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The incidence of human trichinosis in the Vancouver area as determined by the examination of four hundred… Bourns, Thomas Kenneth Richard 1949

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L  (If  THE  INCIDENCE OF HUMAN TRICHINOSIS IN THE  VANCOUVER AREA AS DETERMINED BY THE EXAMINATION OF FOUR HUNDRED DIAPHRAGMS by THOMAS KENNETH RICHARD BOURNS  A Thesis Submitted i n P a r t i a l Fulfilment of the Requirements for the Degree of MASTER OF ARTS i n the Department of ZOOLOGY"  THE  UNIVERSITY OF BRITISH COLUMBIA A p r i l 1949  ABSTRACT  Four hundred diaphragms from Vancouver h o s p i t a l s were examined f o r Trichinae "by direct compression and by digestion technique.  Sixteen, or four percent were found  to contain t r i c h i n a cysts.  By applying the formula  M i 3d , where M i s the mean, and a i s the standard deviat i o n , t h e o r e t i c a l range of the inoidenoe of t r i c h i n o s i s i n the entire population was and 6.98$.  determined to be between  1.03$  L i m i t a t i o n of the amount of tissue examined,  v i z . one gram by compression and ten grams by digestion, and l i m i t a t i o n s imposed by the fact that only diaphragm muscle was  examined, allowed an opportunity f o r certain  l i g h t infections to pass unnoticed.  It i s therefore con-  cluded that the incidence of human t r i c h i n o s i s . i n the Vancouver area i s probably somewhat above four percent. While no reeent infections were discovered, t h i s incidence, which i s over double that found i n Eastern Canada, and i n some oases the intensity of i n f e c t i o n , the highest being seventy cysts per gram of muscle, conclusively show that t r i c h i n o s i s constitutes a serious public health problem i n the Vancouver area.  TABLE OF CONTENTS  Acknowledgements  page  I  Introduction...  1  H i s t o r i c a l Review  3  Natural History  7  Methods  9 9  Basic Techniques Compression Method.  11  Digestion Method  12  Experiment designed t o disoover more s a t i s f a c t o r y d i g e s t i o n technique  13  Results  15  Results of the present survey  15  Results o f an attempt t o i n f e c t .Goldfish with T r l c h l n e l l a s p i r a l i s  ;  Discussion  16 17  P o s s i b l e sources o f e r r o r i n r e s u l t s  17  A n a l y s i s of data and case h i s t o r i e s  20  J  Comparison of l o c a l f i n d i n g s w i t h those obtained elsewhere  23  P o s s i b l e explanations  26  References Plates  f o r l o o a l incidence  Acknowle dgement a The author wishes to express h i s sincere gratitude to the many friends and advisors who m a t e r i a l l y to the survey.  have contributed so  In p a r t i c u l a r he wishes to  thank h i s Mother and Father for t h e i r counsel, encouragement, and f i n a n c i a l aid; who  Sr. J . R. Adams, h i s d i r e c t o r ,  has offered help and constructive c r i t i c i s m throughout  the course of the work;  Dr. W. A. Clemens whose door i s  never olosed to those seeking advice; who  Dr. I. 18 • Moynihan  has frequently offered material, f r i e n d l y encouragement,  and sound advice;  Dr. H. E. Taylor, Dr. H. H. P i t t s ,  Dr. H. K. P i d l e r , and Mr. P. J . Pish, f o r t h e i r  co-operation  and interest i n making available the necessary tissues and h o s p i t a l records; who  Mr. J . W. MacKay, an excellent f r i e n d ,  has oheerfully given of h i s time and knowledge i n pre-  paring the photographs; and Miss P a t r i c i a Bourns who  has  worked long hours typing i n order that t h i s paper might be f i n i s h e d on time.  - 1 -  The Inoidenoe o f Human T r i c h i n o s i s i n the Vancouver Area as Determined by the Examination of Four Hundred Diaphragms  Introduction T r i o h i n o s i s , a disease a f f e c t i n g man and other animals, i s caused by the p a r a s i t i c nematode T r l c h i n e l l a s p i r a l i s , which invades the s k e l e t a l muscle f i b r e s o f i t s host. While the vast m a j o r i t y o f human i n f e c t i o n s remain subc l i n i c a l and are not reoognlzed, the numerous epidemics and f a t a l oases on record d e a r l y i n d i c a t e that the very existance o f t r i c h i n o s i s i n a community c o n s t i t u t e s a serious medical problem. Human i n f e o t i o n i s g e n e r a l l y a t t r i b u t e d t o eating raw o r improperly cooked pork.  S i m i l a r l y , hogs are known  to become t r i o h i n o s e d by eating garbage containing i n f e c t e d pork scraps.  Control i s d i r e c t e d f i r s t of a l l toward  prevention o f hog i n f e o t i o n , and secondly t o treatment of pork destined f o r raw consumption.  Since heat i s known t o  k i l l t r i c h i n a l a r v a e , (Wright and Bozicdvieh 1943), the former type o f c o n t r o l i s exeroised i n Canada by a F e d e r a l law which decrees that a l l garbage used as hog s w i l l must  - 2 -  be cooked.  I t i s noteworthy that no such law exists i n  the U. S. A.  The second type of control, v i z . treatment  of pork to be used i n products normally eaten uncooked, i s governed by law i n both Canada and the U. S. A* (Cameron 1927).  In each case law decrees that pork  destined f o r raw consumption must be treated by heat, cold, or s a l t , i n suoh a way as to k i l l any larvae present.. F i n a l control rests with the cook.  For only pork  which has been oooked u n t i l no natural pink colour remains can be regarded as absolutely safe.  Unfortunately, the  average Canadian or American f e e l s w e l l protected by Government meat inspection.  What he does not r e a l i z e i s  that i n neither country i s there any inspection made f o r the only disease he can commonly contraot from pork, trichinosis. Up to the time of the present survey, nothing has been known of the incidence of human t r i c h i n o s i s i n Western Canada.  Theoretically we are adequately protected  i n our own country.  However we have had no proof that  our control measures have been conscientiously praotised. Furthermore, the c i t y of Vancouver i s located a bare t h i r t y miles from the United States border.  On the basis  of some twelve thousand necropsy examinations, i t has been estimated that one American In every s i x , or a t o t a l of twenty-one m i l l i o n Americans harbour Trichinae within t h e i r bodies.  S t o l l (1947) estimates that there i s three  -  2  -  times as much t r i c h i n o s i s i n the U. S. A* as there i s i n a l l the rest of the world put together*  S t o l l also states  that four and one h a l f per cent of the p o s i t i v e eases found, showed oaunts of f i f t y or more larvae per gram of muscle, i n f e c t i o n of an order " capable of causing pronounced c l i n i c a l symptoms."  5e further estimates that  suoh individuals, i f adults, would be carrying loads.in excess of one m i l l i o n larvae* Besides describing a serious medical problem i n the United States, the foregoing figures represent a tremendous number of meals of pork containing i n f e c t i v e t r i c h i n a e * It seems only reasonable then, to assume that a great number of i n f e c t i o u s meals await the Canadian v i s i t o r * The present survey has been c a r r i e d out to determine the adequacy of our controls, and whether or not t r i c h i nosis constitutes a medical problem i n the Vancouver area.  Historical The history of our knowledge of t r i c h i n o s i s follows a pattern whioh i s oommon i n b i o l o g i c a l progress.  It  began with u n o r i t i o a l awareness, was catapulted into s c i e n t i f i c focus at a single stroke, was  developed along  descriptive l i n e s , and at present i s being further developed p h y s i o l o g i c a l l y .  The fact that t r i c h i n o s i s i s  s t i l l very much i n s c i e n t i f i c focus i s bom  out by the  -  4  -  faot that at least nine surveys have been made at various points i n the l a s t three years, and that at least  one  major work has been published on the subject i n that time. The major work referred to i s a very complete book, t r i c h i n o s i s , by S.  E. o u l d , from which the following G  h i s t o r i c a l summary has been oompiled. The e a r l i e s t existence of T r i o h i n e l l a s p i r a l i s i s obscure.  Many writers however, f e e l that i t s a d a p t a b i l i t y  to a wide range of host species points to a long evolutionary h i s t o r y .  Indeed i t has been suggested that the  ancient Mosaio law which f o r b i d s the eating of swine f l e s h was  Instigated following a number of t r i c h i n o s i s epidemios. Probably the f i r s t recorded observation of Trichina  cysts i s that made by Tledman i n 1821,  i n whioh he described  seeing "Small oval shaped whitish stony concretions" i n the muscles of a cadaver whioh he was  studying.  Credit  for f i r s t observing the nematode i n i t s cyst i s given to S i r •Tames Paget, who  i n 1834  observed " l i t t l e specks i n  the muscles" of a cadaver, and who was examine them microscopically.  curious enough to  S i r Richard Owen, who  was  f a m i l i a r with Paget's work, read a paper before the Zoological Society of London i n 1835  i n whioh he named  the worm Trichina s p i r a l i s . The next important i n 1846, who This was  advanoe was made by Joseph Leidy  discovered Trichinae i n the muscles of a hog.  followed by the works of Herbst  (1851),  - 6-  Virohow (1859) and Leuckart (1860) i n which the anatomy of  the worm was more completely described, and the f a c t  was established that an animal eating triehinous meat may develope t r i c h i n o s i s . In 1860 Zenker discovered a heavy i n f e c t i o n of l i v e Trichinae i n the muscles of the body of a Dresden servant girl.  Subsequently he found adult Trichinae i n the  i n t e s t i n e , and i n f e o t i o n i n the ham and sausage at the house where the g i r l had worked.  Thus he postulated,  t r i c h i n o s i s can be f a t a l t o human beings, the complete l i f e history of the worm takes place i n one host, and human i n f e c t i o n can r e s u l t from eating infected pork. Furthermore he c o r r e c t l y surmised that the larvae were disseminated by way of the chyle ducts and the blood stream. Examination of pork f o r Trichinae was begun i n Germany as early as 1863, and l a r g e l y at the insistence of Virohow was made oompulsary by Prussian law i n 1879.  At the same  time pressure was exerted toward the banning of importation of American pork products. By 1881 most of the Central and Western European countries had such bans i n e f f e c t . In order to regain these markets, the U.* S. A. adopted a pork examination scheme i n 1898.  This was continued u n t i l  1906 when i t was abandoned. Since the name Trichina was already applied to a genus of Diptera, E a i l l i e t , i n 1896, changed the name proposed by Owen, v i z . Trichina s p i r a l i s to T r l o h l n e l l a  spiralis*  Around the same time considerable work was  being done on the l i f e h i s t o r y and diagnosis of the parasite.  In 1897 Brown noticed a high e o s i n o p h i l i a associated  with the disease, and suggested i t s diagnostic value. Many workers, notably F i e l d e r (1864), Cerfontaine (1895) and Askowazy (1895) made l i f e h i s t o r y studies and conc l u s i v e l y showed that the larvae were disseminated through the lymphatics and blood stream to the muscles* The f i r s t Triohina antigen was prepared by Str6bel i n 1911.  This has been improved upon by Bachman (1928),  Augustin and Theiler (1932), McCoy, M i l l e r and Friedlander (1933) and f i n a l l y by B o z i o i v i c h (1939).  In each case  the antigen has consisted of an extraction of dried larvae, the constant improvement being directed t o s p e c i f i c i t y of the antigen. As was previously stated, the common source of human i n f e c t i o n i s trichinosed pork.  The only other possible  source i s the meat of an edible game animal.  Sach a  p o s s i b i l i t y has been mentioned by Steffanson (1948) who reported that Trichinae are prevalent i n polar bears i n Frans Josef Land, where they are used as a food  souroe.  Surveys of animal populations have shown that most carnivorous and omnivorous animals may become Infected by Trichinae*  Gould states that the following animals have  been found n a t u r a l l y Infeoted;  mouse, r a b b i t , beaver,  domestic oat, polecat, palm c i v e t , dog, wolf, coyote, fox, marten, f e r r e t , European and American badger, racoon, polar bear, common bear, and mongoose, while the following have been experimentally infected:  Guinea pig, monkey,  sheep, c a t t l e , horse, young chickens, pigeon, magpie, rook, hamster, gopher, ape, crow, Jackdaw and hawk. Gould expresses skepticism about the published claims of certain writers concerning the finding of n a t u r a l l y infeoted frog, e e l , sparrow and pike.  The p o s s i b i l i t y  of oold-blooded v e r t i b r a t e s aoting as hosts to T r l o h i n e l l a s p i r a l i s p a r t i c u l a r y intrigues the author.  In t h i s con-  nection an experiment was performed i n whioh an attempt was made to Infect g o l d f i s h .  The d e t a i l s of t h i s experi-  ment appear i n a l a t e r section.  Natural History The l i f e cycle of T r l o h i n e l l a s p i r a l i s takes place within the body of a single host, and i s the same regardless of the host speoies.  Infection takes place when meat  containing l i v e larvae i s eaten.  Exoystment of the larvae  occurs i n the stomach within a few hours, a f t e r whioh they migrate to the small i n t e s t i n e where sexual maturation takes place.  This prooess i s completed i n forty-eight  hours, and the females, which are more numerous than the males, have been f e r t i l i z e d at the end of that time.  The  males, having completed t h e i r function, pass out with the faeces, while the females burrow into the i n t e s t i n a l mucosa, where they begin to give b i r t h to l i v i n g young on the sixth day*  It has been estimated that a female Trichina  produces some f i f t e e n thousand larvae.  The vast majority  of these are born p r i o r to the twentieth day.  A diminish-  ing number, however, continue to be produced u n t i l the death of the parent somewhere between the seventh and twelfth week.  The newborn larvae penetrate the i n t e s t i n a l  lymphatic vessels from which they pass to the thoracic duct, the right side of the heart, and f i n a l l y are d i s t r i buted to the s k e l e t a l muscles by the peripheral c i r c u l a t i o n . The invading larvae are apparently guided by a histotropism f o r although some apparently penetrate "foreign" v i s o e r a l and nervous tissues, the vast preponderance enter the endomysia of s k e l e t a l muscle f i b r e s .  In t h i s s i t e the  larvae develop i n s i z e , apparently feeding on necrotic muscle o e l l s , u n t i l the t h i r d week a f t e r invasion when they assume t h e i r c h a r a c t e r i s t i c s p i r a l form, and a closed capsule begins to develop around each.  This capsule be-  comes progressively thicker, and i s two walled.  In a l l  p r o b a b i l i t y i t s outer wall i s derived from host tissues and the inner one from the nematod.e products. as the s i x t h month a c a l c i f i c a t i o n process may  As early begin.  This u s u a l l y starts at the poles of the capsule, and  - 9 -  spreads from there to the l a t e r a l capsule walls and to the worm i t s e l f *  During the processes of encapsulation and  c a l c i f i c a t i o n the worm undergoes no further development. The period of l i f e has been found to vary considerably since some die p r i o r to c a l c i f i c a t i o n , while Langerhans (in Gould) i n 1892 recorded l i v e larvae being present i n c a l c i f i e d cysts 31 years a f t e r i n f e o t i o n .  A second host  may be infected by eating such meat as described above at any time during the l i f e of the larvae a f t e r they become i n f e c t i o u s , which condition i s reached at the time encapsul a t i o n begins. It i s i n t e r e s t i n g to note that the invading larvae demonstrate a preference f o r active muscles with a good blood supply, and that development cannot take place i n any other s i t e .than s k e l e t a l musolature.  The crura and  costal regions of the diaphragm are favorite s i t e s , while i n t e r c o s t a l , laryngeal, tongue, and eye muscles are also heavily infected.  Methods Baslo  Techniques A knowledge of the l i f e h i s t o r y of T r l o h i n e l l a  s p i r a l i s leads to the r e a l i z a t i o n that cases of t r i c h i n o s i s may  be most successfully discovered by an examination of  diaphragm t i s s u e .  Accordingly, f o r the purpose of t h i s  - 10  Survey, diaphragms were collected twice weekly from the Vancouver General Hospital, St. P a u l s Hospital, and ,  Shaughnessy Hospital, i n Vanoouver.  The sampling was un-  biased as f a r as the author was concerned, inasmuch as i t represented a consecutive series of autopsies, broken, only by forgetfulness on the part of the morgue attendants. Hospitals were asked to save as much of the diaphragm as possible. Each sample was examined as soon as possible by two methods.  F i r s t of a l l a one gram portion was  squeezed  between glass plates and examined d i r e c t l y under the microscope x45.  Secondly a ten gram sample was removed,  ground up, and digested i n an a r t i f i c i a l g a s t r i c juice f o r t h i r t y - s i x to forty-eight hours, a f t e r which i t was put into a modified Baermann apparatus.  This was allowed  to stand f o r two hours to allow any larvae to concentrate i n the stem of the funnel.  Subsequently 30 c.c. were drawn  o f f and inspected microscopically. The modifications of these basic techniques used i n the early part of the survey were those employed by T. W. Cameron (1938) Montreal.  These were adopted i n an  e f f o r t to maintain uniformity i n Canadian surveys.  Un-  fortunately certain aspects of the o r i g i n a l technique proved unsatisfactory.  To remedy t h i s s i t u a t i o n tests  and experiments were performed and more suitable methods  M.  11  adopted.  The d e t a i l s of these procedures are discussed  below. Compression Method During the early part of the survey, examination was carried out i n the type of compressor shown i n F i g . I . This consisted of two plates of quarter inch glass, two inches by eight inches, damped together by one-quarter inch bolts running through counter-sunk holes of an inch from the ends of the p l a t e s . proved t h i s devioe to be unsuitable.  three-quarters  Breakages soon  An attempt at  strengthening was made by using a one-half inch plate on the bottom, but the top plate s t i l l broke, and the cost and time l o s t demanded a more p r a c t i c a l arrangement.  To meet  t h i s end the author and h i s father fashioned the compressor pictured i n F i g . I I .  These plates were of one-quarter  inch plate glass four inches by f i v e and a h a l f inches. The frame was made of one-sixteenth  inch brass and leaves  a window two and a h a l f inches by four inches.  This l a t t e r  compressor proved to be e n t i r e l y s a t i s f a c t o r y . When the a l l glass compressor was used, a one gram sample of tissue was weighed out and examined. However a f t e r the brass-framed device had been i n operation f o r a short time the weighing procedure was eliminated.  I t was  discovered that the window held approximately one and a quarter grams of t i s s u e , and subsequently a s p a c i a l  12  c r i t e r i o n was used. Beginning with the two hundred and f i f t i e t h sample Cameron s iodine s t a i n technique was adopted. ,  This pro-  cedure involved staining the muscle i n a 0.5$ solution of iodine f o r ten minutes, and then destaining i n 2^ photographic hypo u n t i l the iodine colour was removed from muscle t i s s u e .  This method was tested on experimentally  infeoted rat tissue and was found to s t a i n l i v e larvae a deep reddish brown. Digestion Method The digestion technique used i n the early examinations was s i m i l a r to that used by Cameron and consisted of digesting ten grams of chopped muscle i n a g a s t r i c juice oomposed of 0.01 grams of Papain i n 30 c o . of normal (8$) saline f o r t h i r t y - s i x hours at 37° C.  A f t e r t h i s time'  the residue was allowed to stand f o r two hours i n a Baermann apparatus.  This apparatus consisted of an eight  inch 60° funnel stopped at the bottom, with a kitohen seive l y i n g i n the funnel, and three layers of cheesecloth l i n i n g the seive.  Unfortunately  no heat controlled room  was a v a i l a b l e , so the procedure used was as follows.  The  funnel was warmed up and f i l l e d with warm (37°) water u n t i l the l e v e l was Just above the inside of the seive.  Then  the digest residue was poured into the seive and the whole was allowed to stand f o r two hours to allow any l i v e larvae to wriggle through the f i l t e r i n g apparatus and down into  -  the stem of the funnel*  13  -  At the end of that time the lower  30 o.c. were drawn o f f and examined x45 under the miorosoope. F i r s t l y papain proved to be conducive to putrefaction and the resultant odour was most offensive to say the least.  A surface f i l m of chloroform  or t o l u o l  diminished  t h i s effeot s l i g h t l y , but neither was completely successful, and the use of either increased the cost of materials. Secondly, the digestions d i d not run to s a t i s f a c t o r y completion.  Probably any l i v e larvae would have been freed,  but a r e s i d u a l muous-like slime resulted i n every case which increased the time required f o r oleanlng up the Baermann apparatus. In order to discover a s a t i s f a c t o r y digestive juice an experiment was set up wherein various concentrations of pepsin and papain were used to digest ground lean beef. The conditions and r e s u l t s of t h i s experiment are found i n Table I . Experiment designed to discover more s a t i s f a c t o r y Digestion Technique In a l l oases the sample was f i v e grams i n weight, and of uniformly lean ground beef.  The g a s t r i c juices were  50 c o . i n volume, the pepsin being dissolved i n 0.01 M . HG1 and the papain i n 0.8$ NaCl.  The solutions were un-  corrected f o r pH and were not s t i r r e d , but were l e f t to incubate i n the 37° oven.  A f t e r twelve, sixteen, twenty,  - 14 -  twenty-four, and t h i r t y - s i x hours of inoubation, the solutions were examined f o r condition, and an estimate was made as to whether or not digestion had proceeded to a degree s u f f i c i e n t to free any larvae that might be present. The r e s u l t s of these tests showed pepsin to be f a r superior to papain.  As opposed to the slimy v i l e - s m e l l i n g  papain residue, pepsin was found to be p r a c t i c a l l y odourl e s s and to leave a clear amber solution with a f i n e precipitate.  A solution of 0 . 2 $ pepsin was found to digest  the beef to completion i n sixteen hours, and was chosen to be used throughout the remainder of the survey, beginning at the one hundred f i f t e e n t h diaphragm sample. The only other departure from the o r i g i n a l technique was the elimination o f the use of a meat grinder.  The  author found that he could save considerable time by cutting the tissue into f i n e pieces with scissors i n preference to using a meat grinder which required cleaning and b o i l i n g . The e f f i c i e n c y of the digestion was not impaired by the modification. In the eases where Trichinae were found to be present, the i n t e n s i t y of i n f e c t i o n was determined on the basis of a compression examination of f i v e grams o f t i s s u e .  The  i n t e n s i t y was expressed i n terms of the number of larvae per gram of diaphragm.  Table I The Bffiolenoy of Various Concentrations of Pepsin and Papain i n Digesting Ground Beef  $ Pepsin  12 hrs.  16 hrs.  20 h r s .  24 hrs.  36 h r s .  0.5  C  C  C  C  C  0.4  N  c  C  C  C  0.3  N  c  C  C  C  0.£  N  c  C  C  C  0.1  I  N  C  C  C  0.08  I  N  C  C  C  0.05  I  I  N  N  c  # Papain  12 hrs.  16 hrs.  20 hrs.  24 hrs.  36 hrs.  1.0  I  N  C  C  C  0.75  I  N  N  N  C  0.5  I  I  N  N  C  0.25  I  I  I  N  N  0.1  I  I  I  I  N  0.075  I  I  I  I  I  0.05  I  I  I  I  I  0.03  I  I  I  I  I  0.01  I  I  I  I  I  Index to Symbols used i n Table I I -  Incomplete  N - Nearly Complete  C - Complete  Results Results of the Present Survey A t o t a l of four hundred diaphragm samples were examined, and of these sixteen, or four per cent were found to oontain Trichina oysts.  This figure remained consistent throughout  the survey inasmuch as the successive groups of one hundred contained four, three, four, and f i v e p o s i t i v e tissues respectively. Theoretical range of the inoidenoe was calculated using the method suggested by Simpson and Roe, who staifce that such a range w i l l be represented by M i 3 a , where M i s the mean and a  i s the standard deviation.  The l a t t e r figure  i s arrived at by using the formula a =<Y npq  (Rieker 1937)  where n i s the t o t a l number of oases examined, p i s the percentage of positive cases, and q i s the percentage of negative oases*  The value o f <x i s found to be 0.99.  When  t h i s figure i s applied to the f i r s t formula i t i s seen that the t h e o r e t i c a l range of incidence i s from 1.03$ to 6.98$. This means that there i s a 95$ p r o b a b i l i t y that the incidence of t r i c h i n o s i s i n the whole population sampled l i e s within t h i s range. Table II shows the d i s t r i b u t i o n of positive oases with respect to the hospitals from whioh they came*  Ten p o s i t i v e  tissues came from the Vancouver General Hospital, two from St. Paul's Hospital, and four from Shaughnessy H o s p i t a l .  - 16 Table I I The D i s t r i b u t i o n of Positive Tissues with Respeot to Source  Source  Number Examined  Number Positive  # Positive  213  10  4.7  87  Z  2.3  Shaughnessy  100  4  4.0  TOTAL  400  16  4.0  Vancouver General St. Paul's  In nine of the p o s i t i v e oases. the cysts were d i s covered by compression method only, while, the remaining seven were detected by both compression and digestive techniques. In only three instances were l i v e larvae found.  The remain-  ing thirteen p o s i t i v e tissues contained cysts i n which the worms were dead or absent, and the worms, cysts, or both were c a l o i f i e d (Figures 5 - 13). Results of an Attempt to Infeot Goldfish with T r l e h l n e l l a spiralis The fact that Trichinae tolerate a wide range of host species tempted the author to consider the p o s s i b i l i t y of i n f e c t i n g a cold-blooded vertebrate.  In an attempt to test  t h i s p o s s i b i l i t y two experiments were set up using Goldfish Carassius auratus L. as subjects. In the f i r s t experiment, approximately two thousand Trichina larvae were forced by means of an eyedroppea? down  17  the throat of each of four f i s h .  A f t e r four days the f i s h  were k i l l e d , the i n t e s t i n e removed, s l i t open, and soraped, and the sorapings examined f o r adult Trichinae.  No worms  were found i n any of the four f i s h . Subsequently approximately three thousand larvae were foroed down the throat of each of s i x f i s h .  To further  expose them to i n f e o t i o n , they were fed f o r four  suocessive  days on ground rat meat whieh was heavily triehinosed. The f i s h were kept i n an aquarium f o r nine weeks a f t e r which they were k i l l e d , skinned, decapitated, and eviscerated.  One gram of musole was  examined by direct compression,  and the remainder of the carcass was  chopped up  and  examined by the routine digestion method. Not a single l a r v a was  discovered by these examinations.  F a i l u r e to infect Carassius auratus with T r i e h i n e l l a s p i r a l i s i s probably a t t r i b u t a b l e to the low body temperature of the f i s h . f i s h may  The r e l a t i v e l y short i n t e s t i n e of Gold-  also play a r o l e however, and the author f e e l s  that s i m i l a r experiments performed on f i s h with a more complex digestive tract might show p o s i t i v e r e s u l t s .  Discussion Possible Sources of Error i n Results On considering h i s methods the writer i s forced to conclude that a c e r t a i n number of p o s i t i v e tissues have been missed, and that the incidence of t r i c h i n o s i s i n t h i s  - 18  area i s a c t u a l l y higher than the four percent indicated. Four possible sources of error exist whioh might have allowed such tissues to escape unnoticed. The f i r s t such p o s s i b i l i t y concerns the fact that the present survey was oonfined to an examination of the diaphragm t i s s u e s .  Evans 1938  ( i n Kerr et al) found an  incidence of 36$ i n one hundred consecutive autopsies at Cleveland, Ohio, but would have reported a 25$ incidence had he oonfined h i s examinations to the diaphragms*  The  remaining 10$ were found i n the sternomastoid and i n t e r costal muscles.  S i m i l a r l y Walker and Breckenridge 1938,  (in Kerr et al) reported an incidence of 33$ i n one hundred autopsies i n Alabama, nine of which were discovered i n intercostal  t  rectus abdominis, and pectoral muscles and not  i n the diaphragm.  Thus the writer concludes that while  the diaphragm i s the most l i k e l y seat of a t r i c h i n a infeotion, any survey which bases i t s results on diaphragm examination alone i s l i k e l y to miss up to 10$ of the p o s i t i v e oases* Seoondly t h i s survey has been c a r r i e d out u t i l i z i n g only one gram of tissue f o r compression examination and ten grams f o r the digestion*  I t seems very possible that one  or more very l i g h t infections may have escaped notice. This contention i s born out by the work of Jacobs 1938 ( i n Kerr et al) who, using direct compression only, examined ten gram samples of one hundred tissues recorded as negative  - 19 -  toy routine one gram sample methods.  In t h i s one hundred  supposedly negative tissues Jacobs found six to be p o s i t i v e . The t h i r d possible means by which certain positive tissues may not have been recognized as such, involves the length of time the digestive mixture was allowed to incubate.  Oursoh (1948) has shown that the v i a b i l i t y of  T r i e h i n e l l a larvae i s d e f i n i t e l y reduced by peptic digestion of longer than twelve hours duration. Thus i t seems not unreasonable to suppose that l i v e larvae may have existed i n some of the tissues and may not have survived the t h i r t y six to forty-eight hours digestion used i n the present survey. F i n a l l y the author acknowledges the fact that h i s Baermann apparatus was not i d e a l .  No incubation room was  at h i s disposal, so, as mentioned previously,  compensation  was attempted by warming the apparatus and f i l l i n g the funnels with warm (37°) water. room temperature  This quickly cooled down to  (^20°) however, and would have decreased  the a c t i v i t y of any larvae present to a very great degree. Thus the author regards h i s apparatus as having been a c t u a l l y a simple sedimentation system rather than a true Baermann apparatus.  (The e f f i c i e n c y of the l a t t e r being  due to the a c t i v i t y of larvae being concentrated.) A further observation was made while larvae from experimentally infected animals were being concentrated. author noted that the angle of the funnels was great  The  £0  enough (60°) to allow many larvae to s e t t l e on the runnel wall.  This s i t u a t i o n probably would not have been so pro-  nounced had the larvae been kept active i n a 37° oonstant temperature room.  Nevertheless, l i g h t i n f e c t i o n s might  have been missed on t h i s acoount, and c e r t a i n l y a more acute angled funnel would have been desirable. Taking into consideration a l l the aforementioned channels by whioh some positive cases may have escaped notioe, the author f e e l s j u s t i f i e d i n presuming the incidence of human t r i c h i n o s i s to be somewhat above four percent. Analysis of data and case h i s t o r i e s An attempt to correlate the incidence of t r i c h i n o s i s with such factors as r a c i a l o r i g i n , m i l i t a r y service, residence, and economic status, had o r i g i n a l l y been planned to be included i n t h i s study.  However the size of the  sample examined and the incompleteness of many of the h o s p i t a l reoords proved t h i s aim to be i n f e a s i b l e .  Further  modification was required by the fact that on six separate occasions, two diaphragms arrived from the Vancouver General Hospital bearing the same autopsy number.  These were  designated i n the laboratory as tissue A and tissue B. Examination revealed that i n four of these pairs, both diaphragms were negative, but i n the case of autopsy number 84£, A was negative and B was p o s i t i v e , while i n autopsy number 848, A was p o s i t i v e and B was negative.  This means  that there i s only a f i f t y percent chance that the h o s p i t a l  - 21 -  records of the numbers 842 and 848 a c t u a l l y r e f e r to the positive tissues. The l i m i t e d data a v a i l a b l e , as seen i n Table III suggests a possible c o r r e l a t i o n between the incidence of t r i c h i n o s i s and c e r t a i n national food customs, such as the eating of raw sausage.  It i s noted that four of the p o s i t i v e  cases i n whioh the n a t i o n a l i t y i s known are from Central or Western European countries.  H a l l (1938) noted a higher  incidence i n Americans of European o r i g i n than i n those of English, Scottish, or I r i s h o r i g i n . The writer, i n h i s study of case h i s t o r i e s , attempted to pick out from the recorded  has  symptoms any con-  d i t i o n s suggestive of the actual i n f e o t i o n .  Here again,  however, he found that f o r the most part h i s e f f o r t s were frustrated by a lack of h i s t o r y i n the h o s p i t a l records. In the documents of the seventy-four  year old I t a l i a n  male the following statement occurs "Brain weighs  1380  grams and on sectioning presents i n the l e f t i n t e r n a l capsule rather scattered areas of softening and one or small c y s t i c spaces suggestive  of old i n f a r c t i o n .  n  two  An  attempt was made to locate the mentioned s l i d e , but i t was found that only lung and kidney s l i d e s of t h i s oase had been saved.  It i s u n l i k e l y that the c y s t i c spaces mentioned  had any connection with the t r i c h i n o s i s condition. larvae have been known to penetrate the spaces may  However  "foreign" tissues, and  Just possibly have been lesions l e f t by such  Table 111  Date Concerning the Positive Cases  Age  Sex  Racial Origin  V.G.H.  74  M  Italian  20  V.G.H.  71  M  Hungarian  6  V.G.H.  50  F  English  ?  No  V.G.H. • V.G.H.  67  M  German  ?  No  6 weeks  M  ?>  V.G.H. *?  69  M  Spanish  Residence i n Vancouver  Military Service  Hospital  Yr.  Yr.  Condition of Larvae Dead. Cysts C a l c i f i e d .  No  Dead and fragmented Cysts & 70 worms c a l c i f i e d * Mixed dead and a l i v e not 1 calcified* Dead. Worms c a l c i f i e d cysts not. 1 Dead or absent. Cysts c a l c i f i e d or n o t . 22 Mixed dead and a l i v e worms & cysts o a l c i f i e d or not. 9 Dead or absent worms & cysts o a l c i f i e d or n o t . 0.2 Dead. Worms & cysts c a l c i f i e d or not. 52.8 Dead & fragmented worms & cysts c a l c i f i e d or n o t . 4.2 Dead. Worms & cysts c a l . or not 2.4 Dead. Cysts c a l . larvae not. 0 . 8  6 weeks  N/A  Pt. A l b e r n i  ?  24  Yr.  No  21  Yr.  ?  Irish  ?  ?  ?  ?  ?  ?  88  M  ?  Yr.  Yes  Shaughnessy  67  M  ?  ?  Yes  Shaughnessy  77'  M  ?  ?  Yes  Dead or absent Larvae c a l . cysts not. Dead. Cysts heavily c a l .  Shaughnessy  79  M  ?  ?  Yes  A l i v e . Cyst c a l .  St. Pauls  72  M  ?  ?  ?  St. Pauls  67  F  ?  Yr.  ?  English  61  F  V.G.H*  35  F  ?  V.G.H.  83  M  V.G.Ho  ?  Shaughnessy  11  10  4  No  37Yro  V.G.H. *?  Number per Gram  0.4 6 .  2  Dead. Cysts & larvae c a l or not Dead. Larvae c a l . , cysts not •  8  * Question marks i n column X indicate' Tissues which may have wrong number. Question marks i n other colums indicate information unavailable i n Hospital records.  0.2  -  invading larvae.  2  2  Brain lesions caused by Trichinae are  discussed by Gould. P a r t i c u l a r note was taken of the number of p o s i t i v e oases having h i s t o r i e s of rheumatic conditions.  In t h i s  regard i t was found that only two of the sixteen had r e corded h i s t o r i e s of a r t h r i t i c conditions and that one of these was found at autopsy to be an o s t e o a r t h r i t i s of the spine.  The other, the sixty-seven year old German male had  a seven year o l d h i s t o r y of a r t h r i t i s i n the pectoralscapular region with pronounced atrophy of the muscles. While the l a t t e r case might possibly have some s i g n i f i c a n c e , these r e s u l t s oertainly do not o f f e r any evidence toward incriminating t r i c h i n o s i s as a f a c t o r i n rheumatic conditions. The appearance of Triohinae i n the diaphragm of a six weeks old baby presented a noteworthy case.  Early  writers believed that intrauterine i n f e c t i o n d i d not take place.  However Roth 1935, 1936,  (In Gould) demonstrated  muscle triohinae i n the foetuses of guinea pigs Infected during pregnancy, and Kuitunen Ekbaum (1941) reported finding four l i v e larvae i n 6.5 grams of diaphragm tissue from a seven months o l d foetus. communication)  Dr. L. Ranta (Personal  desoribed the f i n d i n g of a heavy i n f e c t i o n  of trichinae i n a foetus at autopsy i n Toronto. In the present case, the baby was born f i v e weeks prematurely i n a f r a i l condition, .  He fed poorly and  -  23  -  coughed a l o t , and X-ray showed increasing opacity i n the right lung, autopsy was  Six weeks a f t e r b i r t h the baby died, and performed.  an  The report showed bronchitis,  p a r t i a l obstruction of the r i g h t side of the bronchial tree, and i n s i p i e n t croup. who  The w r i t e r contacted the  physicians  had attended the confinement and the c h i l d ' s i l l n e s s ,  and was  informed that the mother, a seventeen year old  Canadian woman, had suffered no i l l n e s s during pregnancy. The condition of the cysts i s worthy of note inasmuch as the larvae were dead or absent, and many of the cyst walls showed o a l o i f i c a t i o n . i n 1896,  Ehrhardt quoted by Gould, observed  death of muscle larvae as early as the t h i r d week,  and l a r v a l c a l c i f i c a t i o n between the forty-second s i x t i e t h days.  and  S i m i l a r l y Leukart (In Gould) demonstrated  c a l c i f i c a t i o n as early as the eightieth day.  Nonetheless,  the process of c a l c i f i c a t i o n does not usually commence u n t i l a f t e r the sixth month of i n f e c t i o n , and the w r i t e r f e e l s that t h i s case may  indicate a difference i n the rate of  c a l c i f i c a t i o n i n foetus over that i n the adult. Comparison of l o c a l findings with those obtained  elsewhere  The incidence of human t r i c h i n o s i s i s governed by main factors; v i z . care exercised i n r a i s i n g hogs and paring pork products,  two pre-  and food and cooking habits.  Table IV presents a picture of the incidence i n humans i n various areas,  unfortunately no mathematical corre-  l a t i o n s can be drawn because i n certain areas i . e . Germany, no recent surveys have been made, and the figures represent  - 24 -  only approximate values.  Furthermore the s i t u a t i o n i s com-  p l i c a t e d by.the fact that hog surveys which are so i n t i mately t i e d up with human surveys, are i n places lacking, and i n other plaoes not representative of the pork which appears on the public market. locally.  The l a t t e r case exists  The author's information on t h i s subject comes  from personal communications with Sr. I. W. Moynihan  who  has a t r i c h i n o s i s survey of r a t s and hogs i n the Vancouver area i n progress.  The figures to be used represent only  approximate values, since the mentioned surveys aire not complete, and furthermore do not represent a cross section of B r i t i s h Columbia's market pork, inasmuch as they r e f e r l a r g e l y to B r i t i s h Columbia raised hogs, whereas a great portion of the pork consumed i n B r i t i s h Columbia i s raised i n the p r a i r i e Provinces. Table IV Comparison of Local Findings with those Obtained Elsewhere  Size of Sample Germany  ?  U. S. A . California Oregon Washington  5213 129 33 200  Canada Montreal Toronto Vancouver  1359 539 410 . 400  $ Positive 1.5$ 16.1$ 17.8$ 12.1$ 20. $ 2.2$ 1.5$ 1.75$ 14.0$  - 25 -  On analyzing  Table IV we see that the United States of  America has the highest inoidence of any country, having 16.1$  (Wright et a l ) .  the hog incidence  As opposed to t h i s Gould states that  i n the U. S. A* i s between one and two  percent, giving a r a t i o of about one trichinosed hog to ten trichinosed humans. Germany, on the other hand, has a human incidence  of roughly 1.5$ (Gould) and a hog incidence  of 0.00044$ (Todd 1948) or a ratio, of one hog to three thousand persons infected.  From these figures we conclude  that careful hog control as practiced i n Germany, sponsors a much lower human incidence hog  than does a system of poor  control as practiced i n the U. S. A*  The above figures  also demonstrate that food habits conducive to Trichina i n f e c t i o n can so enlarge the r a t i o of hog i n f e c t i o n to human i n f e o t i o n as to s t i l l leave a noteworthy human incidence. The Canadian picture, as based on examination of two thousand, nine hundred ninety-five hogs and one thousand, f i v e hundred f i f t y - n i n e humans, shows a 0.57$ hog infeotion opposed to a 2.2$ human Infeotion, or a ratio: of one trichinosed hog to 3.8 infected humans.  This low r a t i o  would indioate that human t r i c h i n o s i s i n Canada i s l a r g e l y a result of poor hog control, and not of cooking habits. From Table IV i t i s seen that while the 4$ human i n Vancouver i s considerably  incidence  below that of the United  It i s w e l l above that found i n Eastern Canada.  States,  The present  * 26 -  survey i s not s t r i o t l y comparable to that made by Cameron in Montreal, since children under the age of one year were excluded from the l a t t e r . However even i f compensation i s made for t h i s discrepancy, the l o c a l incidence remains over double that found i n Montreal. Possible explanation of l o c a l incidence She fact that human incidenoe i n the Vancouver area i s over twice that found i n Eastern Canada i s explained on two major bases. P i r s t of a l l , present data indicates that the incidence i n hogs i n t h i s area i s considerably higher than that present i n Eastern hogs. Cameron of  0.57$  (1940)  has shown an incidenoe  i n two thousand, nine hundred ninety-five hogs  (mixed grain and garbage feeders) of which nine hundred ninety-five known garbage feeders showed an incidence of 0 . 2 $ . Moynihan, on the other hand, demonstrated that 2 . 2 $ of six hundred ninety-one mixed B r i t i s h Columbia hogs were infected, while  4»0$  of the three hundred seventy-four  garbage feeders included i n the above six hundred ninetyone , were p o s i t i v e .  The trichinosed hogs found by Moynihan  were a l l raised at piggeries managed by Orientals on Lulu Island, near Vancouver. The reader i s asked to bear i n mind the faot that Moynihan's results do not represent an incidence in hogs marketed i n B r i t i s h Columbia, but only those raised and marketed here.  The author has been unable  to obtain a statement of the ratio of P r a i r i e raised hogs  - 27 to B r i t i s h Colombia r a i s e d hogs which are marketed i n B r i t i s h Columbia.  However Moynihan's survey does lend  i t s e l f to the supposition that the incidence i n t o t a l marketed pork i n B r i t i s h Columbia i s somewhat higher than that i n Eastern Canada* The r e l a t i v e l y high incidence i n l o c a l l y r a i s e d hogs i s l i k e l y due to c a r e l e s s feeding by the farmers, but a l s o be a t t r i b u t a b l e to hog i n f e o t i o n from r a t s .  may  Thus f a r  Moynihan has examined the e n t i r e diaphragms of f i v e hundred n i n e t y r a t s by compression only. f i v e or  l6.1$  were t r i c h i n o s e d ,  Of t h i s number n i n e t y 87.3$  of the p o s i t i v e s being  r a t s from around p i g g e r i e s , and the remaining around garbage dumps.  12*7$  from  This incidence i n r a t s i s not unusual  and it... i s d o u b t f u l i f such a c o n d i t i o n i s responsible f o r the high incidence i n hogs, although some hogs probably do become i n f e c t e d by eating these  rats.  The second b a s i s of explanation f o r the high human inoidence i n t h i s area i s found i n considering the high t r a f f i c to the United States*  People i n Eastern Canadian  c i t i e s are l i k e l y to make t h e i r week-end excursions i n an Eastern-Western plane, c o n f i n i n g themselves to other Canadian c i t i e s .  L o c a l l y however, Vancouver i s the sole  large Canadian c i t y on the mainland, and week-end t r a v e l by motor car i s l a r g e l y d i r e c t e d South across the I n t e r n a t i o n a l border.  The 20$ human incidence i n the State of Washington  (Ref. Table IV) would i n d i c a t e a f a i r l y high hog incidence. Then considering that i t takes only one meal of t r i c h i n o u s  - 28 -  pork to cause i n f e c t i o n , i t seems very l i k e l y that considerable of the l o c a l incidence may  be attributed to i n -  feotion contracted across the border. At the time when t h i s survey was being organized, author noted that nearly every medical man  contacted  the ex-  pressed the opinion that no t r i c h i n o s i s problem existed i n the Vancouver area.  This survey however shows not  only  that l o c a l human beings are trichinosed, but also that the incidence and i n some oases the i n t e n s i t y i s s u f f i c i e n t l y high to be considered  a serious medical problem.  It must  be born i n mind that the mere presence of T r i c h i n o s i s represents p o t e n t i a l epidemics and possible f a t a l i n f e c t i o n s * The author believes that i f hog i n f e c t i o n s , which are apparently quite l o c a l i z e d , were adequately controlled, and i f the public was  informed of the t r i c h i n o s i s s i t u a t i o n i n  the United States and discouraged  from eating pork i n that  country, that human t r i c h i n o s i s could be v i r t u a l l y wiped out i n t h i s area.  - Xt -  References Cameron, T. W. M.  Trichinosis Can. Jour. Comp.  Cameron, T. W. M.  Investigations on T r i c h i n o s i s i n Canada. I. A Preliminary Survey of the Incidence of T r l o h i n e l l a s p i r a l i s i n Hogs i n Eastern Canada. Can. Jour. Res., D, 16: pp. 89-92, 1938  Cameron, T. W. M.  Investigations on T r i c h i n o s i s i n Canada. I I I . On the Incidenoe of Triohinosis i n Garbage-Fed Hogs. Can. Jour. Res., D, 18: pp. 83-85, 1940  Cameron, T. W. M.  Studies on Triohinosis. IV. Human Incidence i n Montreal. Can. Jour. Res. 21: pp. 413-414, 1943  Gould, S. E.  Triohinosis, pp. 1-290 Charles C. Thomas and Co., Springfield, I l l i n o i s , U.S.A., 1945  Gursch, 0. F.  E f f e c t of digestion and r e f r i g e r a t i o n on the a b i l i t y of T r l o h i n e l l a s p i r a l i s to infect rats. Jour. P a r a s i t o l . 34: pp. 394-395, 1948  Hall,  C.  Kerr, K. B.,  . 1: pp. 5^14, 1937  Studies on T r i c h i n o s i s , VI. Epidemiol o g i c a l Aspects of T r i c h i n o s i s i n the United States as Indicated by an Examination of 1,000 Diaphragms, f o r Trichinae. U.S. Pab. Health Repts. 53: pp. 10861105, 1938 Jacobs, L., and C u v i l l i e r , E. Studies on T r i c h i n o s i s . XIII. The I n c i denoe of Human I n f e c t i o n with Trichinae as Indicated by Post-Mortem Examination of 3,000 Diaphragms from Washington, D.C., and Five Eastern Seaboard C i t i e s . U.S. Pab. Health Repts. 56: pp. 836855, 1941  - 3 o -  Kuitunen-Ekbaum, E. The Inoidenoe of Trichinosis i n Humans i n Toronto. Findings i n 420 Autopsies* Can. Pub. Health Jour. 22: pp.569-576, 1941 — Rioker, W. E.  The Conoept of Confidence or F i d u c i a l Limits applied to the Poisson Frequency Distribution. Jour. Am. Stat. Assoc. 32: pp. 349 356, 1937  Simpson, G. G. and Roe, A* Quantitative Zoology. pp. 1-407, McGraw-Hill Book Co. Inc., New York and London, 1939 Stefansson, V.  Trichinosis from Polar Bear Meat. A r c t i c 1: pp. 144, 1948  S t o l l , N. R.  This Wormy World. Jour P a r a s i t o l . 33: pp. 1-18, 1947  Todd, F. A*  Triohina Inspection i n Germany. Vet. Med. 43: 1948  Wright, W. H. and Bozicevich, J . Experiments i n the cooking of Garbage f o r the Destruction of Trichinae i n Pork Scraps. ' tT.S. Publ. Health Repts. 58: pp. 396404, 1943 1  Wright, W. H., Kerr, K. B., and Jacobs, L. Studies on T r i c h i n o s i s . XV. Summary of the Findings of T r i o h i n e l l a s p i r a l i s i n a Random Sampling and other samplings of the Population of the United States. U.S.Publ. Health Repts. 58: pp. 12931313, 1943  Fig.. 1  Compressor used during the early part of the survey with broken top plate.  Fig. 2  Compressor used during the l a t t e r part of the survey disassembled,  '  figure  figure  1.  2  fig.  2  Fig. 4  Brass  framed compressor assembled.  partially-  B r a s s framed compressor  assembled.  Fig.  5  T r i c h i n a l a r v a e i n r a t muscle to encapsulation X 8 0  prior  Fig.  6  T r i c h i n a o y s t i n human m u s c l e p o l a r f a t b o d i e s X80  showing  F i g u r e 5.  F i g u r e 6.  C a l c i f i e d c y s t i n human m u s c l e c r a c k e d by c o m p r e s s i o n 180  Human m u s c l e c o n t a i n i n g s p i n d l e s h a p e d cyst. Both l a r v a and c y s t c a l c i f i e d X80  Pig.  6  Heavily calcified larva in uncalcified cyst. Note d o u b l e w a l l o f the c y s t 188  Pig.  10  Human m u s c l e c o n t a i n i n g h e a v i l y c a l c i f i e d c y s t w i t h l a r v a f r a g m e n t e d and calcified X80  Fig.  11  U n c a l o i f i e d cyst containing and c a l o i f i e d l a r v a X80  broken  Fig.  12  Dead and c a l c i f i e d l a r v a w i t h c y s t w a l l p r a c t i c a l l y a b s e n t , i n human m u a o l e X80  Figure  11.  Figure  12,  F i g . 12  Calcified  cyst with  larva  absent  X80  F i g u r e 13.  

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