TH _ AN EPIDEMIOLOGICAL SURVEY OF OVINE COCCIDIOSIS. The-sis. fer fin Dag? a? M. S. .WCWGAM STATE COLLEGE 7.949

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1 i\ 1 IHIHI NIH I HIHI 1 I l li TH _ AN EPIDEMIOLOGICAL SURVEY OF OVINE COCCIDIOSIS The-sis. fer fin Dag? a? M. S..WCWGAM STATE COLLEGE Jack Sa ~s fifi fl [Dunlap 7.949

2 THESIS This is to certifg that the thesis entitled An Epidemiological Survey of Cvine Coccidiosis presented by Jack Sherwin Dunlap has been accepted towards fulfillment of the requirements for,j Lfi. V. degree in_l'_arasitology fl ;' Jiliawlfiuim ajor professor Date, gmmw 1 j 1 3:19 M-795

3 LN EPIDBIIOLOGICAL SURVEY OF OVINE COCCIDIOBIS By Jack Sherwin Dunlap A Submitted to the School of Gredncte Studiee of Michigan State College of Agriculture end Applied Science in pertiel fulfillment of the requirements for the degree of MASTR 0F SCIHCB Depu-tnent ct Bectericlcgy end Public Heelth 1919

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5 TABLE OF CONTENTS Acknowledgement...o...o..oo.o...o.. 1 Introduction... 1 Review of Litereture...o...o...o.. 2 literiele end lethode... 9 Discussion...t...o...o Summary.o...o......; References eeeeeeeeeeeeeeeeeeeeeeeeeeeeeeee 21 n fififit Fm?A} 4-? «ix-v l

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7 Acknowledgement The writer wishes to express his thanks to Dr. P. A. Eakins for his guidance end criticism during this experiment. -1- Acknowledgement is nice nede to the mini Husbendry Depertnent for their essistenc e in cere end hendling of the sheep.

8 INTRODUCTION Cocoidiosis is primarily a disease of sheep in feed lots where large numbers of young animals are held in relatively restricted quarters for the purpose of fattening for market. In the feed lots where outbreaks have occurred they are associated with overcrowding and poor sanitation. Outbreaks do occur in places other than feed lots where there is an overcrowding of young susceptible animls. The source of infection to the lambs is the ewe. The ewes act as carriers, passing a few oocysts at all times. The length of time that is necessary for the sporulation of the oocysts outside the body before they become infective is dependent upon many factors, among them humidity, temperature, and the species of coccidia. Under laboratory conditions this interval has been from twenty-four hours up to one hundred and twenty hours for the different species under optimum conditions. This experiment was undertaken for the purpose of studying the natural infection of lambs as to time and course as shown by oocyst counts. The period at which the first sporulated oocysts were found in the bedding was determined as well as numbers present in the bedding. The sheep under observation were a small flock of crossbred yearlings, ewes, and the lube born to these ewes. The period of observation was from the first of January till the end of August and during this tires the flock was both in the barn and on pasture. At no time during this period of observation were there any deaths in the lambs attributable to cocoidiosis or any symptoms of clinical coooidiosis manifested by the lambs.

9 mm or mummy The discovery of the microscope by Leeuwenhoek and his description of bodies from the bile duct of rabbits is apparently the first observation of a parasitic protozoan. It is believed that the bodies seen by Leeuwenhoek were the oocysts of limeria stiedae. In sheep the first mention of coccidia was made by Leuokart in 1879 and by Rivolta in the same year. In the United States coccidia were first reported from white patches noted in the intestines of sheep by Curtice (1892) and Stiles (1892). Becker (19314.) describing the coccidia of animals listed seven species of coccidia from sheep and goats, five of which are considered valid and two of which require further study. Two new species were added by Christensen, (1938) and the two doubtful species were dropped, leaving seven as the val id number. Honess (1912) described two new species, one from the bighorn sheep and the other from both the bighorn and domestic sheep. The following list of coccidia occurring in sheep is given by Morgan and Hawkins (19148) and it was used as the basis for describing the species in this paper. Bimeria pallida - Christensen, The oocysts measure 12 to 20 microns leug (average 1h.2 microns) by 8 to 15 microns wide (average 10 microns). The oocyst is ellipsoidal to subspherical, it possesses no polar cap and the micrcpyle is imperceptible. A single dark refraction line between oocyst membrane and inner surface of the cyst wall ' can be seen. The early spherical sporont is sparsely and irregularly granular, pale and inconspicuous. Sporulation occurs within 21; hours.

10 -3- Eimeria m; - Iotlan, Mossy and Vajda, The oocysts measure 12 to 22 microns long (average 16.5 microns) by 10 to 18 microns wide (average microns). The shape of the oocyst is ellipsoidal to subspherical with no perceptible micropyle and no polar cap. The cyst wall is demarcated on each side by a heavy black refraction line, giving a characteristic double-contoured appearance. The early spherical sporont is clear cut with refractile granules and globules in an irregular network. The sporulation time is 214 to 148 hours. Dimeria pk kphl-nkimov - Yakimov and Rastegaeva, The oocysts are 20 to 28 microns long (average 23.1 microns) by 15 to 20 microns wide (average 18.3 microns). The shape is ellipsoidal, occasionally slightly ovoid, with the micropyle usually imperceptible and there is no polar cap. A single dark refraction line marks the intersurface between oocyst membrane and inside surface of the cyst wall. The early spherical sporont is pale and may sometimes appear to be a delicate salmon pink. (This coloration may be affected by the light source.) Sporulation occurs in 24 to h8 hours. limeria Houssu and llarotel, The oocysts measure 25 to 33 microns long (average 28.9 microns) by 18 to 2h microns. wide (average 21 microns). The shape is characteristically like a hen's egg with a distinct micropyle at the narrow end. There is no polar cap. The boundary between the oocyst membrane and the inner surface of the cyst wall is Imrked by a single refraction line. The oocyst may appear a delicate salmon pink to a pale yellowish brown. The sporulation time is 214. to 148 hours. limeria arlcingi - Marotel, Oocysts measure 1;? to L2 microns long (average 27 microns) by 13 to 27 microns wide (average

11 18 microns). The shape 1. ellipsoidal with a well developed micropyle covered by a polar cap. A single, heavy, black contour line is present between oocyst membrane and inner surface wall. The early spherical sporont is pale. Sporulation occurs in 214 to 72 hours. limeria granulosa - Christensen, The oocysts are 22 to 35 microns long (average microns) by 17 to 25 microns wide (average 20.9 microns). Their shape is that of a broad shouldered urn or pyriform. There is a distinct micropyle covered-by a polar cap. The boundary between the oocyst membrane and the inner surface of the cyst wall is marked by a single black refraction line. Early spherical sporontstare characteristically dense and uniformly granular. Sporulation time is 72 to 96 hours. limeria intricata - Spiegl, The oocysts measure 39 to 53 microns long (average 147 microns) by 27 to 314 microns wide (average 32 microns). The shape is ellipsoidal with a wide micropyle under the polar cap. A single dark refraction line is present between the oocyst membrane and inner surface of the wall. The wall is characteristically opaque, transversely striated, yellowish brown to dark brown in color, and thickened to 2 to 3 microns. Sporulation time is 72 to 120 hours. Bimeria ah-sa-ta - Honess, The oocysts are to microns long (average microns) by to microns wide (average 23.2 microns). and polar cap present. Their shape is ellipsoidal with a micropyle A single dark refraction line shows at the boundary of the oocyst membrane and the inner surface of the wall. Bimeria crandallis - Honess, Measurements of the oocysts

12 are to microns long (average microns) by to microns wide (average microns). The shape is ellipsoidal with a polar cap present. The oocyst's wall presents a double contoured appearance being demarcated on each side by a dark refraction line. A survey of the literature by Thomson and Hall (1931) revealed that the same species of coccidia were found in sheep and goats. Studies by Baloset (1932) showed that the infections of coccidia in sheep and goats were by species comon to both. Kansas (1942) showed the same to be true of the domestic and bighorn sheep in Wyoming with the exception of _E_. crandgalliis which was only found in the bighorn sheep. The finding of oocysts in domestic lambs at Michigan State College by Hawkins (unpublished data) and by the author, which very closely resemble 5. crandallis. tend to show that this species is also common to both domestic and wild sheep. 1hrtin (1923) reported losses from coccidiosis starting about two weeks after the purchase of the lambs. Outbreaks in the feed lets have been studied by Thorp (1938) and been and Thorp ( 1938) showing that symptoms appear two to three weeks after the lambs' arrival in the lots. The lambs were passing a few oocysts upon their arrival in the lots, and the numbers gradually rose to a peak in one to two weeks, then declined fairly rapidly. Christensen (19140) reported an outbreak in a feed lot with death losses and securing beginning 12 to 16 days after. the start of feeding and ceasing after the clinical symptoms had lasted about two weeks. The mortality was about 3.14 per cent. Studies as to the source of the outbreak showed tint the lambs were passing oocysts upon entering the feed lot, and that the outbreak was not due

13 to oocysts remaining from the previous spring, as these oocysts did not appear viable and were not sporulated. The infection in this case was traced to the feed and the water was not incriminated as there was always a fresh clean supply. Examinations showed that the feeding of silage provided ideal conditions - moisture for the rapid development of sporulated oocysts and fine enough consistency so that the oocysts did not filter through the feed. The feeding of chopped alfalfa, corn and molasses also'was incriminated as a medium.for the sporulation of the oocysts, but the moisture was not as high as in the silage and sporulation'was somewhat reduced. Sporulation was found to take place in fecal pellets before complete dessication occurred, but the number of sporulated oocysts was too low to account for massive infections. In this outbreak the fact that silage was fed in troughs that'were not cleaned provided ideal conditions for the outbreak. Studies on the sporulation and viability of E. arloingi made by Christensen (1939) showed that oocysts in clean water sporulated slowly at near freezing temperatures (0"to 5 C.) and rapidly at roomttemperature (20' to 25 C.). If the temperature was raised to 32 C. sporulation'was accelerated but segmentation'was abnormal, and at h0' C. they failed to show signs of sporulation. In fecal sediment there'was no evidence of sporulation at near-freezing, room, or incubator temperatures even though the oocysts'were covered with a shallow layer of water. However, 20 per cent of the unsporulated oocysts were still viable after ten months at near-freezing temperatures. The failure to sporulate in the presence of fecal sediment'was attributed to lowered oxygen tension resulting from putrefaction. Apparently this is also the reason that sporulation does not take place until the oocysts

14 have been passed from the body and a more favorable oxygen tension is obtained. Inside of fecal pellets in.which the moisture was preserved, oocysts showed little sporulation at near-freezing temperature, abundant and rapid sporulation at room temperature, and no sporulation at he C. The drying of fecal pellets at room temperature produced many sporulated oocysts before dcssicetion wrinkled and shrank the oocysts beyond recognition, and Christensen concluded that drying was probably fatal to oocysts within several days to a few weeks. The pathology of coccidiosis was described by Stiles (1892) as being irregular, slightly elevated,'whitish patches on the mucosa of the small intestine, these patches being three-fourths to one inch in diameter. The epithelial cells lining the villi m. very large and every one contained one or more oocysts. Nocard (1893) reported a case, (read before the International Congress of Hygiene and Demography, London, 1891) from.a three month old lamb in which there were true polypi, which on section showed the glandular epithelial cells to contain parasites. lhrtin (1923) found that lesions from.coccidicsis were confined to the small intestines and were small white areas. visible to the naked eye fron both serous and mucous surfaces. Hieroscopic examination of the small intestine showed large masses of coccidia in various stages of their cycle. The coccidiumnwae at thnt time identified ". h EE 219 Richardson (19h8) reported that the mucous membrane of the small intestine'was thickened and edematous. The capillary vessels are congested and hemorrhages may occur. In the more chronic cases there was a proliferation of the epithelium of the small intestine to form.papilliform.elevations. In the acute type of coccidiosis, Newsom and Thorp (1938) reported enteritis present in

15 ~8- the large intestinee,and the mucous membrane in the cecum frequently was streaked with grey, as fairly typical lesions. In the more chronic cases the intestinal wall is much thickened. Treatment of coccidiosis has been studiedlby Hawkins, :3 3:}. (19143) using sulfaguanidine and sulfasuxidine. These workers found them to be of some value in the treatment of natural outbreaks if the treatment was instigated early in the course of the infection. Boflund and Koffman ( )7 found sulftalyl to be superior to sulfanilamide or nicotine-copper-sulfate when used against coccidiosis. Sulftalyl is 2-ftalylsulfanilamidotiazol. delay or prevent development. The sulftalyl acted upon the oocysts to Christensen and Foster (19143) used sulfaguanidine in experimentally produced coccidiosis in lambs. They reported good results'when used as a prophylactic, 0.2 per cent by weight in the feed for fienty days. When the lambs were fed sulfaguanidine as a therapeutic agent in the concentration of 0.15 per cent in ration upon the first appearance of symptoms their results showed no value in the small number of groups tested. The use of comercially ground sulfur as a prophylactic agent against. natural outbreaks of coccidiosis in feeder lambs was studied by Christensen (191414). The use of sulfur in percentages of 0.5 per cent to 3.0 per cent of the total ration effectively prevented naturally acquired clinical coccidiosis with no visible ill effects noted in the lambs after 72 days of continuous feeding. The rate of fattening compared favorably with the untreated controls.

16 MATERIALS AND MODS The lambs studied in this report were born to crossbred ewes which with yearlings made up an experimental group for the study of sheep parasites. The sheep in this group received no phenothiazine either as a drench or in the salt mixture. With this exception they were fed the same ration'whioh'was used for the college flock, alfalfa hay, grains, bran and corn silage being used in the ration. During the 'winter and early spring months the sheep were housed in an experimental barn in pens providing a center feed rack and a trough and a small water well. 'The pens opened to a small outside enclosure to which the sheep had free access except in the severest weether when.the doors were closed to prevent storms fron blowing into the barn and during nights when the temperature'was very low. Two pens were utilised in the barns, pen A was composed of wethers and pen 3 was composed of yearling and aged ewes. Six lambs born to the latter group were the lambs under observation, the first being born on the 29th of February, l9h8 and the last one on the 26th of March, l9h8, Table IV; Bedding in the pens consisted of straw'and a small amount of hay'which'was pulled from.the racks. Fresh bedding was added as necessary'when it became trampled down, soiled with feces cr'wet. This meant that the height of the bedding was built up, and twice during the winter it was necessary to remove it. The sheep were taken from.the barn the week of the 12th of June and placed on.a clean pasture which had not been used by sheep for at least three years: pens A.end B'were combined on the pasture. A recording thermometer was placed in pen B at the level of the

17 -10- bedding to record the temperature in the barn. The monthly report from the United States Isather Bureau, East Lansing was used for the mean temperature outside the barn, (Table II). kamination of the bedding for oocysts was started on the 29th of December, 1914.? and continued at weekly intervals until the 214th of May, l9h8. The following technic was adapted from the technic of Freitas (19h7). Separate samples were obtained from each pen by gathering the bedding in pails, approximately 1400 grams were taken at random from over the entire surface. Feces in so far as possible 'were shaken out of the bedding. The samples'were taken to the laboratory and mixed so that all sections of the pen were equally represented in the part that was removed. Samples of 200 grams were weighed from each of the pens and placed in large enamel buckets, covered with tap water, and a weighted wire screen was placed on top to keep the straw under water. The buckets were then placed in the I refrigerator to macerate over night. It'was necessary to keep samples in the refrigerator in order that results might not be affected by sporulation after the samples were collected. Atter maceration the straw was removed and placed in a large funnel, which contained a small piece of wire mesh at the narrow end, washed with at least 2000 milliliters ef'water in three washings and allowed to drain. The'wash water was added to the original water from maceration and allowed to settle over night in the refrigerator. The supernatant fluid was then raoved by means of a water aspirator and the sediment was transferred to graduates. Here again the buckets were rinsed three times. Sedimentation was carried out again and the volume reduced further to less than 250 milliliters. Appropriate amounts of the sediment were trans-

18 ferred to 15 milliliter centrifuge tubes and a sugar solution, composed of 500 grams of sugar dissolved in 360 milliliters water, was added to the top of the tube. A cover slip was placed on the top of the tube so that a small bubble was formed under the cover slip. The percentage of sedinent to sugar solution was not over one part to seven; this was found necessary to reduce the amount of debris on the cover slip so as not to obscure the oocysts, and also to prevent too great lowering of the specific gravity of the solution. The material was then centrifuged at 1000 rpm for one minute and the cover slip removed by lifting vertically and placed on a slide. The entire surface of the cover slip was examined, identification was made by the aid of a key (Table I) modified from Morgan and Hawkins (19148). Duplicate counts were made on each sample and the averages were used in Figures 1 and 2. Focal examinations of the lambs started one week after their birth. In order to obtain feces from the young lambs without contamination fromnthe bedding, small cloth pockets, as described by Goldsby and Eveleth (1%), were tied over the anus and allowed to remain there for four to six hours. After the lambs had reached sufficient siso the feces were obtained directly from the rectum. The fecal samples were taken to the laboratory and four gram samples were weighed into 125 milliliter bottles to which 56 milliliters of water were added. The mixture was then stirred for approximately one minute with a triple paddle stirrer which was attached to a l/h inch electric hand drill, adapted from Kausal and Gordon (1941). Fifteen-hundredths milliliter of the suspension was transferred to a 15 milliliter centrifuge tube and sugar solution added to the top; care was exercised that

19 the solution did not run over the top of the tube when the cover slip was placed there and again when the cover slip was removed after one minute centrifugation at 1000 revolutions per minute as described by Stoll (1930) and Hawkins, _e_t_,_1_ (19th). Examination of the entire cover slip was carried out for the oocysts and they were identified. In the cases where the number of oocysts was excessive higher dilutions of the original mixture was made and flotation made again. The average for the six lambs was plotted as to total number and by species in Figures 3, h, 5 and 6. The counts were made with a Bausch and Tomb Compound Binocular microscope using an American Optical Model 370, 100 watt lamp with a Corning Daylight ground glass filter. The microscope was calibrated with an ocular micrometer for checking the lengths during the counts. leasurements of oocysts were made with the aid of Leits Filer ocular micrometer calibrated to tenths of a micron and interpolations were made to hundredths of a micron.

20 -13- DISCUSSION The identification of a parasite by the size and shape of the ovum or cyst is often quite difficult and frequently not accurate as closely related parasites may pass cysts and ova of very nearly the same shape and size. This inaccuracy becomes especially apparent 'when one is surveying a number of samples where mdnute detailed study of each individual is impractical. This is the case with the coocidial oocysts, where the life cycle should be the most important criterion for the determination of the species. Tyszer (1932), in regard to avian coccidiosis, cites certain criteria that should be considered with respect to determining the I.number of species that infect a specific host. In cases where the identification is dependent upon measurements and slight morphological characteristics the acceptance of a new species should depend upon the application of these criteria: single oocyst infections and the determination of the complete life cycles from such infections, determination of the sporulation time, the location of pathology in the animal'with pure infections, and the use of immunity experiments. The use of measurements alone leads one to fallacies as Jones (1932) obtained bimodal curves from.single oocyst infections and this bimodal effect was maintained throughout successive transfers. The use of measurements as a criterion of speciee should only be valid when a sufficiently large number of measurements gives a symmetrical and continuous curve. However, it is doubtful if measurements alone should ever serve to differentiate species. Richardson (l9h8) maintains a similar opinion on the establishment of species on the characters of

21 the oocysts alone. The coccidia in sheep have been separated into nine species based upon size, shape, presence or absence of structures, color and time of sporulation, as the individual life cycles have not been studied. The use of the above criteria in the examination of fecal samples for the per cent infection by species leads to a large number of oocysts being placed in the category of unknown. The unknown group contains not only oocysts distorted by natural conditions but also those that had debris over the ends, lying in a plane other than parallel to the slide, those'with apparent loss of the polar cap, lack of coloration, or failure of the refraction lines to be distinct. It was also noted by the author that changing the light source or by focusing the condensing lens of the lamp one could impart a delicate salmon pink that is associated with the oocysts of E. Egg-fl;- ygkimovi, hence the same lamp was used throughout the survey. The loss of the polar cap by _E_. crandallis would give oocysts an appearance very similar to that of 5. m, and the loss of the polar cap from'g. arloipgi would result in an oocyst of the same general appearance of E. gang-ee-ekimovi. with these facts in mind only the species that were encountered in large numbers or'were identified with certainty are included in the graphs; namely, _E_. arloiggi, E. crandall- _i_ _, g.m! 5. pgllida, g. faurei, and _E_. intricata. In addition to the above species 5. gins-m-zak imovi and 5. granulosa were identified by morphology as being present during the course of the study, however, E was not identified and if present it was class i- fied as 5. arloingi. A11 cysts not showing very definite characteristics of the species are classified as unknown and are included in

22 -15.. the total counts. 5. arloiggi and g.m were found to be the two most frequently encountered species both as to duration of passage and in numbers passed. 5. pallida, E. crandallis and E. M were next in frequency and were found with considerable constancy throughout the period studied. E. intricate appeared sporadically and in small numbers. _E_. granulosa was identified twice during the experiment, once when two oocysts were identified from one 'lamb on the 25rd of April and the second time was when an oocyst from pen B was identified from the bedding as that species on the 10th of May. At no time during the course of observation did any of the lambs pass oocysts of a single species. Measurements of E. arloingi and _E_. crandallis (25 oocysts each) separated on the basis of the double contoured oocyst of E. crandallis gave the following: A E. arlcigi E. crandallis Length to microns to microns Width to microns to microns Mean to microns to microns m range 1.25 to to 1.61 M mean In both species the data obtained here differs from that observed by Honess (19142) in that the oocysts tended to be longer and narrower in each case. kaminati on of the bedding showed that pen A, the yearlings, had higher counts than pen B, the ewes, until the latter part of April

23 -16.. when the lambs were passing oocysts in pen B which was then occupied jointly by the ewes and their lambs, (figure 1). The first sporulated oocysts were observed in pen B on the 12th of April at which time the mean weekly temperature in the barn at bedding level was 149 F. and the mean temperature for the proceeding week was 118 F. This was the first time since the 29th of December that the mean temperature had risen above 115 F., and the mean remained above this temperature for the remaining time of the experiment (figure 2). The demonstration of sporulated oocysts from the bedding the first time on the 12th of April was after one of the lambs had passed oocysts. The oocysts were passed by a single lamb on the 9th of April, a second lamb started passing oocysts on the 16th of April and the remaining four lambs passed oocysts for the first time on the 25rd of April, Table IV. This shows that the method used fa' the detection of the oocysts in the bedding was either not sensitive enough to recover the very low percentage of sporulated oocysts necessary for infection or that sporulated oocysts occurred first in the feed troughs resulting in the infections. A third possibility is that of contamination of the udders of the ewes. Christensen' s work (1959) showed that some sporulation takes place at near-freesing temperature inside of fecal pellets. This small number of sporulated oocysts is apparently sufficient to cause the very early infections. Study of the reports on natural outbreaks tends to show that the life cycle in the sheep is probably less than two. to three weeks as by that time the peak of the oocyst discharge is reached and hence at least one and possibly more cycles, if this occurs, have been completed. Attempting to correlate this with the initial passing of oocysts, it would seem that

24 -17- the lamb was infected some time in the latter part of March, a considerable time before the demonstration of sporulated oocysts in the bedding. However, during this period and for the whole of the month of March the mean weekly temperature was below 140 I. only once. It will be necessary to study the course in a more severe winter to determine how low the temperature would have to be to prevent infections in the lambs. Comparison of the average total counts of the ewes and the lambs (figure 3) showed that the ewes remained at a consistently] low level throughout the period of examination, although they were in the same pen and on the same pasture as the lambs. During the period the lambs were in the barn the counts showed a very rapid rise in number of oocysts from the first till the seventh week at which tine the highest peak was reached. next two weeks. This was followed by a rapid decline during the There was a second rise which reached its peak five weeks after the first and twolve weeks after the initial passing of oocysts; at this time the lambs were on pasture. Further study with coccidia free lambs would be necessary to show whether this apparent immunity of the ewes was due to acquired or an age immunity, or'to a combination of both. kamination by species (figure 4. '5. and 6) show that the peaks of the infections for the different species varied slightly and that there were secondary peaks present. E. arloi_ngi showed its highest peak five weeks after the first oocysts were passed and a second seven weeks later. E. crandallis' highest peak occurred six weeks after its first appearance and the second five weeks later. The initial peak of _E_. parva occurred six weeks after the first identification of that

25 ' species and the second six weeks later. 5. pallida reached its first peak in five weeks and the second four weeks after the first. _E_.M reach/ed its peak in three weeks, and a very slight peak occurred eight weeks later. 5. intricate showed the first peak after three weeks, the second two weeks later and a third four weeks after the second, however, in no case were these peaks very high. The individual lambs showed difference as to age at which the first oocysts were passed varying from five weeks to eight weeks old. The peaks of infection as determined by the number of oocysts varied from one week to four weeks after the first discharge of oocysts. The ages at which the peak oocyst counts were encountered varied from six weeks to twelve weeks with no apparent correlation being possible between the age and time of first oocyst discharge or peak discharge of them due to the small number of animals. The peak discharge of oocysts in the lamb which first passed oocysts was not as high as the succeeding lambs. The lamb which was second in the passing of oocysts reached a peak higher than the preceding lamb but not as high as the four remaining lambs which.11 passed their first oocysts the same week. Hypothetically this could. be partially explained on the grounds that the first lamb acted as a multiplier" of the infection. The fact that the young susceptible 1.2m play the role of "multipliers" is substantiated by figure 1 which shows that the number of oocysts in pan B greatly increase during the time of peak discharge of oocysts by the lambs. Until this time the ewes, acting as carriers, maintained a low level of potential infection in the bedding which was one source of infection for the lambs. The lambs build the

26 -19. potential infection to a higher level without a substantial increase in production of oocyst by the ewes.

27 -20- SUMMARY 1. This study shows that a mean temperature in the barn at bedding level of approximately I49 I". is necessary for the demonstration of sporulated oocysts in the bedding. 2. The bedding is a potential source of cocoidial infection in the young lambs. 3. Natural infections are usually of a mixed nature with E. srlcingi and E. parva being the most frequently encountered species, although all valid species are apparently present. 11. There is a relative immunity developed in the older sheep, with the older sheep acting as carriers and the young lambs acting as "mltipliers' of the infection. 5. There was no apparent correlation between age of lamb and initial discharge of oocysts or peak of oocyst discharge.

28 -21- REFERENCES Baloset, L The Coccidia of the Sheep and Goat's DeveloPmental cycle of Eimeria pig-w-yakimov. Yakimov and Rastegaieva, Tunis Arch. Inst. Pasteur, 2. Becker, Elery R. 193k. Coccidia and Cocoidicsis of Domesticated, Game and Laboratory Animals and of man. Collegiate Press, Inc., Ames, Iowa. 3. Christensen, J. F Species Differentiation in the Coccidia from the Domestic Sheep. Jour. Parasitol. h. 2h: h53-h67. Christensen, J. F Sporulation and Viability of Oocysts of Rimeria arloingi from.thc Domestic Sheep. Jour. Agric. Res. 59: h. 5. Christensen, J. F. 19h0. The source and Awailability of Infective Oocysts in an Outbreak of Cocoidicsis in Lambs in Nebraska Feedlots. Amer. Jour. Vet. Res. 1: Christensen, J. F. 19hh. Sulfur Prophylaxis of Cocoidicsis of Feeder Lambs. Amer. Jour. Vet. Res. 5: 3hl~3h5. 7. Christensen, J. F. and A. 0. Foster. 19h3. Further Studies with Sulfaguanidine in the Control of Ovine Cocoidicsis. vet. use hh91h7. Curtice, C Parasites. Jour. Comp. use. 13: Deem,.A. W}.and F. Thorp, Jr variation in Numbers of Coccidia in Lambs During the Feeding Season. vet. med.

29 314: Freitas, M. G Overwintering of the Nematode Parasites of Sheep with Notes on the Course of the Infection. Thesis for the Degree of M. 8., School of Graduate Studies, Michigan State College. 11. Goldsby, A. I. and D. F. Evcleth Diagnosis of Internal Parasites of Sheep. Vet. Med. 1.11: Hawkins, P. A., C. 1.. Cole and F. Thorp, Jr. 191;}. The Effects of Sulfaguanidine and Sulfasuxidine in a Natural Outbreak of Ovine Cocoidicsis. Vet. Med. 38: Hawkins, P. A., C. L. Cole, H. E. Kline and J. H. Drudge Studies of Sheep Parasites I. The Course of Untreated Nematode Infections. Vet. Med. 39: Hoflund, S. och M. Hoffman On parasitsjukdomar och deras beka'impende hos vara vanligaste husdjur. Sulftalyl, ett gott model not ccccidios. Meddelanden fran Kungl. Veteringrhggskolan I Stockholm, Argang. 21: Honess, R. F Coccidia Infesting the Rocky Mountain Bighorn Sheep in Wyoming with Description of Two New Species. Univ. Wyoming Bulletin No pp. 16. Jones, E. R Size as a Species Characteristic in Coccidia: Variation Under Diverse Conditions of Infections. Archiv fur Prot. 76: Kauzal, G. P. and H. McL. Gordon A Useful Mixing Apparatus for the Preparation of Suspensions of Feces for Helminthological Examinations. Jour. Council Ind.

30 .25- Res. 1h: 30h s Martin, H. M Cocoidicsis in Sheep. North Amer. Vet. in Morgan, B. B. and P. A. Hawkins Veterinary Protosoology. Burgess Publishing 00., Minneapolis, Minn. 195 pp- 20. Newsom, I. E. and F. Thorp, Jr Lamb Diseases in Colorado Feedlots. Bulletin No. hh8. Colorado Agric. Exp. Station L2 pp. 21. Nocard, B Cocoidial Tumours from the Small Intestine of the Sheep. Jour. Path. Bacty. 1: h0hah05. Richardson, U. F. 19hBt Veterinary Protoscology. Oliver and Boyd, Edinburgh: Tweeddale Court. 2h0 pp. 23. Stiles, C. I Notes on Parasites 10- A Case of Intestinal Cocoidicsis in Sheep. Jour. Comp. Med. 13: Stoll, N. R On Methods of Counting Nematode Ova in Sheep Dung. Parasitol. 22: Thomson, J. G. and G. N. Hall Observation on Intestinal Cocoidicsis of Sheep in Northern Nigeria. Jour. Trop. Med. Hyg. 3h: Thorp, F., Jr Some Feedlot Diseases of Lambs. Vet. Med. 33: )... Tyszer, E. E Criteria and Methods in the Investigation of Avian Cocoidicsis. Sci. 75:

31 Table I KEY TO OOCYSTS OF EIMERIA IN SHEEP 1. Polar cap absent POlarctppras'ont-Daub-u Ellipsoidal, micropyle inconspicuous Egg-shaped, micropyle conspicuous at narrow end - - -'. faurei 3. Ellipsoidal, with single refraction line marking innernll----o-u h. Ellipsoidal, with.two dark refraction lines one on each side of wall ~ parva h. Ellipsoidal to subglobular, pallid, microns 1 ng esp-sew-.ea111da Ellipsoidal, microns long 'E. nina-kohl-yakimovi 5. wall transparent, thinner than 2 microns Nell opaque, thicker than 2 microns, transversley striated, dark brown - ~ E. intricata E111P801d81--" C-O-CneI-eeue-o'z. Typically shaped like a broad shouldered urn or pyriform, cap on broad end granulosa Ellipsoidal, with two dark refraction lines one on each side of'wall E. crandallis Ellipsoidal with single refraction line marking innernll oarioini -_rl- a' a e Adapted from Morgan and Hawkins (1948)

32 0 H \cho 0.. O O 0 Table II 'Neekly Mean Temperatures East Lansing, Michigan! Degrees Farenheit Neck ending High Low Average 12 January 19L February 19h l Mkrch 19h April 19h may 19h8 10 V h 31, 7 June 19h8 1h July 19h8 I August 19h8 9 l h h h h h.0 6h h h.0 H p \N e e c sees seen asses sees 8 NO PO O (n00. cm O O O O h ; 21.0 h5.8 hl : h8.5 b3o h , h h Computed from U. 8. Dept. of Commerce, Neather Bureau, Monthly Climatological Summary, East Lansing Station.

33 Table III Lambs Average per cent infection a 9 é: 1%.1351 cran-gallis BEE: 3223 int'gicata n3?" 9 April April pm pm May 13 3S 7 6 h May May LO 28 May L June h June June June July July July July h July h August August "3:33;" e Figures for E. nina-kohl-ekimovi and E. granulcsa are included in the column marked unknown. Figures are expressed as the nearest whole number with the exception of 3. intricata.

34 Table IV Lamb ' ' Date of birth Date of 1'17 oocysts Date of 1'15 peak Date of and peak No. oocyst per gram feces 13 peak February 25 April 21 May 12 June 120, e 6 March 16 April 23 April 28 May 91, e 6 March 9 April 30 April 5 June M45 9 March 23 April 11.; May 5 June 206, March 23 April 30 April 25 June 123, mm: 25 April 21 May 25 June 268,100 a 1131 and 1152 twin lambs

35 PEN A *PEN B 1} z'ua'oj-ajujaajv MARCH APRIL MAY I In FIGURE A A - v- r w _ fl.4 A L A _ a'. : a ' 1'. t :5 A JANUARY FEBRUARY 9N 0038 :10 WVUO 83d SLSAOOO

36 A 8 ' I.) 30 3' 1' lo i APRIL 1 3HN38HVJ $ d FIGURE 2 IN BARN " h \ i I PEN A «- PEN B -- MEAN TEMPERATURE cc 7 23 FEBRUARY JANUARY 1?.1. SJ. SAOOO 031 V'IOBOdS.LNBOBBd

37 GRAM 0F F 55 OOCYSTS PE R E E s s E s H FIGURE 3 LAMBS --- -EWES E 1"» APRIL 1'0 I MAY 0 5' JUNE 1" l '.101"

38 FIGURE 4 FEEEFEEE OOCYSTS PER GRAM PE 5 E F F FE 0F FECES g. ARLOINGI E. CRANDALLIS \ V I 3 In oh if h b I' b". at 1'7

39 , lulubw FIGURE E. PARVA PALLIDA mm m m m m m m m mm mmmmmmm muowu mo Idmo 1 mm I'-"'-"'l-""' '-"-'"-"'-'-"'" "'I o -. J 92 ~11.- " "5 1 AUGUST 4 -l

40 FIGURE 6 mmmmmmmmmmemmmmmmm mwuwu m0 Edmo mum mhm>ooo E. EALLBLL INTRICA TA

41 ... : a 1. \.i...e -rii Fl D'IIIELI'..'\I. It ID ufiaan ui.. qgeglis in)

42 Ha-.....l !.A I... ' : '3.e 111 } W. V...l ii elullllal111r.l. 141 y..1. v.1 I 1 (.14.. \ A 5 1.

43 MICHIGAN STATE UNIVE ITY I l I I I I H I I I I I I I I I I TI'ES O