v e wood d i s t i l l a t e . This reaction i s displayed by flight-experienced beetles i n illuminated or non-illuminated s i t u a t i o n s , or by non-flown beetles i n darkness. (2) Males or females d i s -play a c e n t r i p e t a l response i n a f i e l d of odour a r i s i n g through a substratum. They trace a meandering path for a prolonged period, and repeatedly r e t r i e v e t h e i r general p o s i t i o n when they transgress beyond the boundary of the odour f i e l d . This reaction i s displayed under i l l u m i a t e d conditions by flight-experienced beetles. ( 3 ) Males or females respond anemotactically to a i r streams carrying a t t r a c t i v e host odour. This reaction i s displayed under illuminated conditions by flight-experienced beetles. The e f f e c t of f l i g h t experience on photic re-sponses i s not all-or-none s but apparently involves a r i s e of threshold to l i g h t i n the presence of a t t r a c t i v e host odour, or a lowering of threshold to host: odour i n the presence of l i g h t . The non-response of the beetles to green wood from a l i v i n g tree, and the continued response to mixtures of green and a t t r a c t i v e wood indicate that neither attractants nor repe l l e n t s are present i n the l i v i n g tree. It i s i n f e r r e d that s u s c e p t i b i l i t y of dying trees to t h i s ambrosia beetle owes i t s e l f to chemical changes during the death process. Since both sexes react to the odours, i t i s i n f e r r e d that i n i t i a l attack on a tree involves a primary attractant from the host. •Implications are seen for the r o l e of behaviour i n the ecology of t h i s insect, Other implications are seen for the use of the newly acquired information i n bioassay techniques i n which the insect w i l l serve as a test instrument for chemical studies of host a t t r a c t a n t s . GRADUATE STUDIES F i e l d of Study: Forest Entomology S t a t i s t i c a l Methods i n Forest Research Forestry Seminar Topics i n Wood Anatomy Biochemistry Experimental Zoology Directed Studies -J. W. Wilson J„ R. Campbell J . H. G. Smith Forestry Staff W. S. Hoar Zoology K. Graham PUBLICATIONS Francia, F. C. & Snyder, T. E. I960,, A summary of P h i l i p p i n e termites with supplementary b i o l o g i c a l notes. P h i l i p p i n e J . S c i . 89(1): 63-77. Francia, F. C. 1960. Protecting buildings from termite and fungus damage. FPRI Technical Note No. 10, 1-4. Francia, F. C. 1960. Studies on the control of ambrosia beetles which damage newly-cut timber. P h i l . Lumber-man, 6(2): 5-7. Francia, F. C„ 1958. Powderpost beetles (bukbok) i n -jurious to wood and other forest products. P h i l . Lumberman, 4_(2) : 6-et seq. Francia, F. C. 1957. Pinholes i n logs and lumber. P h i l . Lumberman, _3(5): 16-18. STUDIES QP SOME ASPECTS OP BEHAVIOUR IK THE AMBROSIA. BEETLE TRYP033EMDR01I LINEATUM (OLJ.\\ i^m) by FADSTINO C FBAHCIA. B»S«F«, University of the Biilipjgines, 1952 M.S. j, State University of Hew York, College of Fbnastfcry at Syracuse University, 1957 A THESIS SUBMITTED US EAHHAL FULSILME1C21 OF THS REQUIREMENTS FOR THE I3BGRKB. OF Doctor of Riilosophy i n the Faculty of We accept this thesis as conforming to the required standard THE UNHERSITY OF BRITISH GOIUMBIA. JtQy, 1965 In p r e s e n t i n g t h i s t h e s i s i n p a r t i a l f u l f i l m e n t o f t h e r e q u i r e m e n t s f o r an advanced degree a t t h e U n i v e r s i t y o f B r i t i s h C o l u m b i a , I a g r e e t h a t t h e L i b r a r y s h a l l make i t f r e e l y a v a i l a b l e f o r r e f e r e n c e and s t u d y . I f u r t h e r a g r e e t h a t p e r -m i s s i o n f o r e x t e n s i v e c o p y i n g o f t h i s t h e s i s f o r s c h o l a r l y p u r p o s e s may be g r a n t e d by t h e Head o f my Department o r by h i s r e p r e s e n t a t i v e s , , I t i s u n d e r s t o o d t h a t c o p y i n g o r p u b l i -c a t i o n o f t h i s t h e s i s f o r f i n a n c i a l g a i n s h a l l not be a l l o w e d w i t h o u t my w r i t t e n p e r m i s s i o n . The U n i v e r s i t y o f B r i t i s h Columbia Vancouver 8, Canada^ Date v 7 ABSTRACT Tim factors and mechanisms which consistently deliver newyly over-wintered adults of the ambrosia \"beetle •Prypodendron lineatura (Olivier) to specific host trees under specific conditions have been considered!. Studies and analyses of the behavioural patterns of the beetle im respect to light, host tree factors, and factors of the environment -were made in recognition of the fact that preiimdasary research results demons-trated the predominant role that reactions to light played!, in the behaviour of the beetle. The results of the studies showed that the beetles, before flight, were; strongly phototacstic at temperatures in the range of 5° to 56°C. Bssitive photic response was inhibited i n am increasing percentage of indi-viduals at temperatures above 56°. At 58°0, positive response to light by the beetles ceasedl to exists The non-flown beetles reacted; quickly to angular deviations of alignment im respect to a light source and turned, with almost equal readi-ness toward the light regardless of i t s atmgle. The beetles' responses to light may be classified as follows: (l) not inhibited, the beetles imme-diately, move toward the source of light; (2) ini t i a l l y inhibited positive responsej (5) inhibited positive response, the beetles may or may not imi-c t i a l l y inhibited but the general movement toward She light source i s not direct| (4) completely inhibited response, 'She beetles move with no appa-rent response to light* The photopositive response was found to mask: the other potential capabilities of the non-f lerwn beetles. Exclusion of the photic stimulus from £light-inexperiencedl beetles resulted in response to host odour in an odour f i e l d . PligM eaqaerience was found to modify partially the simple photic reaction of some individuals in the absence of host odour, but the majority of the \"beetles' responses remained unchanged. The behaviour of Telodendron is not strictly stereotyped i a the sense that i t follows a definite pattern. While flight may be normal as & conditioning mechaaasmi preparatory to alighting and host finding, response to an \"attractive1* odour source and subsequent boring behaviour may be exhibited, in the absence of previous flight experience, under certain conditions. i i i TABLE OF CONTENTS Bags ACMOWI^MBNTS .. .. .. 1 LIST OF TABLES; 2 LIST OF ILLUSTRATIONS 5 INTRODUCTION .. 5 MATERIALS AND METHODS . :. 12 The Experimental Insect . . . . . . . . . . . . . . . . . . . . 12 1. General considerations . . . . . . . . . . . . . 12 2. Source of the experimental supply of beetles and methods of handling and storage . . . . . . . . . . . . . . . . . 14 5. Treatment of the beetles for testing . . . . . . . . . . . 15 Host Tree Factors . . . . . . . . . . . . . • • « . ....-••» . 16 H General considerations . . . . . . . . . . . . . . • . • • 17 2. Source of host wood and preparation for testing . . . . . 19 5. Preparation of wood factors as stimuli to the responses ofi^the beetles • . • . • • . . . . . . . . • . . . . . . . 20 The Light Factors 21 1. General considerations . . . . . . • . . . . . . . . . . . . 21 2. Description of the equipment used and methods of testing . 22 Accessory Factors . . . . . . o . . . . . . • • ••• • • • • « •• 26 1. Increasing temperature: its provision and use in the tests . . . . . . . . . . . . »-••*• ... . . . . . • •• 27 2. Airstream: its provision' and use in the tests . . . . . . 50 3. Increasing atmospheric pressure: its provision and use in the tests . • ... ••• •• ... . . » •• . . . . . . « • » « • • 51 Methods of Observation . . . . . . . . . . . . . . . . . . . ... 53 EXPERIMENTAL RESULTS; . . . 56 TABLE QP CONTENTS (continued) Page 1. Responses to white light under normal laboratory conditions . . . . . . . . . . . . . . . . . . . . . . . . . . . 36 2. Photic responses in the presence of air current with or without wood odour • . .. . • • . • . . . . . • • • .. • • • • • .. 41 5. Photic responses in the presence of \"green\" wood factors . . . » 44 4. Photic responses in the presence of \"attractive wood factors ........ 44 5. Responses to background black-and-white patterns . . . . . . . 49 6. Responses to white light under increasing temperature . . . . 49 7. Responses of Trypodendron to various stimuli in the dark . . . 57 DISCUSSION OP RESULTS . . . . . . . . . . . . . . . . . 60 1. Photic behaviour in Trypodendron: its ecological significance and its underlying mechanisms . . . . . . . . . . . . . . . . 60 2. The modifiers of phptic behaviour: their relation to host-finding .... . o . . . . . .. . . . . . . . . . . . . . . » . 63 3. The behaviour of Trypodendron: its use in bioassay techniques 72 CONCLUSIONS . 74 LTTERATURE CITED . 78 ACMCWIMXJMENIS To Dr. Kenneth Graham, professor of forest entomology, University of British Coumbia, for his invaluable advice, encouragement and kindness throughout the conduct of the experiments and preparation of this thesis, I am deeply indebted* To Dr. John A* Chapman of the Canada Department of Forestry, Forest Entomolpgy and Pathology Laboratory, Victoria, B. C , for his cooperation, courtesies and kindness in furnishing biological specimens and in offering helpful suggestions, I am deeply grateful. To the members of the Graduate Committee: Drs. J. <3F. H. Campbell, P. G. Haddock, W. S. Hoar, R. W. Wellwood, and D. J. Wort for their interest in the conduct of the work on this thesis, grateful acknowledgments are extended. I also give my thanks to many individuals including Dr. A. Kozak, Hessrs. John Walters, J. M. Kinghorn, J. T. Welf, and P. Demko for giving help in various ways, and to several institutions including the National Research Council of Canada, British Columbia Loggers Association, and the i '• Department of Zoology, University of British Columbia, for their assistance and permission to use laboratory equipment and space. Especially to the External Aid Office, Government of Canada, Ottawa, and the Forest Products Research Institute, Republic of the Philippines, for jointly granting a Colombo Plan Fellowship which permitted the pursuance of the work on this thesis, I extend my wholehearted thanks. 1 LIST OP TABLES Table l o . Page 1. Summary of responses of flight-inexperienced male and female Trypodendron to a horizontal beam of light under normal laboratory conditions ,. 57 2. Summary of responses of flight-experienced male and female Trypodendron to a horizontal beam of light under normal laboratory conditions . 40 5. Photic responses of adult Trypodendron under increasing temperature . . . . . . . . . . . . . . . . . 56 4. Photic responses of adult Trypodendron as influenced by gravity and increasing temperature . . . . . . . . . . 57 2 LIST OP ILIiU^TRATIONS Figure Ho. Page 1. Trypodendron lineatum (Olivier) adult female beetle. Actual length 3 to 3.5 mm. (After Graham, 1963) . . . . . . . 13 2. A Reichert microscope illuminator \"Lux FNI\" operating through a variable resistor with an amperage indicator . . 25 5. Diagram illustrating plan of view of the i n i t i a l starting points or angular displacements of the beetle Trypodendron in tests for its responses to a horizontal beam of white light . . . . . • 25 4. Apparatus used for testing responses of Trypodendron to light in a temperature gradient 28 5. Apparatus and set-up for testing photic responses of Trypo-dendron in relation to gravity and increasing temperature . 29 A 6. Apparatus for testing photic responses of Trypodendron under increased atmospheric pressure . . . . . . 55 A 7. Scheme of programme of experiments on the photic responses of Trypodendron lineatum (Olivier) in relation to various host wood factors and conditions of the environment ....... 55 8. A to H. Representative paths of response to a horizontal beam of light by newly overwintered adult females of Trypodendron lineatum (Oliv.) . . . . . . . . . . . . . . . 58 9. A to H. Representative paths of response to a horizontal beam of light by newly overwintered adult females of Trypodendron lineatum (Olivier),. . . . . . . . . . . . . . 39 10. A to D. Representative paths of response to an overhead source of light by newly overwintered adults of Trypoden-dron lineatum (Oliv.) . . . . . . . . . . . 42 11. A to D. Representative paths of flight-inexperienced and flight-experienced adult Trypodendron in response to a gentle stream of air without odour or with attractive host wood odour . . . . . . . . . . . . . . . . . . . . . . . . 45 12. A to D. Representative paths of responses to light and to an airstream with or without attractive host wood odour by flight-experienced as well as flight-inexperienced adults of Trypodendron placed on a perforated area on a sheet of paper above a piece of \"green\" wood . . . . . . . . . . . . . . . . . . . 45 5 LIST OF ILLUSTRATIONS (continued) Figure Fo. Page 13. A to D. Representative paths of responses to light and to an airstream with or without attractive host wood odour \"by flight-inexperienced as well as by flight-experienced adults of Trypodendron in direct contact with a piece of M green\" wood 46 14. A and B. Representative paths of responses to light and to an attractive piece of wood under perforated area on sheet of paper by flight-inexperienced and flight-experienced Trypodendron beetles . . . . . . . . . . . . . . 50 15. A and B. Types of response to light and to an airstream without attractive wood odour by flight-inexperienced and flight-experienced Trypodendron beetles on a perfo-rated sheet of paper above attractive host wood. . . . . 51 16. Representative paths of response to light by flight-inexperienced and flight -experienced adult Trvjspdendron beetle placed on a perforated sheet of paper above attractive host wood and subjected to a gentle stream of air with attractive wood odour . . . . . . . . . . . . . 52 17« A and B. Representative paths of response to light by the flight-inexperienced or flight-experienced Trypodendron beetle in contact with a piece of attractive host wood . 55 18. A and B. Representative paths of response of flight-inexperienced and of flight-experienced adult Trypodendron beetle to light and to a stream of air with attractive wood odour. . . . . . . . . . . . a . . . . . . . . . . . . 54 19. Representative paths of response of the flight-experienced Trypodendron beetle placed in contact with a piece of attrac-tive wood in relation to light in the presence of an air-stream with or without attractive host wood odour. . . . 55 20. Effect of increasing temperature on the photic, responses of Trypodendron lineatum (Olivier)-.. . . . . . . . . . . 58 21. Comparison of effects of gravity and increasing temperature on the photic responses of Trypodendron lineatum (Olivier) 58 4 IMrRQDUCREIQN Among the general problems concerning ambrosia, beetles are those which pertain to their host tree relations. More specifically i s the ques-tion as to the factors and meahaiaifflms which consistently deliver these insects to host trees of specific kinds and conditions. The question i s prompted by the fact that ambrosia beetles show remarkable ability in dis-covering felled, wir^tbrown, or dying trees im a forest, or logs i n water storage, \"selecting\" them as to botanical identity and state of morbidity in the sequence of changes between l i f e and death. They are even able to detect these special requirements despite heavy coverage of the log surface with substances objectionable or confusing to human olfactory sensation, such as creosote and kerosene, or despite a residue of the insecticide lindane which ultimately Trills those beetles as. they crawl over the log surface. Two associated problems are posed by the foregoing observations. One concerns the identity of the intrinsic and extrinsic factors and the behavioural patterns and mechanisms involved!, i n the process of host disco-very and discrimination. The other concerns the nature of the facttes i n a log and the mechanisms of their origin which account for the differential attack by these beetles on logs of different kinds and conditions. These problems are of both academic and practical importance. Considerations of the general problems, in turn, pose subsidiary questions. Does attack involve random trial-and-error sampling by the beetles, or does the insect detect i t s host from a remote point? Of the various conceivable differences between living and dying trees., which of them should be capable of providing essential cues for the beetles? If 5 6 detection is from remote points, •which of the possible phenomena, i n a dying tree may act at a distance, and which of the psa-mi.ac conceivable senses; in insects may take part in remote detection? If odours play an important role, as may be inferred from certain f i e l d studies, (Chapman, 1962, 1965), are they absent from the healthy living tree, or merely occluded within the cells until mechanical alterations, such as permeability or hydrostatic pressure changes, release them s&en the tree begins to die, or does the change also involve the disappearance of chemical repellents? If attractants are normally absent, why do they not develop in the living tree, and why do they form as i t begins to die? If a chemoal process, i s iwolveii, does i t involve simple oxidation, simple cleavage, condensation, polymerization, molecular rearrangement, or abnormal metabolism? If abnormal metabolism i s involved, how? may normal and/or abnormal metabolism be ar t i f i c i a l l y modified!, to accelerate or retard the formation of chemical attractants? i f chemical attractants are formed, what are their group or specific chemical natures? Can their attractancy be obscured by other substances presented simulta-neously with them to the insects?. The foregoing questions are of a factual and observational nature, some having practical implications . There is to be considered also the conceptual aspect which pertains to the relationships of ambrosia beetle behaviour to the general theory of animal behaviour. Kennedy (1956, 1958) refers to three types of hypothesis concerning the nature of instinctive behaviour. One is the simple physiological one such as Loeb favoured, according to which reflex and instinctive action do not differ in prineijftLe and changes, of behaviour pattern are due primarily to changes, in the reflex thresholds brought about by changes in the state of receptors and effectors. 7 A second hypothesis, \"based on the work of reflex physiologists, notably Sherrington, also recognizes no difference of principle between reflex and instinctive action but attributes changes of threshold primarily to the integrative activity of the central nervous system. A third hypothesis, advanced by the modem ethologists, draws a distinction of principle between reflex and instinctive action, the latter depending on an internal drive-generating mechanism which, even -vftien triggered by an external stimu-lus, has nothing in common with reflex: mechanisms. The general study comprises several largely distinct but not entirely mutually exclusive tasks. These consist of analysis of behaviour in the insect, experimental modification of living and dying trees, and chemical and histochemical analysis of changes in chemical precursors and products of trees as they gradually die. The objective of the present phase of the undertaking i s to investigate the behaviour of the insect, particularly i n relation to the circumstances affecting its response to host tree factors. The present investigations pertain to the ambrosia beetle species Trypodendron lineatum (Olivier) which has been an object of much concern and research in recent years. Earlier studies on this insect (Graham! and Werner, 1956) revealed that the reproductively mature beetles, freshly removed from their overwintering quarters i n spring, are entirely dominated by a strong photopositive response. In the presence of light, they do not bore into the kind of wood that i s normally being attacked by non-captive individuals in the field, in darkness, they bore readily into attractive wood, or into non-attractive wood to which gas-distilled extracts of attractive wood are applied. This contrasts with events in the f i e l d 8 where attack takes place in broad daylight. Subsequent observations (Graham, 1959) showed that the ability of the beetles to bore into wood in daylight depends on their f i r s t having a period of flight. This appears to simulate one stage in the natural sequence of events in the f i e l d . Balfour and Iteramonov (1962) contend- that flight i s not absolutely neces-sary. They found that under severe confinement, a small percentage of beetles w i l l attack wood while exposed to light without hawing been pre-conditioned by flight. Further experience by Grahamt (1965), however, sustains the original basic conclusion that the in i t i a l l y strong photo-positive response in overwintered adults of Trypodendron is a major obstacle to their boring-in activity until they have experienced flight. It i s conceivable, however, that Balfour and J&ramonov's observations signify that in certain populations or in certain samples of beetles from themi different thresholds of responsiveness to host factors already exist among the various individuals and that the threshold is lowered for a l l individuals by the flight experience. It should be noted that in their caution of precluding the possibility of their beetles having had flight experience, Balfour and Paramonov used specimens which had been extracted ar t i f i c i a l l y from brood logs and stored until spring. Thereby they denied the beetles the normal prehibernation flight. Further investigations (Graham, 1961) indicated the importance of air-swallowing as a mechanism by which photic reactions are subdued to a point where responsiveness'to host factors is released. Mnirmmn periods of flight for this purpose were, however, not ascertained. The discoveries to this point left various details of behaviour in Trypodendron untouched. 9 No evidence exists to indicate that T. linea-tum discovers i t s host tree or log \"by a random, trial-and-error process. During the attac-king phase of i t s life, i t appears never to settle on hosts other than those of a particular species in a particularrstate between the living and the dead. These are the logs into which they will bore to establish their broods. It is thus evident that the mechanism of detection operates over an appreciable distance. Entomologists are familiar with three forms of perception which enable insects to receive ^ information at a distance, namely vision, audition, and olfaction. Certain evidence exists also for a sense of perception of bioelectric fields (tew, 1961, 1962) even though no structure for their per-ception is known. Of the various properties of plants which emit information that is detectable at a distance, two classes of factors are well-known. One is odour j the other i s of a visual nature, viz. contrast, form, and colour. Perhaps bioelectric fields should be considered as being a possible source of information to insects (Haw, 1961, 1962). As to the significance of voltage gradients (Parr, 1945) or bioelectric oscillations (Scott, 1962) as a source of signals for insects, nothing is known. It is not entirely inconceivable that movement of fluids and gases in a tree may be accompanied by subsonic or supersonic propagations of low intensity, but experimental evidence i s lacking. The fact that Trypodendron, during the attacking phase i n i t s l i f e cycle, appears not to settle on the \"wrong\" kind of logs or trees indicates that visual cues are not primarily or predominantly involved in host disco-very. This is not to deny that sight is probably involved in their guidance mechanism. Oh the other hand, the important role of odour in the guidance of I D insects i s well-known. Furthermore, hidden log experiments in the f i e l d (Chapman, 1962, 1965) have demonstrated convincingly that odour must be a prime factor in host discovery by Trypodendron. With this evidence, i t appears prudent to examine as far as possible the role of odours before testing for the operation of factors whose importance or existence for insects i s uncertain. The foregoing observations on T. lineatum accordingly indicated, the i n i t i a l dominance of the photic response and the release of a chemo-tactic response by exclusion of light, or by flight experience, the latter operating by inducing the swallowing of air. It was also evident that an olfactory guidance mechanism is involved, and volatile chemicals extracted from attractive wood are capable of inducing attack (boring-in) on otherwise non-attractive wood. An olfactory response to host factors was suggested, by their tendency for an unstable aggregation about odour sources from attrac-tive wood provided that light is excluded and that the air in the immediate surrounding of the beetles was not excessively dry (Graham and Werner, 1955) • The knowledge about Trypodendron now raised as many questions as i t had answered. It was not known how long a period of flight i s required for the appearance of a change in the response to host factors i n the presence or absence of host factors. Heither was i t known whether the effect of flight on the photic and host factor responses i s all-or-none, or whether i t involves a progressive change of thresholds. It was not known either whether the beetles could orientate to a stationary odour gradient, or to an airstream carrying odour. The laboratory experiments implicating odour i n behavioural effects were at this point quite meagre. By the same token, there were no satisfactory criteria by which olfactory responses 11 could \"be detected, assessed, or clearly segregated from the other responses which are incorporated into the more complex: reaction of boring into wood. The objectives of the present investigations were to explore factors and mechanisms of the orientation behaviour of Trypodendron lineatum (Olivier) during that stage in its l i f e when i t searches out, selects, and attacks logs. This period is in the overwintered reproductively mature adults. The immediate study is confined to pedestrian beetles because i t appeared that much of their behaviour could best be gained under this circums-tance. The study of photic reactions is here limited to the photopic (light adapted) state. Special attention i s directed at photic reactions and factors which modify these. MATflRTATfi AND METHODS Trypodendron lineatum (Olivier) (Fig* l ) of the family Scolytida© was the species of ambrosia beetle used i n these studies. 1. General considerations.-The species T. lineatum i s , of a l l the thousand or more world ; species of ambrosia beetles, one of the most suitable for study. It occurs over an extremely wide geographic range, being found in the temperate coni-ferous forestcof Europe, Asia, and North America (Hadorn, 1955; Chamberlin, \\ 1939; Novak, 1963; Bletchly and White, 1962). A considerable body of l i t e -rature is available as background information on its general habits, biology, physiology, and ecology (Hadom, 1955; Chamber l i n , 1959; Chapman, 1955, 1958, 1962, 1965; Chapman and Dyer, 1960; Chapman and Kinghorn, 1961; Dyer and Chapman, 1965; Graham, 1959, 1961; Graham and Werner, 1959; Nijholt and Chapman, 1964; Novak, 1965a, 1963b; Prebble and Graham, 1957; Radinsky and Daterman, 1964a, 1964b). Its habit of overwintering during a period of i t s adult l i f e in the forest l i t t e r , duff, or under flakes of bark on trees adja-cent to logging or logged-over areas (Hadorn, 1933; Novak, 1965; Chapman, 1960; Ghapmajn and Kinghorn, 1961; Dyer, 1962, 1965) permits collection of biologically uniform material in sufficient quantity for convenient experi-mentation in the laboratory. This contrasts with most other ambrosia beetles which are far less synchronous and do not accumulate in an intermediate storage medium,but search new logs almost directly following emergence from the brood log which nurtured them. Trypodendron adults are amenable to pro-longed storage under refrigeration without showing marked loss of vigour or ability to f l y or walk provided they are not exposed to desiccation of 12 13 Fig, 1 . Trypodendron lineatum (Olivier) adult female \"beetle. Actual length 5 to 3 .5 mm., (After Graham, 1965) 14 more than 25 per cent of their body weight (Nijholt and Chapman, 1964). The visible differences between the sexes render them easy to define for the purpose of the experiments. The versatility of Trypodendron in accepting a wide range of genera of coniferous trees offers considerable freedom of choice in the kinds of wood which may satisfy certain needs of experimental conve-nience. The same diversity in respect to acceptable host species may provide increased opportunities for discovering the essential chemical attractants by correlative evidence. 2. Source of experimental supply of beetles and methods, of handling and storajjey'-The source of the beetles was in the Gowichan Lake district of Vancouver Island, British Columbia. The time of collection was in late March and early April, 1964, just prior to the beetles* normal emergence as. reproductively active adults from their overwintering quarters. Collections were obtained within a. zone of about 100 meters inside the margins of standing forest adjacent to areas in which logging had been in progress about 18 months previously. Here the brood of new beetles, generated during the season preceding the collection, occurs in the duff layer near the base of standing trees, and under loose flakes of bark on the lower stem of very old Douglas f i r trees. Old adults may occasionally be found among the population of overwintering new adults at a proportion of about 1 to 50 per cent (Chapman and Mijholt, 1965). Bark flakes and forest duff containing the beetles were freed of coarse material and stored in plastic bags in a refrigerator at a temperature of about 0 to 5°C. Whenever beetles were required for the experiments, they were obtained by spreading the beetle-containing medium, a couple of handfuls 15 at a time, i n a shallow pan floating on 'cater at a temperature of about 50°G. As the beetles became activated, they were transferred to amai^ jars con-taining moist crumpled paper towelling or bits of moist moss to prevent loss of body moisture and to minimize mutual contact among the beetles, as other-wise the beetles bite off each other's front tarsi. Specimens thus mutilated are hardly able to walk normally and become useless for the purposes of the experiment. Care also had to be exercised in intercepting the beetles before they could take off i n flight as i t i s known that this factor modifies their responses to light in the presence of odour from attractive wood (Graham, 1961) and to attractive odour in the presence of light. 3. Treatment of the beetles for testing.-The supply of beetles used in the experiments was segregated* as to sexes. They were placed in jars with moist paper or moss. To prevent the beetles from becoming very active in the presence of light at room temperature, the jars containing them were kept in a thermos jug containing ice. Beetles were removed from the jars as needed in the experiments. The beetles were tested for their photic behaviour immediately after removal from cold storage and without allowing them to experience flight. Later, the same beetles were given a period of captive flight by means of a procedure described by Chapman (1956). This was done for the purpose of studying details of the effect of flight experience on photic orientation as well as observing the time patterns of flight activity per se. Each beetle was glued by its pronotum to the underside of the tip of .^iicardboard-insect-mounting point. The beetle was affixed in such a way that i t was horizontal and that its wings were not obstructed in motion by the paper point or any glue. It was also noted that interference of glue with anten-16 mal movement suppressed a l l effort of flight. Four or five beetles indivi-dually affixed to separate points, each set on an entomological pin, were observed concurrently. At every one minute interval, the flight activity of each beetle was observed and recorded horizontally across graph paper ruled at ten lines per inch. Positive activity on each beetle was indicated by an ttx\" sign, while inactivity was represented by a blank square. Total flight as well as periodicity of intermittent flight were thus known. The beetles were kept under these flight-testing conditions for various periods of time, from 5 minutes to 6 hours. To prevent possible loss of body moisture during the flight period and exposure to light, the piece of balsa wood to which the pins supporting the beetles were stuck was kept i n a shallow pan of water. After the prescribed period of flight, the beetles were removed from the card points to their respective vials, care being exercised so as not to cause injury to the beetles, and again tested for photic response, one at a time, or for responses to some other stimuli concurrently with the light factor. Host Tree Factors Like the beetles i n being important as both objects and instruaents in the study of behaviour i s the host tree. The wood ofcthe:hbstitree i s eventually to be investigated for products of biochemical changes, factors and mechanisms which are involved in rendering the wood attractive to ambrosia beetles. It is noteworthy that the attractive condition for the ambrosia beetles develops develops in a tree from the effects of injuries and exposure after a certain period of time. Before the attractant factors can be clearly interpreted preparatory to isolation and evaluation, i t is necessary to . elucidate the accessory conditions required to induce aa effective response of the insect to the host. For this purpose, portions of known attractive and non-attractive host materials were provided as reference standards of host factors. 1. General considerations. -The recorded host trees of Trypodendron are Douglas f i r (Pseudo-tsuga menziesii /SfLrbJ? Franco), amabilis f i r (Abies amabilis Jjtovygjfeorb.), grand f i r (A. grandis ^Douglj7 Forb.), western hemlock (Tsuga. heterophylla Sarg.), spruce (Picea spp.), pine (Pinus spp.) in British Columbia (Erebble and Graham, 1957) ; noble f i r (Abies procera), larch (igrix lepto-lepis) t spruces (Picea abies and P. sitchensis), pines (Pinus sylvestris and P. radiata) in Scotland (Bletchly and White, 1962), and several species of spruce in Europe (Novak, 1965b). Host tree material of Trypodendron provides several different kinds of stimuli which may affect this insect in various ways. These include the tactile stimulus arising from contact with solid object. The solid surfaces may offer various topographic features varying from smooth plaques and deep narrow fissures as exist in bark texture to relatively flat and coarse surface as exists on the cut or sawn ends of logs. The colour may also vary considerably but i t s stimulating character i s unknown. Odour i s perhaps the most important stimulus to consider because mostainsects react to i t i n a very remarkable manner (Dadd, 1963; Dethier, 1965; Budden-brock, 1958; Kennedy and Booth, 1959a, 1959b, 1960; Roedear, 1965; (Sara and Vite, 1962). En Trypodendron^ the odour of a suitable host tree appears to be a guiding factor (Chapman, 1965). Taste is a factor to which most insects have a low threshold of sensitivity (Dethier, 1965) and which may 18 affect the reactions of the beetles as they begin to chew into wood. Another conceivable stimulating factor i s the visual form or mass of the host (Dethier, 1957) but its influence on Trypodendron is not known. As i t is the purpose of the study to deal with factors involved in the attraction of the beetles to their host, material was chosen especially to provide a source of the odour stimulus. Accordingly, features of bark topography and colour and the property of taste played no part in the selection of the speoific host for the preparation of the samples. There were other considerations in the choice of wood. One is the choice of tree species among the potential or known host species of Trypo- dendron. Fbr experimental convenience, certain preferences had to be made, since in the particular area where the experimental beetles were obtained, western hemlock and Douglas f i r are not only the most commonly available trees but are also usually the most heavily attacked by Trypodendron (Graham and Werner, 1956). Of these two trees, Douglas f i r is noteworthy for its white sapwood region which becomes attacked by the beetles and i s easily differentiated from the reddish region which is immune to attack. Also, Douglas f i r wood had been a subject of chemical investigation for i t s attractant properties (Graham and Werner, 1956). Douglas f i r , however, possesses a limited sapwood layer which prohibits obtaining convenient siae of tranverse slices for testing attack response; of the beetles and, more important, appears to change from the non-attrative \"green\" condition to the attractive \"ripe\" condition in a shorter time than does hemlock. This instability is disadvantageous where alteration of host material condition during the course of the experiment is undesirable. For this reason, western hemlock appeared preferable as a source of wood in studies of 19 attack responses and as a source of odour in studies of orientation to light and air flow. Another consideration in the choice of wood concerns i t s state of attractiveness. The state of attractiveness can be measured only by the response of the beetles. The chemical identity of the attractants is not yet known. In so far as attractiveness to the beetles is concerned, however, a host tree exists in several conditions ranging from a '\"green'* state as exists in the wood of a newHy felled tree to a \"ripe\" state which i s highly favoured by the beetles and to a \"spent\" state in which the attraotant factors are no longer present as in logs, wood, or finished dry lumber. The attractive or \"ripe\" state develops during winter in the sapwood of the unconverted host trees felled during autumn and winter. Spring-felled trees with tops retained, or cut only into long log lengths remain unattrad* tive to the beetles through the spring flight period of Trypodendron but become highly attractive i f cut into short log sections at the time of felling (Johnson, 1964). 2. Source of host wood and preparation for testing. -The \"ripe\" and the \"green\" wood materials came from the same loca-l i t y as the experimental beetles. The attractive state of the wood samples was assured by taking them fromi polyethylene-sheet-covered portions of logs of host trees felled in early winter and allowed to remain in the woods until attacks occurred on the uncovered portions. These \"ripe\" samples as well as \"green\" samples from spring-felled host trees were stored as thick slabs i n plastic bags in a freezer at about -17.8°C (0°F). Samples of both \"ripe\" and rtgreen\" wood for testing attack responses of the beetles were prepared by debarking the frozen slabs, 20 squaring the edges, and planing the surfaces with a woodworking \"jointer\" machine. The shavings obtained from the planing operation were saved in plastic bags and stored in the freezer. 5. Presentation of wood factors as stimuli to the beetles' responses The method used for evaluating the olfactory response consisted of testing for the retention of the beetles at an odour source while they were subjected to the potential attraction of a beam of horizontal light or a broad f i e l d of light whose source is overhead. A sheet of paper, finely perforated with pinholes at the center, was placed over a small section of either \"ripe\" or \"green\" wood, about 5.0 cm. by 7.0 cm. by 1.5 cm., as desired. The beetles were placed individually on the perforated area to examine the ability and behaviour of the beetles in orienting to a gradient of odour emanating from the wood beneath the paper without involving a stream of air. Another condition under this series of tests ;consisted of associa-ting wood with a stream of air alone or with \"ripe\" wood odour borne by a stream of air. The purpose of the test was to determine the possible occur-rence of an anemotactic response prompted by the host odour. This would concern an alternative mechanism to the odour-gradient theory of host finding. In another series of tests, the. beetles were allowed physical contact with the piece of either \"ripe\" or \"green\" wood. This series would provide not only thigmotactic behaviour responses on physical contact with the piece of wood but also chemotactic behaviour responses as a result of tarsal contact and/or oral tasting of the host wood. In addition, the effect of contact with the source of the attractive wood odour may provide a different response. The jtfigot Factors In its most comprehensive aspects, the photic behaviour of an insect would be concerned with the directional changes and time patterns of its orientation reactions under diverse conditions of a photic environment. Light may be varied as to intensity level, specific wavelength, wavelength balance in mixed colours, degree and plane of polarization, discreteness or diffuseness, and angle and plane of incidence relative to the axis of the insect. \"Various contrast and colour patterns, both stationary and moving, in the visual f i e l d may influence the responses (Reiohardt, 196l). A l l of these are possible elements and variables to which Trypodendron is subjected in its natural environment. In addition, the insect may be expected to respond differently in the dark-adapted and light-adapt edl states as demons-trated by Raymont (1959) with the beetle Dineutes. The present studies employed mainly a heat-filtered, unidirectional white light of constant intensity and colour temperature, projected in a beam toward the beetles and a broad f i e l d of overhead light provided by ceiling fluorescent lamps. 1. General considerations.-Light is an important factor of the environment to which insects react either positively or negatively under certain conditions. Light i s defined as that portion of the electro-magnetic spectrum ranging in wave-length from about 200 to 10,000 m/u or about 2000 to 100,000 %l The light visible to human eyes ranges from 4000 to 7500 %. while that to a bee's eyes ranges from 3000 to 6500 A (Carthy, 1965). In terms of colour which the different wavelengths represent to human eyes, the bee's eyes can distin-guish colour in the ultraviolet range and not in the far red range of the spectrum. As a token stimulus, light is \"wsry effective In that i t s reflective and absorptive characteristics provide insects some kind of information about their environment. Changes in light intensity associated with shadows of moving objects notify the insects of the presence of prey or enemy. Because infrared wavelengths usually accompany visible light under aerial conditions, brighter illumination means a warmer, drier environment, and a lower illumi-nation may mean a cooler, damper one (Brown and Erosser, 1962) • 2. Description of the equipment used and methods of testing.-The source of white light was a Reichert microscope illuminator \"Lux FNI\" (Pig. 2) operating through a variable resistor with an amperage indicator. Its normal rating was 50 watts at 6 volts and 5 amperes. This equipment gave control over light in respect to focus, aperture, intensity, and to a certain extent, colour temperature. Because the wavelength balance (colour temperature) of light from a tungsten filament changes with voltage of input current, a fixed setting of the rheostat was adopted for an input of 4.0 amperes. The incident light to which the beetles were exposed was measured by a \"Ehotovolt\" photometer model 200 M as ranging from 25 to 50 footcandles. Colour temperature was judged to be similar to that of a 40-watt incandescent bulb operating at normal voltage, the value of which is rated at 2760° Kelvin. To insure that the responses to the horizontal beam of light were purely photic and not thermal, a heat f i l t e r from a Bausch and Lomb projector was interposed. The overhead source of light was provided by four \"General Elec-tric \" cool-white F40 GW fluorescent ceiling lamps at a distance of about two metres above the experimental table. The incident light furnished by these lamps ranged from 50 to 60 footcandles as measured by the ;. ; Fig. 2. A Reichert microscope illximinator \"Lux SKI\" operating through a variable resistor with an amperage indicator. It was used as the source of a horizontal beam of white light. A Bausch and Lomb heat f i l t e r i s shown in front of the illuminator. 24 \"Ehotovolt\" photometer. The simple photio reactions were tested by placing the beetles, one at a time, at a central point oh a sheet of white writing paper. A narrow beam of heat-filtered light, as defined earlier, was projected horizontally across the surface of the paper. The beetle, i t s sex rioted, was aligned with the axis of the beam. As the beetle was allowed to walk, a pencil line was traced behind i t until i t reached the edge of the paper, whereupon i t was returned to the starting point, lax some tests, the o beetles were aligned in eight different directions, 45 apart, in respect to the Light source. In other tests, only four directions at 90° intervals were tried. These starting points are illustrated in Fig. 5. The syste-matic testing of angular displacement was provided in recognition of the fact that certain insects react at different rates to different axial displacements relative to the light sour<5o,(Euddenbrock,:,1958; Fraenkel and Gunn, 1961). Besides the systematic tests which began with a fixed starting direction, other tests with different i n i t i a l directions of align-ment were tried in order to examine for possible adaptation of the insect during successive tr i a l s . After the beetles were each exposed to a hori-zontal beam of light, they were tested for their responses to a broad fi e l d of light. Essentially the same positions on the white sheet of paper were used in aligning the beetles. The photic reactions were tested in the presence of other factors such as wood odours, air currents, increasing temperature, gravity, and atmospheric pressure. The beetles included both flight-experienced and flight-inexperienced individuals. Previous observations indicated that the individual beetles 25 T Direction of light 1 V 5 Fig. 3. Diagram illustrating plan of view of the i n i t i a l starting points or angular displacements of the beetle Trypodendron in tests for i t s responses to white light. 26 walked toward a point source of a horizontal team of light and began attempts to f l y when light was present i n a l l directions as provided by the ceiling lamps. These observations led to undertaking a test to determine the responses of the beetles to light coming from below the horizontal plane of sight of the beetles. For this purpose, the beetles were indi-vidually placed on a sheet of transparent glass below which a light was placed. Following this test, a combination of light coming from above and below the beetle was tested to determine the responses of the beetles. The orientation behaviour and responses to host wood factors ware also studied in the absence of light. The beetles were individually placed in a. cardboard box, about 50 cm. by 45 cm. by 15 cm., painted black inside. The beetles were placed on a predetermined position on a piece of paper inside the box which at times contained either an attractive or a \"\"green1* piece of wood as a particular test demanded; the box was afterwards covered for a period of about five minutes; then i t was opened to determine the posi-tion of the beetles. Another condition of testing consisted of passing wood odour through a hole in one side of the box. Accessory Factors The behaviour of insects under natural conditions i s often stereo-typed, described in terms of inborn pattern of behaviour commonly known as. instincts (Dethier and Stellar, 1964; Scott., 1963; Fraenkel and Gunn, 1961). Inborn reflexes, and reflexes \"conditioned\" by previous experience may occur but are often masked by inhibitors and integrated in such a way that they serve the needs and purposes of the insect as a whole. According to Wigglesworth (1953), \"stimuli which the insect can learn to associate wiihi the presence of food, or with the location of i t s nest, are judged to be perceptible. Such experiments, of course, need the greatest care in their interpretation; for the insect may be guided by some other stimulus that has been overlooked by the experimenter; indeed, in the course of a single expe-riment, i t may cease to be oriented by one stimulus and come to depend upon another; and there are many who hold that the whole perceptual experience of the organism i s integrated, as i t were, into a pattern, and not to the isolated stimuli of which i t is composed.'* While certain aspects of specific responses, or the aggregate behaviour of a species may be recognized as being peculiar to i t in various ways, the component responses to internal (physiological) cues and external stimuli may show considerable fl e x i b i l i t y as a result of modifying accessory factors. These factors have to be considered. 1. Increasing temperature; its provision and use in the test3.^-Earlier observations (Graham, 1959) indicated that photic reversal of Trypodendron occurred when the temperature was in the region of 55° to 58°C but i t was desired to evaluate the effect in a more quantitative way. Several methods of test were employed. The fi r s t method made use of a glass tube, 2.0 cm. in diameter and 85 cm. in length, stoppered at the ends, placed in a temperature gradient apparatus (Pig. 4). The gradient appa-rat us consisted of a long shallow copper trough, to the under surface of which were soldered two lengths of copper tubing, one at either end of the trough and not intercommunicating. The tubing was folded in a zig-zag manner to give maximum contact with the trough. Hot water could be passed through one tube and cold through the other. Flow of the hot and cold water respectively was regulated to produce a temperature gradient from 15°C at one end and 40°0 at the other. The glass tube into which the beetles 28 Fig. 4. Apparatus used for testing responses to light in a temperature gradient. 29 were placed was coated with a transparent red varnish, except for annular clear zones of about 2.0 cm. in width spaced about 15.0 cm. apart. The red zones were provided to create conditions which would constitute darkness to the insects, yet would enable the observer to see through. A strip of moist tissue paper was placed inside the glass tube to provide a crawling surface for the beetles. The beetles were put in groups of five, sexes noted, at the colder end of the tube; then, stoppered. A beam of white light was projected successively into each clear zone for five minutes at a time beginning at the colder end. The number of beetles aggregating at the lighted zone was recorded in relation to the temperature at the particular point in the gradient. Another arrangement of apparatus was designed to relate the effect of temperature and of gravity to the photic responses of the beetle. This consisted of a water bath apparatus and a wide-mouth glass jar whose outer surface was painted flat black, except for two clear areas, each about three centimeters square. One of these clear areas was about two centimeters below the rim of the mouth of the jar. The other area was at the opposite side, near the base. A thermometer was fitted through the black rubber stopper and strips of moistened tissue paper, about 2.0 cm. in width and 11.0 cm. in length, were attached to the bottom surface of the rubber stopper so that when the jar. was stoppered, the tip of the thermometer was halfway down the jar and the strips of paper touched the interior bottom surface. The apparatus i s shown in Fig. 5. Groups of five beetles, sexes noted, were put inside the jar. After being stoppered, the jar was immersed in a water bath. The water bath apparatus consisted of a cylindrical glass vessel containing warm 29 A Fig. 5. Apparatus and set-up for testing responses of Trypodendron in relation to gravity and increasing temperature. 50 water whose temperature was varied from 15° to 40°G. A beam of white light was projected into the upper clear window so that the beetles had to climb or crawl up the paper strips (against gravity) to react to light. Once the number of the beetles responding positively to light had been recorded, the beam of white light was moved to the lower window. The beetles' movement would then be toward the force of gravity as they responded positively to light. The test was performed for every five-degree rise i n temperature until 40°©. 2. Airstream with or without wood odour; its provision and use in the teats.-Anemotaxis is the term applied to reactions or orientation in an air current (Praenkel and Gunn, 1961). Many species of insects, on taking to flight, head into the wind. Drosophila f l i e s are known to walk and orien-tate very accurately into a wind containing the smell of mashed frui t . In Trypodendron beetles, the typical and characteristic flight orientation i s against the wind (Chapman, 1962). Praenkel and Gunn (1961) state that i t is probable that many olfactory reactions are possible only with the air of air currents and that the means of reaching the source of odour i s to go against the wind or air current. To test for the anemotactic and anemo-olfactory effects on the photic reaction of the beetles, a gentle stream of air flowing at 0.5 meter per second (as measured by a \"Wallac-Thermex GGA2C\" thermo-anemometer), f i r s t without bearing wood odour, and then with odour in i t , was directed to the beetles, i t s sex noted, as i t was being tested for its reaction to light. The odour was provided by passing the airstream through a bottle containing either a mass of shavings from attractive wood or sawdust from unattractive \"green\" wood or various combinations of attractive shavings and \"green\" sawdust respectively. 51 Another teat concerned the effect of air streaming in an odour-f i l l e d environment. The purpose was to test for the operation of aneap-tactic response that is prompted by the host odour. Accordingly, the beetles were brought into an odour fi e l d by placing them on perforated paper over attractive wood and exposing them to a stream of either non-odorous or wood-odour-carrying air flowing in the direction of the light source. TJests of response to non-odorous and odour-bearing airstreams ware also made with beetles placed inside a dark box. Non-flown as well as flown (flight-experienced) beetles were tested. 5. Increasing atmospheric pressure: its provision and use in the tests.-Experiments were devised to test for one of the possible mechanisms by which the air bubble in the ventriculus of the flight-experienced beetles (Graham, 1961) may function in the modification of the photic response. One possibility is that the gas bubble functions by crowding the internal space, thereby creating haemostatic pressure; which i n turn relays the effect as a signal to a pressure receptor centre. Another possibility i s that a signal is received more directly by stretch receptors in the wall of the ventri-culus in which the bubble occurs. A third possibility i s that the crowding of internal space may stimulate stretch receptors in the body wall. In the absence of morphological evidence for stretch or pressure receptors in Trypodendron, the question must be approached experimentally. In experiments with spruce budworm larvae, Wellington (1948) demonstrated that photic reversal in flaccid individuals could be brought about by such turgor-causing factors as feeding, injection of fluid into the gut or into the haemolymph, or the imposition of external pressure by means of ligaturing. It would be of both academic and practical experimental interest to know whether the olfactory and other responses of Trypodendron can be studied while keeping the insect under pressure, in lie u of the other-wise necessary conditioning flight (Graham, 1961). It was assumed that i f pressure, and not: stretohv i s the mechanism involved in the change of photic response of Trypodendron, its effect could be made manifest in beetles placed under elevated! atmospheric pressure. A compression chamber was constructed consisting of \"Lucite\" plastic tubing, 40.0 cm. long and 5.0 cm. in outside diameter and with a wall thiokness of about 6.0 mm. It was fitted with a hose connection leading to a 4-way connector. The other passages in the connector were Joined respectively to a pressure gauge, an air bleeder valve, and a nitrogen tank. Nitrogen was used as the compressed gas, on assumption that the normal quantity of atmospheric oxygen -in the chamber was s t i l l available to the insects, whereas compressed air would have increased concentration of oxygen. This assumption follows that of Dalton's Law of partial pressure of gas mixtures. In order to judge the effect of pressure on the photic response, i t was necessary, in accordance with other findings of the investigation, to provide attractive host tree material as a source of an opposing stimulus to the photic one. Accordingly, a small sample of attractive wood was placed in one end of the compression chamber, non-flown beetles were introduced, the chamber sealed at the ends and placed in the retaining steel \"cradle11. Pres-sure was built up to a maximum of 50 psi (about two atmospheres). A beam of heat-filtered light was projected into the apparatus to cause the beetles to accumulate at the odour source, then the light was moved to the opposite end to compete with the odour for response of the beetles. 55 The apparatus for testing the response of the beetles to light under increasing atmospheric pressure is shown in Fig. 6. Methods of Observation The broadest problem of photic behaviour of Trypodendron may be considered.to relate to the nature and strength of orientation and other responses under various conditions of light, alone, or in combination with other factors. The consideration should include the difference in sex and kind of conditioning of the insect. For the study of behaviour, i t was necessary to choose appropriate criteria by which responses could be interpreted. The responses of insects to effective external stimuli are variously manifest in changes of the state of activity or inactivity and in their spatial and temporal patterns. Mani-festations of responses to stimuli are observable in movements of appendages or of regions of the body and displacement of the animals as a whole. Trypo-dendron i s capable of slight movement of its head in a vertical plane, of extending, waving, and folding back its antennae, of manipulating its mouth-parts for chewing or ingesting food, of waving its legs randomly i f the beetie is held aloft, of coordinating them in walking on a surface, or of folding them against the body in feigned death, of elevating the elytra and unfolding its membranous wings, of restoring them to their resting position, of vibra-ting elytra and wings in flight movements, and of manipulating the external genitalia as occasion befits. Variables in these, conceivably include the dimensions of time, space and force, giving rise to such derived values of displacement of parts, or of the whole insect, such as rate of walking or of wing beat of flight, and various degrees of consistency i n direction or activity in these. The variables would give rise also to such forms of 33 A Pig. 6. Apparatus for testing photic responses of Trypodendron under increased atmospheric pressure. 54 measurement as rates of change, and periodicities in functions, change of orientation angle in respect to a direction stimulus, and observations on the assumption of postures such as feigned death or preparatory stance for flight, boring into wood, feeding and mating. Preliminary experience in handling the beetles suggested that certain of the foregoing manifestations of response to stimuli would be most meaningful of the single or integrated reactions of the insect to light, darkness, contact, pressure, gravity, air flow, temperature, and chemi-cal and physical factors in the host tree. The kinds of response that were sought included the beetles' alignment and directed movement in respeot to directional stimuli, turning activity, retention at an odour centre or other centres of stimuli, attempts to f l y or actual flight and the compound responses of the whole insect which are manifest in i t s attempt to bore into wood. Group responses were observed quantitatively in the aggregation of beetles when a number of them were exposed simultaneously to an experimen-tal situation as in a temperature gradient apparatus. The experiments were conducted in a darkened room. The primary purpose of the darkness in these studies was not with the intention of adapt-ing the insect to the condition of scotopic vision, which would involve the use of beam intensities far below the present photopic requirements, but to avoid extraneous sources of light that might interfere,with the normal response of the beetle to a given stimulus. The experiments were conducted in a series which started in May, 1964. The scheme of the programme of experiments conducted in these studies of the behaviour Trypodendron is presented in Fig. 7. 55 Pig» 7. Schema of the programme of experiments on the photic responses of Trypodendron lineatum in relation to various host wood factors and conditions of the environment o n O V E R W I N T E R E D A D U L T S TIDAE) o t F l o w n . I A f t e r F h g K t U . W H I T E L S 6 B T PUEB^HT 0 ' F l £ L D o r L I O M T J H O R S J S O N T A i . |/ B £ A W O P JUI G U P -j ^ S r a e f t , \" W o o d Pr&seyCF *fte'ct N^ t»»» Contact 8 i n s e c i in Contact j -j\"\" W t ' - l r k o u t A i r Sfrcocvr^ A i r S o u r c e A n i i p o c t a t t o L i g h t j -. loot/ 4 *Aitraci.\\y/&\" \\Noottv. (2) vertical black-and-white broad lines, (5) diagonal black-and-white broad lines, (4) a l l black, or (5) a l l white background pattern while the beetles were individually exposed to a horizontal beam of white light did not produce results which showed any marked preference or tendency by the beetles to.respond to any of the patterns. 6. Responses to white light under increasing temperature.-Tests of the photic responses of flight-inexperienced adult Trypodendron at temperatures ranging from 14° to 40°Q, with the beetles inside a painted glass tubing placed on a temperature gradient apparatus, indicated that the peak of activity characteriaed by excitability, positive phototaxis, and attempts to f l y occurred between 26° and 28°G. At 58°<3 the beetles ceased to become attracted to light entirely. The results of 10 tests, using 10 beetles in each test, are shown in table 5. In the tests for responses of the beetles to light, antipodal to gravity, under increasing temperature, the range of temperature under which the responses of the beetles were studied was from 25° to 40°C In these tests, the peak of activity of the beetles was noted at about 29°C. The maximum temperature which registered a positive photic response by the beetles was 36°G. In only one instance out of 10 tests did a beetle appear at the lighted window at 56°G. Mating occurred occasionally between 25° and 35°G. These pairs were indifferent to light. In the tests for responses of the beetles to light positioned sympodally with gravity, the range of temperature registering the highest 50 Pig. 14 (A and B). Representative paths of responses to light and to an attractive piece of wood under perforated area on sheet of paper by flight-inexperienced (indicated by thin line) as well as flight-experienced (indicated by thick line) Trypodendron lineatum. Direction of light Figure 14 A. Specimen; female beeile Ammeter at 4.0 amperes Light, heat filtered Duration of flight: 30 min. c Direction of light 51 Pig. 15 (A and B). Types of response to light and to an airstream without attractive wood odour by flight-inexperienced (indicated, by thin line) and flight-experienced Trypodendron beetle on a perforated sheet of paper above an attractive host wood. Direction of light Direction of light Figure 15 B. Specimen: Female beetle Ammeter\" at 4.U amperes Light, heat filtered Duration of flight: 15 min. Direction of air stream without wood odour 16. Representative paths of response to light by flight-inexperienced (thin line) and f light -experienced (thick line) adult Trypodendron beetle placed on a perforated sheet of paper above attractive host wood and subjected to a gentle stream of air with attractive wood odour. Direction of air stream with \"ripe\" wood odour 53 Fig. 17 (A and B) „ Representative paths of response to light by the flight-inexperienced (thin line) or by the flight-experienced (thick line) Trypodendron beetle in contact with a piece of attractive host wood. Tracings were made of paths as beetle wandered away from the wood0 Figure 17 A. Specimen: female beetle Ammeter at 4.0 amperes Light, heat filtered Duration of flight; 5 hours Direction of light Figure 17 B. Specimen: male beetle Ammeter at 4.0 amperes. Light, heat filtered Duration of flight: 1.0 hour 54 Pig„ 18 (A. and B)« Representative paths of response of flight-inexperienced (indicated by thin broken line) and of flight^sxperienced (indicated by thick broken line) adult Trypodendron beetle to light and to a stream of air with attractive wood odour 0 Beetle ms in contact with wood and i t s path was traced whenever i t left the wood. Direction of light I I \\ \\ \\ 1 v \\ / y / .W/ J2—i. \"Ripe?., wood '\",r-^ ( V \\ Figure 18 A. Specimen: female beetle Ammeter at 4 . 0 amperes Light, heat filtered Duration of flight: 5 hours Direction of air stream with attractive wood odour ion of light Figure 18 B. Specimen: male beetle Ammeter at 4.0 amperes Light, heat filtered IXu-atioa of flight: 40 min. S e c t i o n of ^ 19« Representative paths of response of the flight-experienced Trypodendron beetle placed in contact with a piece of attrac-tive wood in relation to light in the presence of an airstream without (indicated by dotted line) or with attractive host wood odour (indicated by dashed line). Direction of light Figure 19 Specimen: female beetle Ammeter at 4.0 amperes Light, heat filtered Duration of flight: 45 min. s response to air without odour. - - - = response to air with attractive wood odour 4 * . i \\ I : i : I Direction of 56 percentage of beetles responding to light was 25° to 28°C. The maximum temperature registering the least number of beetles responding positively to light was S6°C. In some tests, increasing temperature caused the beetles to become very active or \"excited\". Attempts to f l y were noted i n many instances. Thermal paralysis and death of several beetles occurred between 56° and 58°C. The summarized data on the responses of the beetles to light in relation to gravity and increasing temperatures are shown in table 4. They are also illustrated in Pigs. 20 and 21. Table 5. Photic responses of adult Trypodendron under increasing temperature in a glass tubing on a gradient apparatus. Temperature range Per cent of beetles responding G positively to light 14 - 16 55 17 - 19 75 20 - 22 83 25 - 25 83 26 - 28 85 29 - 51 61 32 - 54 52 35 - 57 56 58-40 ' 0 57 Table 4. Photic responses of adult Trypodendron as influenced by gravity and increasing temperature M Temperature range Per cent of beetles responding positively to light G a. Light antipodal : b. Light sympodal to gravity with gravity 25-25 81 85 26 - 28 85 85 29 - 51 ,86 70 52 - 54 58 44 55 - 57 18 17 58-40 0 0 1/ Average of 10 tests per temperature rangej 10 beetles per test.; 7. Responses of Trypodendron to various stimuli in the dark.-Under normal laboratory conditions, newly overwintered Trypodendron adults, having flight experience or not, moved about in a random manner when placed inside a dark box. The same results were obtained when a stream of air was allowed in and out of the box through tiny holes. However, when the airstream carried attractive wood odour, the beetles tended to approach the hole through which the air stream was introduced. Physical contact with a piece of \"green\" wood placed inside the box did not cause an aggregation or positive response of the beetles. The majority of the beetles had moved away from the piece of wood after the allot-ted time of five minutes allowed for each test. Again the introduction of an airstream without wood odour did not change the i n i t i a l behaviour of the beetles. Positive response, however, was observed when the airstream carried attractive wood odour. Fig. 20. Effect of increasing temperature on the photic responses of Trypodendron lineatum (Olivier). Fig. 21. Comparison of effects of gravity and increasing temperature on the photic responses of Trypodendron lineatum (Olivier). Temper at tare range in degree centigrade 5 9 Attractive wood inside the box caused an aggregation of the beetles on or near the wood. During the period of observation, some of the beetles; crawled under the wood while others were either away from i t or at the base of contact of the wood with the sheet of paper. No boring behaviour was observed during the period allowed for each test. However, in a test in which the beetles were oonf ined to a piece of wood covering i t with a suitable stacking dish and left overnight, preliminary borings were observed. DISCUSSION QP RESULTS 1. Photic behaviour in Trypodendron; i t s ecological significance and i t s underlying mechanisms.-The response of Trypodendron adults to light at the end of their overwintering period has been shown by these studies and by previous other observations (Graham, 1959) to be strongly photopositive. Furthermore, the tests have shown that the beetles' ability to orientate to the source of light i s in no way affected by their i n i t i a l angular displacement. The expression of positive phototaxis appears essential at this particular stage of the l i f e cycle of the beetles. After overwintering, residence in the forest l i t t e r or loose bark not only ceases to function advantageously for the insect, but would frustrate its survival i f special sequences in beha-viour did not cause i t to disperse and react to host factors* The presence of a narrow horizontal beam of light attracts Trypo-dendron. „ Positive response is often characterized by an increase in the locomotory speed toward the source of light. At some point in the expression of positive phototaxis or at a certain distance from the source of light, the beetles may be stimulated to f l y . Another condition which stimulates flight is the presence of a broad overhead f i e l d of light as provided by ceiling lamps or by skylight. The importance of an overhead source of light i s shown by the fact that when the beetles were, exposed to light coming from below their horizontal plane of sight, flight did not occur * Restoration of an overhead source of light caused initiation of flights. In the laboratory, escaped beetles were observed flying continuously about the ceiling lights, at times even when their heads in touch with the fluorescent light tubes. The light intensity close to the light tube was 60 61 of the order of 500 footcandles. In nature, ordinary sunlight produces an iiTuminance of some 10,000 to 14,000 footcandles (Piatt and Griffiths, 1964). Under f i e l d conditions, the beetles do not f l y upward into the sky. At most/ they have been observed flying up to about six to seven meters from the ground i n forest areas with an elevation of about 400 meters above sea level (Chapman, 1962). One conceivable explanation for the beetles* not f l y -ing toward the sun may be that they f l y , like many other insects, at a certain angle to the rays of light. This type of light reaction is termed light compass reaction (Fraenkel and Gunn, 1961). It i s also possible that there is an upper limit in the threshold of light intensity which the photoreceptors of the beetles can tolerate. The occasional negative phototactic response among positively phototactic beetles has been observed (see table l ) . Why i t should exist among flight-inexperienced beetles which generally are positive in their light responses remains among other questions to be answered. It is conceiva-ble that in these individuals, diapause influences are s t i l l partly operative, or reproductive maturity is delayed. Positive phototaxis in Trypodendron is not a unique behaviour. Photic attraction is involved in various activities among different insects. Fireflies, for instance, though nocturnal in activity also exhibit positive phototaxis in attracting or locating a mate (KcElroy, 1964). In the moths, however, i t is s t i l l unexplained as to why these nocturnal insects are attrac-ted to light at night but not to sunlight in the day. Winged termites, on the other hand, utilize their brief positive phototactic behaviour as a means to disperse and possibly interbreed with members from different termite colonies of the same species. Little i s known directly of the mechanisms which trigger light attraction in Trypodendron although much has already been studied and written about photoreception in many species of insects (Birukow, 1961; Booth, 1965j Burtt and Catton, 1956, 1962; Grescitelli and Jahn, 1959, 1942; Dethier, 1955d; Fingerman, 1952; Goldsmith, 1964; Rogers, 1962; Ruck, 1965; de Vries, 1956; de Wilde, 1962). One of the factors known to influence the expression of phototaxis in some insects i s hormone concentration. Beetsma et al (1962) found that injections of an extract of the abdomen of male cecropia moth into fourth and f i f t h instar larvae of a hawk moth shifted the balance between photopositive and photonegative tendencies towards a definite photopositive response. In the Colorado potato beetle, activity of the corpora allata has been correlated with positive phototaxis (de Wilde, 1959) . Photonegative response was also demonstrated by implantation of active prothoracic glands into the body cavity of the caterpillars. It i s conceivable that in some of the newly overwintered Trypodendron adults the prothoracic gland s t i l l exerts an inhibiting influence on the photic beha-viour of the beetle. In some other insects, light attraction is tied with homeostatic processes which regulate the physiological and biochemical functions of the various internal organs. Thus the larvae of several species of tent cater-pillars, Malacosoma distria, M. pluviale and of the f a l l webworm Hyphantria textor on becoming hungry are attracted to light and crawl up to the top branches where there is young foliage to feed on (Sullivan and Wellington, 1955; Wellington et al, 1954). A similar light attraction enables the adults of white pine weevil, Pissodes sbrobi, to climb to the terminal shoots or leaders which they infest (Sullivan, 1959). These observations suggest possible approaches to the study of physiological mechanisms in photic responses in Trypodendron. One important observation on the behaviour of Trypodendron that needs derivation from known physiological characteristics of organ systems in insects i s the mechanisms which stimulate the beetle to f l y . It appears that the amount and intensity of light falling on i t s photoreceptors may be involved in the flight-initiating mechanisms. It i s also possible that the photoreceptors have a threshold number which triggers the flight response. Pielou (1940) believes that in the beetle Tenebrio molitor, there i s a relation between threshold of response and the number of sensiHa stimulated. The failure by a broad f i e l d of light coming from below the horizontal plane of sight of the beetles to stimulate the beetles to f l y seems to support the foregoing statement. Another explanatiork relevant to the beetle's failure to f l y in the presence of \"sunken\" light involves differences in the thresholds of stimulation or in the ability to discri-minate light between the dorsal and ventral part of the eyes of insects whose eyes are divided into two portions (Burkhardt, 1964). Trypodendron has divided eyes. 2. The modifiers of photic behaviour; their.-relation to host^fihding.^ The failure of a non-odour-bearing airstream to modify the activity of Trypodendron beetles moving toward the source of light indicates that the airstream alone has no inhibiting effect on the phototactic beha*r _ viour of the beetles, nor does i t e l i c i t a positive anemotactic response. This conforms with statements by Dethier (1957) in his review on the orien-tation by some flying insects that air current alone does not initiate 64 orientation. This statement was supported by observations made on the behaviour of diverse kinds of insects such as Drosophila, a f l y ; Geotrupes ^ - i. stercorarius, a beetle; and Bombyx mori, a moth. ,In these insects, i t is odour which initiates orientation to air currents. In our own studies, pedestrian Trypodendron adults responded to an airstream carrying attractive wood odour. Their positive phototactic behaviour was immediately inhibited on their being exposed to an airstream carrying attractive wood odour. Their response, however, was variable in the duration of time they stayed at the odour source. The beetles approached the orifice of the air source but eventually left i t to proceed toward light. On the basis of this observation, one may postulate the need for other kinds of stimuli that must be present in order to retain the beetles at the odour source. The failure of the beetles to react to an airstream when they are already surrounded by odour appears at f i r s t to be contrary to expecta-tions. It does not appear to correlate with their ^ response to an odorous airstream brought into an odour-free \"field\". It also tends to contradict the suggestion that an anemotactic response occurs. It should be remembered, however, that these studies pertain to a pedestrian situation i n which the tactile stimulus of the insect in contact with a'surface may under certain conditions inhibit an anemotactic response. If the beetles reacted other-wise, they Would be biological failures, abandoning attractive host mate-r i a l every time a breeze blew across i t . At least two considerations might be discussed in connection with the foregoing statements: (l) conditions within the beetles and (2) condi-tions within the host materials. : 65 Granting that a l l conditions within the host material meet the requirements of the beetles, i t seems that the beetles s t i l l must go through a series of stereotyped responses starting with positive phototaxis, flight, orientation to an airstream carrying attractive wood odour, approach to the source of the odour, settling and crawling before being able to bore into the host material. Graham (1959) indicated that flight exercise could effect an inhibition of the phototactic response of the beetles. Flight-exercised beetles were no longer strongly photopositive, and when put in contact with an attractive pieoe of wood, the beetles behaved normally by exploring the bark contours and eventually boring into wood. While the present study did confirm that flight and the presence of attractive wood inhibit the positive phototactic behaviour of Trypoden- dron, i t did not, however, show such strong depression of the phototactic reaction, nor the strong tendency to bore into attractive wood. The reason for the disparity of results i s not clear. It may have to be sought i n differences in the population stock, conditions of storage or unrecognized differences in the environmental conditions during the experiments. There appears to be present in the behaviour pattern of Trypo-dendron another mechanism of responses leading to the boring into wood. For instance, Graham (1959) observed that beetles which had no previous flight experience when kept in the dark with a piece of attractive wood exhibited the boring behaviour „ Chapman (1959) and Dyer and Chapman (1965) reported that newly overwintered,,flight-inexperienced beetles could be forced to bore into attractive logs but not into \"green\"/logs when confined to the upper surface of logs in small aluminum rings partly imbedded in the 66 surface of the logs. While Graham's observations noted borings made in the dark, Chapman and Dyer's did not note whether the borings were made during the day or during the night. Neither, however, excluded the possible effect of locomotion by prolonged walking or crawling on responsiveness to light or to its host as observed by Johnson (1958) in connection with the settling responses in aphids. The importance of flight experience preparatory to host attraction under normal conditions has been noted in insects other than Trypodendron. In aphids, Kennedy (1955, 1958) and Kennedy and Booth (1965a, 1965b) observed that flight experience \"primes\" or promotes the settling response on a host leaf. A suitable host leaf itself was considered to possess an inhibitory stimulus that would cause the aphid to remain. If the leaf were unsuitable, the insect would take to flight again. The \"excitability of* the.;settling responses increased as an aftereffect,of flight in whioh the excitability of flight itself (measured by the rate of climb) was not falling and was even increasing.\" As regards the effect of flight on phototaacis, Kennedy (1965) believes that, in aphids, \"at first.the phototaacis is positive to a l l light intensities, but that as flight proceeds i t becomes negative to bright light while s t i l l remaining positive to weaker light, until finally i t becomes negative even to the dimmest light and the reversal can then be said to be complete.\" Several suggestions have been advanced to explain the underlying mechanisms which cause the settling and feeding behaviour as an aftereffect of flight. ,Evans and ^ethier (1957) and Hudson (1958) are of the opinion that flight, especially in the blowfly and the honeybee, causes a diminution 67 of blood trehalose and glucose levels resulting in the lowering of the thres-hold for feeding. Johnson (1958) also suggests that respiratory metabolism has an effect on the settling responses as he found that aphids increased readiness to settle after a brief anaesthesia with carbon dioxide but not' with ether. These suggestions appear to come under the \"peripheral1* hypo-theses which, according to Kennedy and Booth (1963b), are \"versions of the chain-reflex theory of behaviour assuming that the link lies not in the central nervous system but in some cumulative physiological consequence of the locomotor activity which provides 'feed-back' •\" Kennedy (1Q65) believes, however, that the mechanism lies in the cental nervous system and is controlled by i t , \"over and above any peripheral feed-back from flight such as air swallowing and gut distension seem to provide.\" As to the composition and make-up of the set of stimuli that must be present in the host material before a \"conditioned\" or \"primed\" insect could show a favorable response, Beck (1965) suggests that they are bioche-mical and biophysical in nature. He gives the following classification of stimuli which influence different feeding responses: Type of response , Evoking stimulus 1 Positive a f Negative Orientation Orientation Biting or piercing Maintenance of feeding Attractant Arrestant Incitant J Stimulant T I J | 1 Bepellent Repellent Suppresant Deterrent Beck defines an \"attractant\" as any stimulus which attracts the insect by orienting movement1 toward the source. An \"arrestant\" is said to cause an insect to cease locomotion in close contact with the source, while 68 an \"incitant\" i s a stimulus which evokes a biting or piercing response. The negative responses are caused by a set of stimuli, termed differently, which prevent or inhibit the positive responses. A considerable amount of work has been done on host-finding and selection by many phytophagous and parasitic insects (Beck, 1957$ 1960, 1965, 1965; Brues, 1946; Dethier, 1941, 1947a, 1947b, 1951, 1952a, 1952b, 1955a, 1953b, 1954, 1955, 1957, 1962, 1963; Dethier and Rhoades, 1954; Fraenkel, 1955, 1959; Ham&mura, 1959; Harris, 1960; Howe, 1950; Ibbotson and Kennedy, 1959; Kennedy, 1955, 1958; Kennedy and Booth, 1951, 1965a, 1965b, 1964; Kennedy et al, 1959a, 1959b, 1961; Idpke and Fraenkel, 1956; Loschiavo et a l , 1965; MacGregor, 1948; Merker, 1955; Ohnesorge, 1955; Painter, 1958, 1965; Thompson and Parker, 1927; Thorsteinson, 1955, 1958a, 1958b, 1960; de Wilde, 1958). In Trypodendron, neither the nature of the so-called \"incitant\" which would cause the beetle to begin boring, nor the other sets of stimuli in the series, nor their corresponding inhibitory counterparts are known. The effect of increasing temperature in the environment on the phototactic behaviour of various insects has always been that of reversal (Jack and Williams, 1937; Green, 1954; Pertunnen, 1959; Pertunnen and Palo-heimo, 1965, 1964; Rudinsky and Vite, 1956; Wellington, 1948; Wellington et al, 1951, 1954). The results of the tests on Trypodendron show that the beetles shun the light at temperatures above 58°C. The term \"photic reversal\" in the sense of a permanent change from photopositive to photonegative or vice versa as a result of high temperature may not be an applicable term for Trypodendron. Photic inhibition may be a more appropriate term because the beetle ..does not really become negatively 69 phototactic as indicated by the fact that they s t i l l respond positively to light when returned to ordinary temperatures. The apparent effect of high temperature under f i e l d conditions on the photic response of insects may not actually be a reversal of phototaxis but a different response which prompts the insects to escape from heat which is correlated with the most intensely illuminated condition. Should, however, the temperature in the shaded area be higher than or equal to that in the exposed area, the insect would likely, as does Trypodendron move about, even venturing to light, in search of a favorable temperature. Photic reversal seems to persists, as i s manifest in flown ter-mites or in Trypodendron as i t begins to bore a tunnel into wood, or i n the last instar larva of the hawk moth as i t digs into the ground in order-to pupate (Gilbert, 1964). Positive geotaxis i s often correlated with negative phototaxis, especially in many plant-climbing insects. This is particularly true of the larvae of lepidopterous and of hymenopterous insects. However, many of the non-plant-climbing insects may become photonegative in their response to light without becoming positively geotactic. This is indicated by the photonegative responses of the beetles at high temperatures (up to 57°C) when light was sympodal with gravity (see table 4). The failure of the elevated atmospheric pressure to simulate the effect of the swallowed gas bubble i n reducing the photic response showed that pressure per se is not involved. One must therefore contemplate the possibility that stretch effects on the gut wall or on the body wall are involved (Pinlayson and Lowenstein, 1955). 70 Insects are generally classified as either diurnal, crepuscular, or nocturnal in habits depending on the time of the day they are most active. While Trypodendron is definitely diurnal in habits, expecially in so far as emergence and flight are concerned, they are nevertheless active in the absence of light. The activities of these beetles in the dark include copulation, orientation by crawling toward and attractive odour source, boring into a suitable piece of wood, feeding and oviposition. While the behaviour of the beetles in the dark in relation to a suitable piece of wood or to:an attractive'bdour is similar to the beha-viour of the.flight-experienced beetle ;ih\\the:.presence:rbfilight^tha intrin-sic motivating stimulus acting on the beetle in the dark may be entirely different from the \"priming\" effect of flight. As already reported, flight results in the expenditure of energy by the insect (Pringle, 1965j Sacktor, 1965), accumulation of a gas bubble in the ventriculus (Graham, 1961), and consequent physiological and/or physical effects (Chapman, 1956) responsible for lowering some thresholds which permit the beetle to settle on and bore into a suitable piece of wood. What \"primes\" the beetle i n the dark is not known. It appears that the physiological, biochemical, and physical changes brought about by flight are too much for the flight-inexperienced\" beetle to attain or obtain in the dark. These observations suggest that there must be other mechanisms involved and that the importance of the central nervous system as stipulated by Kennedy (1965) in governing the behaviour of the beetles is not to be overlooked. The retention of beetles on perforated surfaces separated from attractive wood by a shallow space, as well as their ready response to a stream of air passing over attractive' wood but not over green wood can be 71 explained only on the assumption that odour is the prime factor involved* The similarity of responses of males and females to the odour from attractive hut non-attacked -wood demonstrates the existence of an effective primary attractant originating from the wood alone. This is not to say, however, that secondary attractants may not play an important role in increas-ing subsequent frequency of attack (Rudinsky and Daterman, 1964a, 1964b) • The retention of the flown beetles in a f i e l d of odour above attractive wood is characterized by their decisive return to the f i e l d of odour after transgressing the boundary of that f i e l d into an odourless; area (see Pig. 15). This reaction suggests either a memory for position, or a direction-reversal when the olfactory stimulus diminishes or f a i l s , or an ability to detect a gradient within a very short radius. Dethier (1957) mentions that in Bombyx mori a strong drop in odour concentration while the insect is responding to an odour stream results in elimination of stream orientation. The continued photopositive response of T. lineatum even when placed in contact with, green wood, or in a f i e l d of odour from i t ^ or in a directed stream of odour from i t allows several possible inferences. Either attractants are not present in stimulating concentrations or they are present but are obscured or rendered ineffective by repellents or olfactory inhibitors. Bow the inhibition of the photic response of flight-experienced beetles when brought into contact with attractive wood or odour from i t may signify the formation de novo of olfactory attractants from non-attractive or even from repellent precursors, or i t may signify the disappearance of repellents which prohibit the action of pre-formed attrac-tants, or i t may mean that repellents are in i t i a l l y present which convert 72 to attractants. The questions are largely resolved by consideration of the fact that olfactory attraction of attractive wood was retained even when i t was admixed with a large proportion of \"green\" wood. This indicates that i f repellents or olfactory inhibitors exist, they are not present in significant quantities in \"green\" wood. This being so, one may then conclude that attrac-tants are not present in \"green\" wood, for i f they were, their effect would have been manifest because there is evidently nothing in \"green\" wood to prohibit any attractants in i t from eliciting a response. 5. Aspects of behaviour of Trypodendron which are useful in bioassay techniques.-Previous investigations on attractancy of materials for T. lineatum depended on the beetle boring into wood. The technique consisted of placing female beetles in darkness on either bark-covered slabs of wood or half-inch thick transverse sections of wood which were treated in various.ways with test chemicals. It may be assumed that a boring-in response depends not only upon attractancy for the olfactory sense, but also upon factors of texture and taste. The boring-in test is therefore not ideally suited for detecting purely olfac-tory stimuli. It has other disadvantages. It is suitable only for female beetles, since i t is only the females of this species which initiate and carry out tunnelling. It is therefore costly of potential experimental specimens. Furthermore, for statistical reasons, each t r i a l should be based on the propor-tional responses of at least 10 individuals introduced simultaneously. Another disadvantage is that there is a considerable time lag in obtaining a decisive indication of response, and this interval of lag varies considerably between individuals. A period of about 24 hours is often required before the responses of the beetles can be decided. During this interval, pronounced chemical changes 73 may occur i n the material being tested. I f texture, taste, and other factors as yet not understood, are unsatisfactory, olfactory attractants may not y i e l d a response. On the other hand, the test has the . advantage of not requiring close and constant observation of the beetles. I t has a further advantage i n that the holes bored by the beetles provide a permanent record of t h e i r response. This type of test can be set up with non-flown females confined i n the dark with the test material. This bypassing of the need f o r f l i g h t that would be required f o r study of responses i n the l i g h t s i m p l i f i e s the procedure and ensures more uniformity of the test population of beetles. The present researches have opened up two new p o s s i b i l i t i e s of experi-mental situations f o r t e s t i n g olfactory attractants f o r T. lineatum. One depends upon the c e n t r i p e t a l response of beetles i n a f i e l d of odour. The other depends upon anemo-olfactory orientation and locomotion of beetles i n an odour-laden airstream. Both have the advantage of a b r i e f e r period of observation than the boring-in response. The c e n t r i p e t a l response i s the one which i s characterized by the beetles repeatedly r e t r i e v i n g t h e i r p o s i t i o n within a f i e l d of odour orig i n a t i n g i n the substratum, This reaction would serve the purposes of a bioassay tech-nique i n which unknowns are introduced into the substratum. The length of the meandering path and/or the duration of contact with the odour f i e l d would be compared with values obtained with the insect i n an odour-free f i e l d . This technique would have the economical feature of permitting the u t i l i z a t i o n of both sexes f o r studies of the primary host attractant. I t also would y i e l d information from specimens studied i n d i v i d u a l l y . Tracings of the path t r a v e l l e d by a beetle and/or the duration of contact with the odour f i e l d provide one possible quantitative measure of attractiveness. This study must be car r i e d 73 A out either with flight-experienced beetles illuminated with a horizontal beam of light, or with non-flown beetles observed under a dark red light to which they are insensitive. The anemo-olfactory response would provide an immediate measure of attractiveness. A test would depend upon directing an odour-laden airstream toward flown or flight-experienced beetles in the presence of an opposing illumination source, or toward non-flown beetles under a dark red light. The method would permit the utilization of both sexes when primary host attrac-tants are to be studied. This method would be the least time-consuming of a l l . It is conceivable that with this type of test, the manipulation of intensity of the opposing light source would provide a means for quantitative expression of olfactory attraction. Strength of olfactory attraction would be measured in terms of the strength of light necessary to inhibit the olfactory influence. In practice, however, the test might consist of the converse manipulation in which the odour is attenuated while the light is kept constant. CONCLUSIONS The present researches have cleared away some of the primary obsta-cles to a comprehensive understanding of the behaviour of T. neatum (Oliv.). In turn, the patterns of its behaviour assume importance in terms of i t s biology and ecology. These studies have also removed the primary obstacles to the use of the insect as an instrument for bioassaying the attractant nature of chemical substances isolated from the host trees. This laying of foundations for bioassaying techniques also opens up opportunities for the study of factors affecting the formation of attractants in wood in as much as tests for attractancy wi l l no longer depend on the beetles boring into intact wood. Now i t should be possible to experiment with wood that is in a mechanically disintegrated state, which i s more amenable to chemical procedures. The following conclusions pertain to the pedestrian situation for the beetles in their non-diapausing conditipn as they occur in spring, prepared for normal emergence, flight and attack. Males and females were similar in responses except in those situations which involve boring into wood, as only the females perform this operation. 1. The beetles, before flight, are strongly photopositive at tempe-ratures in the range of 5° to 56°C. Positive phototactic response is inhi-bited in an increasing percentage of individuals at temperatures above 36°. At 58°C, positive response to light by the beetles ceases to exist. 2. The non-flown beetles react quickly to angular deviations of alignment in respect to a light source and turn with almost equal readiness toward the light regardless of its angle. 3. The beetles' responses to light, before flight, may be classi-fied as follows: l) not inhibited- the beetles immediately move or even 74 75 run toward the source of light; 2) i n i t i a l l y inhibited positive response^ the beetles may head away from light at the start of movement, but immediately turns about and move toward the source of light; 5.) inhibited positive res-ponse r the beetles may or may not at the start of movement be inhibited by light but the general direction of movement toward the source of light i s oblique; and 4) completely inhibited responser the beetles move with no apparent response to light. 4 . The photopositive response largely masks the other potential capabilities of the non-flown beetles. A photopositive response is retained regardless of the relative direction of light in respect to the earth's gravitational field. Its effect is retained, albeit delayed, despite proxi-mity of the insect to a host odour souroe. 5. Exclusion of the photic stimulus from flight-inexperienced beetles unmasks their capabilities of responding to host odour in an odour field. 6 . Flight experience partially modifies the simple photic reaction of some individuals in the absence of host odour, but the majority remain unchanged. 7. Flight experience prepares the beetles for a response to host factors in the presence of light. As brief a flight as five minutes has produced this effect in some individuals, '/ 8. A response to odour from attractive wood becomes manifest under various conditions: (a) Non-flown beetles display a response in darkness in the form of a \"fluid\" aggregation of part of a population free to move in an area containing an odour fi e l d caused by either the presence of a piece of _ ,. ...... . 76 attractive wood or the introduction of a stream of air carrying attractive wood odour. (b) Plight-experienced beetles display a positive olfactory response under white light by lingering for an extended period of time over an odour f i e l d in opposition to a beam of light. (c) Plight-experienced beetles display a positive olfactory response under white light by heading upstream in a current of air carrying that odour in opposition to a beam of light. (d) Hon-flown beetles display an inhibited olfactory response under white light but exhibit a positive olfactory response in darkness. 9. Hon-flown and flight-experienced beetles take to flight in the presence of a broad overhead f i e l d of light. They do not take to flight when light arrives from below the horizontal plane of vision of the beetles but they do so in the presence of a combination of lights above and below. 10. As pedestrians, the beetles do not respond to patterns of black-and-white lines in their immediate background as they do to a source of attractive wood odour. This fact implies that vision i s not employed as much as olfaction in host-finding. 11. Non-flown beetles under pressure of 30 psi do not respond to host odour while exposed to white light. The implication i s that the presence of gas bubble in the ventriculus accumulated during flight does not produce the change of photic response in the presence of odour by a pressure effect. This conclusion leaves the possibility of its acting by stimulating stretch receptors i n the gut wall. It also denies the experimenter this condition for studying olfactory responses in non-flown beetles. 77 12. When odour already surrounds a beetle, the latter generally does not react to an airstream. Some of the beetles which behave otherwise may not have the threshold of response sufficiently lowered. 15. The responses of this insect to the conditions studied here explain some of the important mechanisms of its ecology. The i n i t i a l strong positive response to light sponsors dispersal of the adult population in spring and empowers the insects with the capacity for territorial coverage. 14. The behaviour of Trypodendron i s not strictly stereotyped in the sense that i t follows a definite pattern. While flight, for instance, may be a normal conditioning or priming mechanism for the alighting or olfactory response, response to odour sources and the subsequent boring behaviour may be accomplished, in the absence of previous flight experience, in the dark. 15. The response of the flight-experienced beetle to an odour-bearing airstream probably provides a simple mechanism for host discovery, in which the host odour may act merely as a releaser of an anemotactic res-ponse. 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Additional conditions apply, see Terms of Use https://open.library.ubc.ca/terms_of_use."@en ; ns0:scholarLevel "Graduate"@en ; dcterms:title "Studies of some aspects of behaviour in the ambrosia beetle, Trypodendron lineatum (Olivier)"@en ; dcterms:type "Text"@en ; ns0:identifierURI "http://hdl.handle.net/2429/38385"@en .