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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 Partial Fulfilment of the Requirements for the Degree of MASTER OF ARTS in the Department of ZOOLOGY" THE UNIVERSITY OF BRITISH COLUMBIA April 1949 ABSTRACT Four hundred diaphragms from Vancouver hospitals were examined for Trichinae "by direct compression and by digestion technique. Sixteen, or four percent were found to contain trichina cysts. By applying the formula M i 3d , where M i s the mean, and a i s the standard devia-tion, theoretical range of the inoidenoe of trichinosis in the entire population was determined to be between 1.03$ and 6.98$. Limitation of the amount of tissue examined, viz . one gram by compression and ten grams by digestion, and limitations imposed by the fact that only diaphragm muscle was examined, allowed an opportunity for certain light infections to pass unnoticed. It i s therefore con-cluded that the incidence of human trichinosis.in the Vancouver area i s probably somewhat above four percent. While no reeent infections were discovered, this incidence, which i s over double that found in Eastern Canada, and in some oases the intensity of infection, the highest being seventy cysts per gram of muscle, conclusively show that trichinosis constitutes a serious public health problem in 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 Basic Techniques 9 Compression Method. 11 Digestion Method 12 Experiment designed to disoover more sat i s f a c t o r y digestion technique 13 Results 15 Results of the present survey 15 Results of an attempt to infe c t .Goldfish with T r l c h l n e l l a s p i r a l i s ; 16 Discussion 17 Possible sources of error i n res u l t s 17 Analysis of data and case h i s t o r i e s 20 J Comparison of l o c a l findings with those obtained elsewhere 23 Possible explanations f o r l o o a l incidence 26 References Plates Acknowle dgement a The author wishes to express his sincere gratitude to the many friends and advisors who have contributed so materially to the survey. In particular he wishes to thank his Mother and Father for their counsel, encourage-ment, and financial aid; Sr. J. R. Adams, his director, who has offered help and constructive criticism throughout the course of the work; Dr. W. A. Clemens whose door i s never olosed to those seeking advice; Dr. I. 18 • Moynihan who has frequently offered material, friendly encouragement, and sound advice; Dr. H. E. Taylor, Dr. H. H. Pitts, Dr. H. K. Pidler, and Mr. P. J. Pish, for their co-operation and interest in making available the necessary tissues and hospital records; Mr. J. W. MacKay, an excellent friend, who has oheerfully given of his time and knowledge in pre-paring the photographs; and Miss Patricia Bourns who has worked long hours typing in order that this paper might be finished on time. - 1 -The Inoidenoe of Human Trichi n o s i s i n the Vancouver Area as Determined by the Examination of Four Hundred Diaphragms Introduction Triohinosis, 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 of i t s host. While the vast majority of human infections remain sub-c 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 indicate that the very existance of t r i c h i n o s i s i n a community constitutes a serious medical problem. Human infe o t i o n i s generally a t t r i b u t e d to eating raw or improperly cooked pork. S i m i l a r l y , hogs are known to become triohinosed by eating garbage containing infected pork scraps. Control i s directed f i r s t of a l l toward prevention of hog infeotio n , and secondly to treatment of pork destined for raw consumption. Since heat i s known to k i l l t r i c h i n a larvae, (Wright and Bozicdvieh 1943), the former type of control i s exeroised i n Canada by a Federal law which decrees that a l l garbage used as hog s w i l l must - 2 -be cooked. It i s noteworthy that no such law exists in the U. S. A. The second type of control, v i z . treatment of pork to be used in 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 for raw consumption must be treated by heat, cold, or salt, in suoh a way as to k i l l any larvae present.. Final 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 feels well protected by Government meat inspection. What he does not realize i s that in neither country i s there any inspection made for 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 trichinosis in Western Canada. Theoretically we are adequately protected in our own country. However we have had no proof that our control measures have been conscientiously praotised. Furthermore, the city of Vancouver i s located a bare thirty 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 six, or a total of twenty-one million Americans harbour Trichinae within their bodies. S t o l l (1947) estimates that there i s three - 2 -times as much trichinosis in 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 half per cent of the positive eases found, showed oaunts of f i f t y or more larvae per gram of muscle, infection of an order " capable of causing pro-nounced 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 million larvae* Besides describing a serious medical problem in the United States, the foregoing figures represent a tremendous number of meals of pork containing infective trichinae* It seems only reasonable then, to assume that a great number of infectious meals await the Canadian vi s i t o r * The present survey has been carried out to determine the adequacy of our controls, and whether or not t r i c h i -nosis constitutes a medical problem in the Vancouver area. Historical The history of our knowledge of trichinosis follows a pattern whioh i s oommon in biological progress. It began with unoritioal awareness, was catapulted into sc i e n t i f i c focus at a single stroke, was developed along descriptive lines, and at present i s being further de-veloped physiologically. The fact that trichinosis i s s t i l l very much i n sci 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 in the last three years, and that at least one major work has been published on the subject in that time. The major work referred to i s a very complete book, trichinosis, by S. E. Gould, from which the following histori c a l summary has been oompiled. The earliest existence of Triohinella s p i r a l i s i s obscure. Many writers however, feel 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 evolu-tionary history. Indeed i t has been suggested that the ancient Mosaio law which forbids the eating of swine flesh was Instigated following a number of trichinosis epidemios. Probably the f i r s t recorded observation of Trichina cysts i s that made by Tledman in 1821, in 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 in i t s cyst i s given to Sir •Tames Paget, who i n 1834 observed " l i t t l e specks in the muscles" of a cadaver, and who was curious enough to examine them microscopically. Sir Richard Owen, who was familiar with Paget's work, read a paper before the Zoological Society of London in 1835 i n whioh he named the worm Trichina s p i r a l i s . The next important advanoe was made by Joseph Leidy in 1846, who discovered Trichinae in the muscles of a hog. This was followed by the works of Herbst (1851), - 6 -Virohow (1859) and Leuckart (1860) in which the anatomy of the worm was more completely described, and the fact was established that an animal eating triehinous meat may develope trichinosis. In 1860 Zenker discovered a heavy infection of l i v e Trichinae in the muscles of the body of a Dresden servant g i r l . Subsequently he found adult Trichinae in the intestine, and infeotion i n the ham and sausage at the house where the g i r l had worked. Thus he postulated, trichinosis can be fatal to human beings, the complete l i f e history of the worm takes place in one host, and human infection can result from eating infected pork. Furthermore he correctly surmised that the larvae were disseminated by way of the chyle ducts and the blood stream. Examination of pork for Trichinae was begun in Germany as early as 1863, and largely at the insistence of Virohow was made oompulsary by Prussian law in 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 in effect. 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 pro-posed by Owen, v i z . Trichina s p i r a l i s to Trlohlnella s p i r a l i s * Around the same time considerable work was being done on the l i f e history and diagnosis of the para-sit e . In 1897 Brown noticed a high eosinophilia associated with the disease, and suggested i t s diagnostic value. Many workers, notably Fielder (1864), Cerfontaine (1895) and Askowazy (1895) made l i f e history studies and con-clusively 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, Mil l e r and Friedlander (1933) and f i n a l l y by Bozioivich (1939). In each case the antigen has consisted of an extraction of dried larvae, the constant improvement being directed to specificity of the antigen. As was previously stated, the common source of human infection i s trichinosed pork. The only other possible source i s the meat of an edible game animal. Sach a possibility has been mentioned by Steffanson (1948) who reported that Trichinae are prevalent in polar bears in 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 naturally Infeoted; mouse, rabbit, beaver, domestic oat, polecat, palm civet, dog, wolf, coyote, fox, marten, ferret, European and American badger, racoon, polar bear, common bear, and mongoose, while the following have been experimentally infected: Guinea pig, monkey, sheep, cattle, 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 naturally infeoted frog, eel, sparrow and pike. The possibility of oold-blooded vertibrates aoting as hosts to Trlohinella  spi r a l i s particulary intrigues the author. In this con-nection an experiment was performed i n whioh an attempt was made to Infect goldfish. The details of this experi-ment appear in a later section. Natural History The l i f e cycle of Trlohinella s p i r a l i s takes place within the body of a single host, and i s the same regard-less 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, after whioh they migrate to the small intestine where sexual maturation takes place. This prooess i s completed in 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 their function, pass out with the faeces, while the females burrow into the intestinal mucosa, where they begin to give birth to l i v i n g young on the sixth day* It has been estimated that a female Trichina produces some fifteen thousand larvae. The vast majority of these are born prior 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 intestinal 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 skeletal muscles by the peripheral circulation. The invading larvae are apparently guided by a histotropism for although some apparently penetrate "foreign" visoeral and nervous tissues, the vast preponderance enter the endomysia of skeletal muscle fibres. In this site the larvae develop in size, apparently feeding on necrotic muscle oells, u n t i l the third week after invasion when they assume their characteristic spiral 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 probability i t s outer wall i s derived from host tissues and the inner one from the nematod.e products. As early as the sixth month a calcification process may begin. This usually 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 calcification the worm undergoes no further development. The period of l i f e has been found to vary considerably since some die prior to calcification, while Langerhans (in Gould) in 1892 recorded l i v e larvae being present i n calc i f i e d cysts 31 years after infeotion. 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 after they become infectious, which condition i s reached at the time encapsu-lation begins. It i s interesting to note that the invading larvae demonstrate a preference for active muscles with a good blood supply, and that development cannot take place in any other site .than skeletal musolature. The crura and costal regions of the diaphragm are favorite sites, while intercostal, laryngeal, tongue, and eye muscles are also heavily infected. Methods Baslo Techniques A knowledge of the l i f e history of Trlohinella  s p i r a l i s leads to the realization that cases of trichinosis may be most successfully discovered by an examination of diaphragm tissue. Accordingly, for the purpose of this - 10 Survey, diaphragms were collected twice weekly from the Vancouver General Hospital, St. Paul ,s Hospital, and Shaughnessy Hospital, in Vanoouver. The sampling was un-biased as far 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. First of a l l a one gram portion was squeezed between glass plates and examined directly under the microscope x45. Secondly a ten gram sample was removed, ground up, and digested in an a r t i f i c i a l gastric juice for thirty-six to forty-eight hours, after which i t was put into a modified Baermann apparatus. This was allowed to stand for two hours to allow any larvae to concentrate in the stem of the funnel. Subsequently 30 c.c. were drawn off and inspected microscopically. The modifications of these basic techniques used i n the early part of the survey were those employed by T. W. M. Cameron (1938) Montreal. These were adopted in an effort to maintain uniformity in Canadian surveys. Un-fortunately certain aspects of the original technique proved unsatisfactory. To remedy this situation tests and experiments were performed and more suitable methods 11 adopted. The details 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 in Fig. 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 three-quarters of an inch from the ends of the plates. Breakages soon proved this devioe to be unsuitable. 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 lost demanded a more practical arrangement. To meet this end the author and his father fashioned the compressor pictured in Fig. II. These plates were of one-quarter inch plate glass four inches by five and a half inches. The frame was made of one-sixteenth inch brass and leaves a window two and a half inches by four inches. This latter compressor proved to be entirely satisfactory. When the a l l glass compressor was used, a one gram sample of tissue was weighed out and examined. However after the brass-framed device had been i n operation for a short time the weighing procedure was eliminated. It was discovered that the window held approximately one and a quarter grams of tissue, and subsequently a spacial 12 criterion was used. Beginning with the two hundred and f i f t i e t h sample Cameron,s iodine stain technique was adopted. This pro-cedure involved staining the muscle in a 0.5$ solution of iodine for ten minutes, and then destaining in 2^ photo-graphic hypo u n t i l the iodine colour was removed from muscle tissue. This method was tested on experimentally infeoted rat tissue and was found to stain li v e larvae a deep reddish brown. Digestion Method The digestion technique used in the early examinations was similar to that used by Cameron and consisted of digesting ten grams of chopped muscle in a gastric juice oomposed of 0.01 grams of Papain in 30 c o . of normal (8$) saline for thirty-six hours at 37° C. After this time' the residue was allowed to stand for two hours in a Baermann apparatus. This apparatus consisted of an eight inch 60° funnel stopped at the bottom, with a kitohen seive lying in the funnel, and three layers of cheesecloth lining the seive. Unfortunately no heat controlled room was available, so the procedure used was as follows. The funnel was warmed up and f i l l e d with warm (37°) water u n t i l the level was Just above the inside of the seive. Then the digest residue was poured into the seive and the whole was allowed to stand for two hours to allow any l i v e larvae to wriggle through the f i l t e r i n g apparatus and down into - 13 -the stem of the funnel* At the end of that time the lower 30 o.c. were drawn off and examined x45 under the miorosoope. F i r s t l y papain proved to be conducive to putrefaction and the resultant odour was most offensive to say the least. A surface film of chloroform or toluol diminished this effeot slightly, but neither was completely successful, and the use of either increased the cost of materials. Secondly, the digestions did not run to satisfactory com-pletion. Probably any l i v e larvae would have been freed, but a residual muous-like slime resulted i n every case which increased the time required for oleanlng up the Baermann apparatus. In order to discover a satisfactory digestive juice an experiment was set up wherein various concentrations of pepsin and papain were used to digest ground lean beef. The conditions and results of this experiment are found i n Table I. Experiment designed to discover more satisfactory Digestion  Technique In a l l oases the sample was five grams in weight, and of uniformly lean ground beef. The gastric juices were 50 c o . in volume, the pepsin being dissolved in 0.01 M . HG1 and the papain i n 0.8$ NaCl. The solutions were un-corrected for pH and were not stirred, but were l e f t to incubate in the 37° oven. After twelve, sixteen, twenty, - 14 -twenty-four, and thirty-six hours of inoubation, the solutions were examined for condition, and an estimate was made as to whether or not digestion had proceeded to a degree sufficient to free any larvae that might be present. The results of these tests showed pepsin to be far superior to papain. As opposed to the slimy vile-smelling papain residue, pepsin was found to be practically odour-less and to leave a clear amber solution with a fine pre-cipitate. A solution of 0 . 2 $ pepsin was found to digest the beef to completion in sixteen hours, and was chosen to be used throughout the remainder of the survey, beginning at the one hundred fifteenth diaphragm sample. The only other departure from the original technique was the elimination of the use of a meat grinder. The author found that he could save considerable time by cutting the tissue into fine pieces with scissors in preference to using a meat grinder which required cleaning and boiling. The efficiency of the digestion was not impaired by the modification. In the eases where Trichinae were found to be present, the intensity of infection was determined on the basis of a compression examination of five grams of tissue. The intensity 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 in Digesting Ground Beef $ Pepsin 12 hrs. 16 hrs. 20 hrs. 24 hrs. 36 hrs. 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 in Table I I - Incomplete N - Nearly Complete C - Complete Results Results of the Present Survey A total 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 five positive 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 latter figure i s arrived at by using the formula a =<Y npq (Rieker 1937) where n i s the total number of oases examined, p i s the percentage of positive cases, and q i s the percentage of negative oases* The value of <x i s found to be 0.99. When this figure i s applied to the f i r s t formula i t i s seen that the theoretical range of incidence i s from 1.03$ to 6.98$. This means that there i s a 95$ probability that the incidence of trichinosis i n the whole population sampled l i e s within this range. Table II shows the distribution of positive oases with respect to the hospitals from whioh they came* Ten positive tissues came from the Vancouver General Hospital, two from St. Paul's Hospital, and four from Shaughnessy Hospital. - 16 -Table II The Distribution of Positive Tissues with Respeot to Source Source Number Examined Number # Positive Positive Vancouver General 213 10 4.7 St. Paul's 87 Z 2.3 Shaughnessy 100 4 4.0 TOTAL 400 16 4.0 In nine of the positive oases. the cysts were dis-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 positive tissues contained cysts in which the worms were dead or absent, and the worms, cysts, or both were caloified (Figures 5 - 13). Results of an Attempt to Infeot Goldfish with Trlehlnella  s p i r a l i s The fact that Trichinae tolerate a wide range of host species tempted the author to consider the possibility of infecting a cold-blooded vertebrate. In an attempt to test this possibility 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 1 7 the throat of each of four f i s h . After four days the fis h were k i l l e d , the intestine removed, s l i t open, and soraped, and the sorapings examined for adult Trichinae. No worms were found in any of the four f i s h . Subsequently approximately three thousand larvae were foroed down the throat of each of six f i s h . To further expose them to infeotion, they were fed for four suocessive days on ground rat meat whieh was heavily triehinosed. The f i s h were kept in an aquarium for nine weeks after which they were k i l l e d , skinned, decapitated, and eviscera-ted. 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 larva was discovered by these examinations. Failure to infect Carassius auratus with Triehinella  s p i r a l i s i s probably attributable to the low body tempera-ture of the f i s h . The relatively short intestine of Gold-fish may also play a role however, and the author feels that similar experiments performed on f i s h with a more complex digestive tract might show positive results. Discussion Possible Sources of Error i n Results On considering his methods the writer i s forced to conclude that a certain number of positive tissues have been missed, and that the incidence of trichinosis in this - 18 area i s actually 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 possibility concerns the fact that the present survey was oonfined to an examination of the diaphragm tissues. Evans 1938 (in 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 his examinations to the diaphragms* The remaining 10$ were found i n the sternomastoid and inter-costal muscles. Similarly Walker and Breckenridge 1938, (in Kerr et al) reported an incidence of 33$ in one hundred autopsies in Alabama, nine of which were discovered in intercostal t rectus abdominis, and pectoral muscles and not in the diaphragm. Thus the writer concludes that while the diaphragm i s the most l i k e l y seat of a trichina infeo-tion, 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 posi-tive oases* Seoondly this survey has been carried out u t i l i z i n g only one gram of tissue for compression examination and ten grams for the digestion* It seems very possible that one or more very light infections may have escaped notice. This contention i s born out by the work of Jacobs 1938 (in 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 this one hundred supposedly negative tissues Jacobs found six to be positive. The third 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 incu-bate. Oursoh (1948) has shown that the v i a b i l i t y of Triehinella larvae i s definitely reduced by peptic digestion of longer than twelve hours duration. Thus i t seems not unreasonable to suppose that li v e larvae may have existed in some of the tissues and may not have survived the thirty-six to forty-eight hours digestion used in the present survey. Finally the author acknowledges the fact that his Baermann apparatus was not ideal. No incubation room was at his 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. This quickly cooled down to room temperature (^20°) however, and would have decreased the activity of any larvae present to a very great degree. Thus the author regards his apparatus as having been actually a simple sedimentation system rather than a true Baermann apparatus. (The efficiency of the latter being due to the activity of larvae being concentrated.) A further observation was made while larvae from experi-mentally infected animals were being concentrated. The author noted that the angle of the funnels was great £0 enough (60°) to allow many larvae to settle on the runnel wall. This situation probably would not have been so pro-nounced had the larvae been kept active i n a 37° oonstant temperature room. Nevertheless, light infections might have been missed on this acoount, and certainly 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 feels j u s t i f i e d in presuming the incidence of human trichinosis to be somewhat above four percent. Analysis of data and case histories An attempt to correlate the incidence of trichinosis with such factors as r a c i a l origin, military service, residence, and economic status, had originally been planned to be included in this study. However the size of the sample examined and the incompleteness of many of the hospital reoords proved this aim to be infeasible. 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 in the laboratory as tissue A and tissue B. Examination revealed that in four of these pairs, both diaphragms were negative, but in the case of autopsy number 84£, A was negative and B was positive, while in autopsy number 848, A was positive and B was negative. This means that there i s only a f i f t y percent chance that the hospital - 21 -records of the numbers 842 and 848 actually refer to the positive tissues. The limited data available, as seen i n Table III suggests a possible correlation between the incidence of trichinosis and certain national food customs, such as the eating of raw sausage. It i s noted that four of the positive cases in whioh the nationality i s known are from Central or Western European countries. Hall (1938) noted a higher incidence in Americans of European origin than in those of English, Scottish, or Irish origin. The writer, in his study of case histories, has attempted to pick out from the recorded symptoms any con-ditions suggestive of the actual infeotion. Here again, however, he found that for the most part his efforts were frustrated by a lack of history in the hospital records. In the documents of the seventy-four year old Italian male the following statement occurs "Brain weighs 1380 grams and on sectioning presents in the l e f t internal capsule rather scattered areas of softening and one or two small cystic spaces suggestive of old infarction. n An attempt was made to locate the mentioned slide, but i t was found that only lung and kidney slides of this oase had been saved. It i s unlikely that the cystic spaces mentioned had any connection with the trichinosis condition. However larvae have been known to penetrate "foreign" tissues, and the spaces may Just possibly have been lesions l e f t by such Table 111 Date Concerning the Positive Cases Hospital Age Sex Racial Residence i n Military Condition of Number per Origin Vancouver Service Larvae Gram V.G.H. 74 M Italian 20 Yr. No Dead. Cysts C a l c i f i e d . 4 V.G.H. 71 M Hungarian 6 Yr. No Dead and fragmented Cysts & 70 worms cal c i f i e d * V.G.H. 50 F English ? No Mixed dead and alive not 1 calcified* V.G.H. 67 M German ? No Dead. Worms ca l c i f i e d cysts • N/A not. 1 V.G.H. 6 weeks M ?> 6 weeks Dead or absent. Cysts calcif i e d or not. 22 V.G.H. *? 69 M Spanish Pt. Alberni ? Mixed dead and alive worms 37Yro & cysts oalcified or not. 9 V.G.H. *? 61 F English 24 Yr. No Dead or absent worms & cysts oalcified or not. 0 . 2 V.G.H* 35 F ? 21 Yr. ? Dead. Worms & cysts c a l c i f i e d or not. 5 2 . 8 V.G.H. 83 M Irish ? ? Dead & fragmented worms & cysts calcified or not. 4 . 2 V.G.Ho ? ? ? ? ? Dead. Worms & cysts c a l . or not 2 . 4 Shaughnessy 88 M ? 11 Yr. Yes Dead. Cysts c a l . larvae not. 0 . 8 Shaughnessy 67 M ? ? Yes Dead or absent Larvae c a l . cysts not. 0 . 4 Shaughnessy 77' M ? ? Yes Dead. Cysts heavily c a l . 6 Shaughnessy 79 M ? ? Yes Alive. Cyst c a l . . 2 St. Pauls 72 M ? ? ? Dead. Cysts & larvae cal or not 8 St. Pauls 67 F ? 10 Yr. ? Dead. Larvae cal., cysts not • 0 .2 * 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. - 2 2 invading larvae. Brain lesions caused by Trichinae are discussed by Gould. Particular note was taken of the number of positive oases having histories of rheumatic conditions. In this regard i t was found that only two of the sixteen had re-corded histories of ar t h r i t i c conditions and that one of these was found at autopsy to be an osteoarthritis of the spine. The other, the sixty-seven year old German male had a seven year old history of a r t h r i t i s in the pectoral-scapular region with pronounced atrophy of the muscles. While the latter case might possibly have some significance, these results oertainly do not offer any evidence toward incriminating trichinosis as a factor in rheumatic conditions. The appearance of Triohinae in the diaphragm of a six weeks old baby presented a noteworthy case. Early writers believed that intrauterine infection did not take place. However Roth 1935, 1936, (In Gould) demonstrated muscle triohinae in the foetuses of guinea pigs Infected during pregnancy, and Kuitunen Ekbaum (1941) reported finding four li v e larvae in 6.5 grams of diaphragm tissue from a seven months old foetus. Dr. L. Ranta (Personal communication) desoribed the finding of a heavy infection of trichinae in a foetus at autopsy in Toronto. In the present case, the baby was born five weeks prematurely in a f r a i l condition, . He fed poorly and - 23 -coughed a lot, and X-ray showed increasing opacity in the right lung, Six weeks after birth the baby died, and an autopsy was performed. The report showed bronchitis, partial obstruction of the right side of the bronchial tree, and insipient croup. The writer contacted the physicians who had attended the confinement and the child's illness, and was informed that the mother, a seventeen year old Canadian woman, had suffered no illness 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 oaloification. Ehrhardt quoted by Gould, observed in 1896, death of muscle larvae as early as the third week, and larval calcification between the forty-second and sixtieth days. Similarly Leukart (In Gould) demonstrated calcification as early as the eightieth day. Nonetheless, the process of calcification does not usually commence u n t i l after the sixth month of infection, and the writer feels that this case may indicate a difference in the rate of calcification in foetus over that in the adult. Comparison of local findings with those obtained elsewhere The incidence of human trichinosis i s governed by two main factors; v i z . care exercised in raising hogs and pre-paring pork products, and food and cooking habits. Table IV presents a picture of the incidence in humans in various areas, unfortunately no mathematical corre-lations 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 situation i s com-plicated by.the fact that hog surveys which are so i n t i -mately tied up with human surveys, are in places lacking, and in other plaoes not representative of the pork which appears on the public market. The latter case exists locally. The author's information on this subject comes from personal communications with Sr. I. W. Moynihan who has a trichinosis survey of rats and hogs i n the Vancouver area in progress. The figures to be used represent only approximate values, since the mentioned surveys aire not com-plete, and furthermore do not represent a cross section of British Columbia's market pork, inasmuch as they refer largely to British Columbia raised hogs, whereas a great portion of the pork consumed i n British Columbia i s raised in the prairie Provinces. Table IV Comparison of Local Findings with those Obtained Elsewhere Size of Sample $ Positive Germany ? 1.5$ U. S. A . 5213 16.1$ California 129 17.8$ Oregon 33 12.1$ Washington 200 20. $ Canada 1359 2.2$ Montreal 539 1.5$ Toronto 410 . 1.75$ Vancouver 400 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 ) . As opposed to this Gould states that the hog incidence in the U. S. A* i s between one and two percent, giving a ratio 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 in Germany, sponsors a much lower human incidence than does a system of poor hog control as practiced in the U. S. A* The above figures also demonstrate that food habits conducive to Trichina infection can so enlarge the ratio of hog infection to human infeotion 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, five hundred fifty-nine 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 ratio would indioate that human trichinosis in Canada i s largely a result of poor hog control, and not of cooking habits. From Table IV i t i s seen that while the 4$ human incidence in Vancouver i s considerably below that of the United States, It is well above that found in Eastern Canada. 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 la t te r . However even i f compensation i s made for this discrepancy, the local incidence remains over double that found i n Montreal. Possible explanation of loca l incidence She fact that human incidenoe in 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 in this area i s considerably higher than that pre-sent in Eastern hogs. Cameron (1940) has shown an incidenoe of 0 . 5 7 $ 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 in the above six hundred ninety-one , were posi t ive . 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 in 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 Pra i r ie raised hogs - 27 -to B r i t i s h Colombia raised 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 raised hogs i s l i k e l y due to careless feeding by the farmers, but may also 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 . Thus f a r Moynihan has examined the entire diaphragms of f i v e hundred ninety rats by compression only. Of t h i s number ninety-f i v e or l 6 . 1 $ were trichinosed, 8 7 . 3 $ of the p o s i t i v e s being rats from around piggeries, and the remaining 12*7$ from around garbage dumps. This incidence i n rats i s not unusual and it... i s doubtful i f such a condition i s responsible f o r the high incidence i n hogs, although some hogs probably do become infected by eating these r a t s . The second basis 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, confining themselves to other Canadian c i t i e s . Locally 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 la r g e l y directed South across the International border. The 20$ human incidence i n the State of Washington (Ref. Table IV) would indicate a f a i r l y high hog incidence. Then considering that i t takes only one meal of trichinous - 28 -pork to cause infection, i t seems very l i k e l y that con-siderable of the local incidence may be attributed to i n -feotion contracted across the border. At the time when this survey was being organized, the author noted that nearly every medical man contacted ex-pressed the opinion that no trichinosis problem existed in the Vancouver area. This survey however shows not only that local human beings are trichinosed, but also that the incidence and in some oases the intensity i s sufficiently high to be considered a serious medical problem. It must be born in mind that the mere presence of Trichinosis represents potential epidemics and possible fat a l infections* The author believes that i f hog infections, which are apparently quite localized, were adequately controlled, and i f the public was informed of the trichinosis situation in the United States and discouraged from eating pork in that country, that human trichinosis could be v i r t u a l l y wiped out in this area. - Xt -References Cameron, T. W. M. Cameron, T. W. M. Trichinosis Can. Jour. Comp. . 1: pp. 5^14, 1937 Cameron, T. W. M. Cameron, T. W. M. Gould, S. E. Gursch, 0. F. Hall, Kerr, K. Investigations on Trichinosis in Canada. I. A Preliminary Survey of the Incidence of Trlohinella s p i r a l i s i n Hogs i n Eastern Canada. Can. Jour. Res., D, 16: pp. 89-92, 1938 Investigations on Trichinosis i n Canada. III. On the Incidenoe of Triohinosis i n Garbage-Fed Hogs. Can. Jour. Res., D, 18: pp. 83-85, 1940 Studies on Triohinosis. IV. Human Incidence i n Montreal. Can. Jour. Res. 21: pp. 413-414, 1943 Triohinosis, pp. 1-290 Charles C. Thomas and Co., Springfield, I l l i n o i s , U.S.A., 1945 Effect of digestion and refrigeration on the a b i l i t y of Trlohinella s p i r a l i s to infect rats. Jour. Parasitol. 34: pp. 394-395, 1948 Studies on Trichinosis, VI. Epidemio-logical Aspects of Trichinosis i n the United States as Indicated by an Examination of 1,000 Diaphragms, for Trichinae. U.S. Pab. Health Repts. 53: pp. 1086-1105, 1938 B., Jacobs, L., and Cuvillier, E. Studies on Trichinosis. XIII. The Inci-denoe of Human Infection with Trichinae as Indicated by Post-Mortem Examination of 3,000 Diaphragms from Washington, D.C., and Five Eastern Seaboard Cities. U.S. Pab. Health Repts. 56: pp. 836-855, 1941 C. - 3 o -Kuitunen-Ekbaum, E. The Inoidenoe of Trichinosis in Humans in Toronto. Findings in 420 Autopsies* Can. Pub. Health Jour. 22: pp.569-576, 1941 — Rioker, W. E. The Conoept of Confidence or Fiducial 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. Arctic 1: pp. 144, 1948 St o l l , N. R. This Wormy World. Jour Parasitol. 33: pp. 1-18, 1947 Todd, F. A* Triohina Inspection in Germany. Vet. Med. 43: 1948 Wright, W. H. and Bozicevich, J. Experiments in the cooking of Garbage for the Destruction of Trichinae i n Pork Scraps. 1 ' tT.S. Publ. Health Repts. 58: pp. 396-404, 1943 Wright, W. H., Kerr, K. B., and Jacobs, L. Studies on Trichinosis. XV. Summary of the Findings of Triohinella 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. 1293-1313, 1943 Fig.. 1 Compressor used during the early part of the survey with broken top plate. Fig. 2 Compressor used during the latter part ' of the survey disassembled, f i g u r e 1. figure 2 f i g . 2 Brass framed compressor p a r t i a l l y -assembled. F i g . 4 Brass framed compressor assembled. F i g . 5 T r i c h i n a l a r v a e i n r a t muscle p r i o r to e n c a p s u l a t i o n X 8 0 F i g . 6 T r i c h i n a oyst i n human muscle showing p o l a r f a t bodies X80 Figure 5. F i g u r e 6. C a l c i f i e d c y s t i n human muscle cracked by compression 180 Human muscle c o n t a i n i n g s p i n d l e shaped c y s t . Both l a r v a and cyst c a l c i f i e d X80 P i g . 6 H e a v i l y c a l c i f i e d l a r v a i n u n c a l c i f i e d c y s t . Note double w a l l o f the c y s t 188 P i g . 10 Human muscle 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 fragmented and c a l c i f i e d X80 F i g . 11 U n c a l o i f i e d cyst c o n t a i n i n g broken and c a l o i f i e d l a r v a X80 F i g . 12 Dead and c a l c i f i e d l a r v a with cyst w a l l p r a c t i c a l l y absent, i n human muaole X80 F i g u r e 11. F i g u r e 12, F i g . 12 C a l c i f i e d c y s t w i t h l a r v a absent X80 F i g u r e 13. 

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