- Email: [email protected]
Experimental Eimeria debliecki infections in nursing and weaned pigs
Veterinary Parasitology, 25 (1987) 39-45 Elsevier Science Publishers B.V., Amsterdam - - Printed in The Netherlands
39
Experimental E i m e r i a debliecki Infections in Nursing and Weaned Pigs DAVID S. LINDSAY, BYRON L. BLAGBURN and TIMOTHY R. BOOSINGER
Department of Pathology and Parasitology, College of Veterinary Medicine, Auburn University, AL 36849 (U.S.A.) (Accepted for publication 24 September 1986)
ABSTRACT
Lindsay, D.S., Blagburn, B.L. and Boosinger, T.R., 1987. Experimental Eimeria debliecki infections in nursing and weaned pigs. Vet. Parasitol., 25: 39-45. Three litters of six, 3-day-old nursing pigs were inoculated via a stomach tube with 8.0 X 105, 1.6 X 106 or 5.0 X 106 sporulated oocysts of Eimeria debliecki and four groups of six, 4-week-old weaned pigs were inoculated with 8.0 X 105, 1.6 X 106, 5.0 X 106 or 1.0 X 107 sporulated oocysts of E. debliecki to determine its pathogenicity. Clinical coccidiosis or deaths did not result from infections. Infections were confined to the jejunum and occasionally the duodenum. Microscopic lesions of mild to moderate villous atrophy were observed in one nursing pig given 5.0 X 106 oocysts and three weaned pigs given 1.6 x 106, &fiX 106 a n d 1.0 X 107 oocysts and examined 5 days post-inoculation. Pathogenic bacteria or viruses were not demonstrated in any pigs. Results of this study indicate that E. debliecki is not a cause of neonatal or weaning diarrhea in pigs.
INTRODUCTION
Neonatal coccidiosis in pigs is a common intestinal disease problem on swine farms in swine-producing areas of the world (Lindsay et al., 1983; Stuart and Lindsay, 1986). The disease is most often associated with Isospora suis infection in young pigs ( Stuart et al., 1978, 1980, 1982), however, reports of coccidiosis caused by Eimeria spp. in pigs of various ages have appeared in the literature (Biester and Murray, 1929; Swanson and Kates, 1940; Boch and Wiesenhutter, 1963; Rommel and Ipczynski, 1967; Rommel, 1970; Coussement et al., 1981, Hill et alo, 1985). In most of the reports that implicated Eimeria spp. as a cause of clinical coccidiosis, the authors did not examine infected pigs for bacterial or viral pathogens. Therefore, the role of Eimeria spp. as a cause of diarrhea or illness in pigs remains unclear. Surveys conducted in the U.S.A. indicate that 60-90% of swine living on pastures or dirt lots are infected with Eimeria spp. (Vetterling, 1966a; Greiner 0304-4017/87/$03.50
© 1987 Elsevier Science Publishers B.V.
40 et al., 1982; Lindsay et al., 1984 ). One study that involved only sows indicated that 90% were infected with Eimeria spp. and that 61% of these sows were passing oocysts ofEimeria debliecki in their feces (Lindsay et al., 1984 ). Because of the prevalence of E. debliecki in sow feces, oocysts should be present in most farrowing houses and, therefore, available to infect nursing pigs. The present study was conducted to determine if an isolate of E. debliecki (Lindsay et al., 1985a) was capable of causing disease in neonatal nursing or weaned pigs. MATERIALS AND M E T H O D S
Oocysts Oocysts of E. debliecki were sporulated, cleaned of fecal debris and enumerated for inoculation as previously reported (Lindsay et al., 1985a). Oocysts were stored in 2.5% (w/v) potassium dichromate solution at 4°C for < 60 days before use.
Pregnant sows with known breeding dates were obtained from a producer in southwest Georgia, U.S.A. These sows had undergone gestation on dirt lots and were positive for Eimeria spp. upon fecal examination. Sows were housed separately in Rockefeller isolation rooms and were confined in farrowing crates that were on elevated plastisol-coated floors. Sows were allowed ad libitum consumption of water and feed containing a coccidiostat (Ampro125%, Merck & Co., Rahway, NJ, U.S.A.). The feces of all sows were examined by fecal flotation and were negative for coccidial oocysts by 1-6 days prior to farrowing. Three litters of nursing pigs and four groups of 4-week-old weaned pigs were used for experimental E. debliecki infections. Common production practices were used in the management of nursing and weaned pigs. All 4-week-old pigs were obtained from sows that had farrowed in Rockefeller isolation rooms and were managed as sows used for nursing pig studies. The pigs were weaned at 3 weeks of age, housed in the farrowing crate and fed feed that did not contain a coccidiostat and water ad libitum.
Experimental inoculation Three litters of six, 3-day-old nursing pigs were inoculated via a stomach tube with 8.0X105 (Litter A), 1.6X106 (Litter B), or 5.0X106 (Litter C) sporulated oocysts of E. debliecki. Four groups of six, 4-week-old weaned pigs were inoculated via a stomach tube with 8.0 × l0 s ( Group A ), 1.6 × 106 ( Group B), 5.0×10 e (Group C), or 1.0X107 (Group D) sporulated oocysts of E.
41
debliecki. Two to four pigs per experimental litter or group were not inoculated with E. debliecki and served as littermate controls for clinical signs o f disease. Examination of pigs Pigs were observed daily for clinicalsigns of coccidiosis.One inoculated pig from each litterof nursing pigs or each group of weaned pigs was killed and examined at necropsy on post-inoculation days (PID) 4, 5, 8, 10, 12 and 14. Tissue samples were taken from the duodenum, every 50 c m throughout the small intestine, cecum, spiral colon, lung, liver,kidney, spleen, mesenteric lymph node and stomach. These tissues were fixed in 10% neutral buffered formalin, embedded in paraffin, sectioned at 6/~m and stained with hematoxylin and eosin for histologicexaminations. Fecal samples were taken from the cecum and fecal flotationsusing Sheather's sugar solution were examined for coccidial oocysts and ova of Strongyloides ransomi. Measurements of 20 mature macrogamonts and 20 microgamonts were made from histological sections of a nursing and weaned pig inoculated with 5 × l0 s sporulated E. debliecki oocysts and examined PID 5. Measurements were made with a calibrated ocular micrometer and are expressed in micrometers with the mean followed by the range in parentheses. Non-fixed portions of the jejunum were examined for transmissible gastroenteritis virus, rotavirus and Escherichia coli bearing pilus antigens K88, K99 or 987P (determined by immunofluorescence tests). Additional non-fixed portions of jejunum were bacteriologically cultured on blood agar and incubated anaerobically to detect Clostridium perfringens and cultured in selenite broth and brilliant green sulfa agar and incubated aerobically to detect Salmonella sp. Littermate control pigs were examined only for clinical signs of disease, none were examined at necropsy. RESULTS
Nursing pigs None of the inoculated nursing pigs developed clinicalsigns of coccidiosisor died. At necrospy, milk curds were present in the stomachs and the feces were of normal consistency. Pathogenic bacteria and viruses were not isolatedfrom any of the nursing pigs. Littermate control pigs did not develop clinicalsigns of disease. Developmental stages of E. debliecki were observed in nursing pigs of Litter A examined PID 4 and 5, of Litter B examined PID 5 and 8, and of Litter C examined PID 4 and 5. The majority of developmental stages present in nursing pigs examined PID 4 were firstand second generation meronts, whereas
42
Fig. 1. Tissue section of jejunum from a nursing piglet experimentally infected with E. debliecki. Note the two macrogamonts (MG) and their close association with the lamina propria. Hematoxylin and eosin stain. Bar: 20/~m.
those examined PID 5 had mainly second generation meronts and sexual stages. Only sexual stages and oocysts were observed in nursing pigs examined PID 8. Twenty mature macrogamonts measured 19.3 X 14.2/~m (14-24 X 12-17) and 20 microgamonts measured 11.5 X 9.1 pm (8-15 X 8-11 ). None of the nursing pigs examined PID 10, 12, or 14 had developmental stages of E. debliecki in tissue sections. Highest concentrations of parasites were present in sections of the anterior jejunum. The duodenum of three pigs (Litter A, PID 4; Litter C, PID 4 and 5) had parasites, whereas none were observed in the ileum of any pig. Developmental stages of E. debliecki were located in enterocytes lining the distal two thirds of the villi. Parasites were located in the apical cytoplasm above the host cell nucleus and infected cells often appeared to bulge into the lamina propria ( Fig. 1 ) ; however, parasites were not observed in connective tissue cells of the lamina propria. Developmental stages were not observed in the cecum, colon or in extra-intestinal tissues from any nursing pig. Mild villous atrophy of the anterior jejunum, in association with large numbers of developing sexual stages, was observed in one pig (Litter C,PID 5) given 5.0 X 106 oocysts. No lesions of coccidiosis were observed in any other nursing pigs. Fecal flotations were negative for ova ofS. ransomi. Flotations from all nursing pigs examined PID 4 and 5 contained fully sporulated oocysts of E. debliecki, whereas those examined PID 8, 10, 12, and 14 contained unsporulated oocysts of E. debliecki.
43
Weaned pigs None of the inoculated weaned pigs developed clinical signs of coccidiosis or died. At necropsy, the stomachs contained ingested feed and the feces were of normal consistency. Pathogenic bacteria or viruses were not isolated from any weaned pigs. Littermate control pigs did not develop clinical signs of disease. Developmental stages ofE. debliecki were observed in weaned pigs of Groups A, B, C, and D examined PID 4 and 5 and in pigs of Groups A and D examined PID 8. The types of developmental stages observed in weaned pigs on PID 4, 5 and 8 were identical to those found in nursing pigs on these days. Twenty mature macrogamonts measured 18.4 X 13.6 ttm (16-22 X 10-16 ) and 20 microgamonts measured 10.2X7.8 ttm (8-12X6-10). None of the weaned pigs examined PID 10, 12, or 14 had developmental stages of E. debliecki in tissue sections. Highest concentrations of parasites were in the anterior jejunum. The duodenum of five pigs ( Group A, PID 8; Group B, PID 4 and 5; Group C, PID 5 and Group D, PID 4) had parasites, whereas none were observed in the ileum of any of the weaned pigs. The distribution of parasites along the villi and location within enterocytes was identical to that observed in nursing pigs. Developmental stages were not observed in the cecum, colon or in extra-intestinal tissues from any of the weaned pigs. Mild to moderate villous atrophy in association with large numbers of developing sexual stages was observed in sections of the anterior jejunum of pigs examined PID 5 from Groups B, C and D. No lesions of coccidiosis were observed in the other weaned pigs. Fecal flotations were negative for ova of S.ransomi. Flotations from all weaned pigs examined PID 8 and 10 contained unsporulated oocysts of E. debliecki, whereas all other flotations were negative for E. debliecki. DISCUSSION Biester and Murray (1929) reported that E. debliecki (probably a mixture of many Eimeria species) caused diarrhea and emaciation in experimentally infected weaned pigs. They also reported constipation as a sequel to infection and that the wall of the cecum and colon was grossly thickened at necropsy. Material in the cecum and colon often firmly adherent to the mucosal surface. Comparisons of the results of the present study to those of Biester and Murray are difficult because they did not state the age or the previous history of the pigs that they infected. Also, they did not state the numbers of oocysts used for experimental infections. In the present study, constipation and gross lesions were not observed. Boch and Wiesenhutter (1963) reported that daily inoculations (up to 28 days) of 4.0X 105-2.0X 106 oocysts (80% E. debliecki, 15% E. scabra and 5% E. polita) into 8-14-day-old nursing pigs caused diarrhea, emaciation and
44 reduction in weights. Diarrhea began by PID 7-16 and lasted for up to 20 days in one pig that died. They did not sacrifice any pigs to examine for microscopic lesions of coccidiosis, nor were attempts made to demonstrate the presence of any other enteropathogens. The presence of E. scabra in their inoculum may have influenced the results of their study, because it has been reported to cause coccidiosis in pigs under experimental ( Rommel, 1970) and natural conditions (Hill et al., 1985 ). The results of the present study and those of Vetterling (1966b) indicate that E. debUecki is not a pathogen of nursing or weaned pigs. Vetterling infected seven 2-week-old weaned pigs with 2.0 X l0 s oocysts of E. debliecki. These pigs did not develop clinical signs of disease, had no gross lesions and only mild microscopic lesions in the small intestine. Measurements of mature macrogamonts and microgamonts of E. debliecki in the present study are in close agreement with those reported by Vetterling (1966b). The location of developmental stages and their distribution in the small intestine of these pigs were also similar to those observed by Vetterling. We did, however, find developmental stages of E. debliecki in the duodenum of several pigs, a site that was not previously reported as being parasitized. Additionally, we have demonstrated that higher dosages of oocysts of E. debliecki do not cause clinical disease in nursing or weaned pigs. Coccidia vary in their inherent ability to produce disease and it would appear that E. debliecki is a non-pathogenic species. The numbers of oocysts of E. debliecki given to nursing pigs in our study are much higher than those needed to produce clinical signs of coccidiosis and deaths in pigs infected with I. suis (Stuart et al., 1980, 1982; Lindsay et al., 1985b). The results of the present study, in which known bacterial and viral pathogens of baby pigs were found not to be present, indicate that infections by E. debliecki alone will not cause disease in baby pigs and that E. debliecki is not a cause of neonatal or weaning diarrhea in pigs. The sporulated oocysts observed in nursing pigs on PID 4 and 5 came from orally inoculated oocysts given to the pigs at 3 days of age. Some oocysts normally do not excyst while passing through the digestive tract of a host and can be found in the feces after inoculation. However, it is unusual to find them for such a long period of time as observed in t h e present study. This may be due to young pigs having slow cecal transit time. This would also explain why nursing pigs had unsporulated oocysts in their cecal contents on PID 12 and 14, while weaned pigs were negative on these days. ACKNOWLEDGEMENTS The authors thank Dr. Lauerman for performing the bacteriological and virological examinations. This study was supported in part by a grant from the College of Veterinary Medicine (FAHDR-AL-V-145) to the authors. Pub-
45
lished as College of Veterinary Medicine publication 1830 and Alabama Agricultural Experiment Station Journal No. 5-861026.
REFERENCES Biester, H.E. and Murray, C., 1929. Studies in infectious enteritis of swine IV. Intestinal coccidiosis.J. Am. Vet. Med. Assoc., 75: 705-740. Boch, J. and Wiesenhutter, E., 1963. Beitrag Zur Klarung der Pathogenitat der Schweinekokzidien. Tierartztl. Vmsch,. 18: 223-235. Coussement, W., Horrens, A.M., Ducatelle, R., Berghen, P. and Geeraerts, J., 1981. Diarrhee bij zuigende biggen geassocieerd met Eimeria neodebliecki.Diergeneeskd. Tijdschr,. 40: 384-395. Greiner, E.C., Taylor, C, Frankenberger, W.B. and Belden, R.C, 1982. Coccidia of feralswine from Florida. J. Am. Vet. Med. Assoc., 181: 1275-1277. Hill,J.E.,Lomax, L.G., Lindsay, D.S. and Lynn, B.S,. 1985. Coccidiosis caused by Eimeria scabra in a finishing hog. J. Am. Vet. Med. Assoc., 186: 981-982. Lindsay, D.S., Current, W.L, Ernst J.V. and Stuart, B.P., 1983. Diagnosis of neonatal porcine coccidiosis caused by Isospora suis.Vet. Med. Small Anita. Clin., 78: 89-95. Lindsay, D.S., Ernst, J.V, Current, W.L., Stuart, B.P. and Stewart, T.B., 1984. Prevalence of oocysts of Isospora suis and Eimeria spp. from sows on farms with and without a history of neonatal coccidiosis. J. Am. Vet. Med. Assoc., 185: 419-421. Lindsay, D.S. Blagburn, B.L., Current, W.L. and Ernst, J.V., 1985a. Development of the swine coccidium Eimeria deblieckiDouwes, 1921 in mammalian cell cultures. J. Protozool., 32: 669-671. Lindsay, D.S, Current, W.L. and Taylor, J.R., 1985b. Effects of experimental Isospora suis infection on morbidity, mortality, and weight gain of nursing pigs. Am. J. Vet. Res., 46:1511-1512. Rommel, M., 1970. Verlauf der Eimeria scabra und E. politainfektion in vollempfanglichen Ferkeln und Lauferschweinen. Berl. Muench. Tieraerztal. Wochenschr., 83: 181-186. Rommel, M. and Ipczynski, V., 1967. Der Lebenszyklus des Schweinekozids Eirneriascabra (Henry, 1931 ). Berl. Muench. Tieraerztl. Wochenschr., 80: 65-70. Stuart, B.P. and Lindsay, D.S., 1986. Coccidiosis in swine. Vet. Clin. North Am. Food. Anita. Pract., 2: 455-468. Stuart, B.P., Lindsay, D.S. and Ernst, J.V., 1978. Coccidiosis as a cause of scours in baby pigs. Proc. Int. Syrup. Neonatal Diarrhea, 2: 371-382. Stuart, B.P., Lindsay, D.S., Ernst, J.V. and Gosser, H.S., 1980. Isosporasuis enteritisin pigs.Vet. Pathol., 17: 84-93. Stuart, B.P., Gosser, H.S., Allen, C.B. and BedeU, D.M., 1982. Coccidiosis in swine: dose and age response to Isospora suis.Can. J. Comp. Med,. 46: 317-320. Swanson, L.E. and Kates, K.C, 1940. Coccidiosis in a litterof pigs. Proc. Helminthol. Soc. Wash., 7: 29-30. Vetterling, J.M., 1966a. Prevalence of coccidia in swine from six localitiesin the United States. Cornell Vet., 56: 155-166. Vetterling,J.M., 1966b. Endogenous cycle of the swine coccidium Eimeria deblieckiDouwes, 1921. J. Protozool., 13: 290-300.
39
Experimental E i m e r i a debliecki Infections in Nursing and Weaned Pigs DAVID S. LINDSAY, BYRON L. BLAGBURN and TIMOTHY R. BOOSINGER
Department of Pathology and Parasitology, College of Veterinary Medicine, Auburn University, AL 36849 (U.S.A.) (Accepted for publication 24 September 1986)
ABSTRACT
Lindsay, D.S., Blagburn, B.L. and Boosinger, T.R., 1987. Experimental Eimeria debliecki infections in nursing and weaned pigs. Vet. Parasitol., 25: 39-45. Three litters of six, 3-day-old nursing pigs were inoculated via a stomach tube with 8.0 X 105, 1.6 X 106 or 5.0 X 106 sporulated oocysts of Eimeria debliecki and four groups of six, 4-week-old weaned pigs were inoculated with 8.0 X 105, 1.6 X 106, 5.0 X 106 or 1.0 X 107 sporulated oocysts of E. debliecki to determine its pathogenicity. Clinical coccidiosis or deaths did not result from infections. Infections were confined to the jejunum and occasionally the duodenum. Microscopic lesions of mild to moderate villous atrophy were observed in one nursing pig given 5.0 X 106 oocysts and three weaned pigs given 1.6 x 106, &fiX 106 a n d 1.0 X 107 oocysts and examined 5 days post-inoculation. Pathogenic bacteria or viruses were not demonstrated in any pigs. Results of this study indicate that E. debliecki is not a cause of neonatal or weaning diarrhea in pigs.
INTRODUCTION
Neonatal coccidiosis in pigs is a common intestinal disease problem on swine farms in swine-producing areas of the world (Lindsay et al., 1983; Stuart and Lindsay, 1986). The disease is most often associated with Isospora suis infection in young pigs ( Stuart et al., 1978, 1980, 1982), however, reports of coccidiosis caused by Eimeria spp. in pigs of various ages have appeared in the literature (Biester and Murray, 1929; Swanson and Kates, 1940; Boch and Wiesenhutter, 1963; Rommel and Ipczynski, 1967; Rommel, 1970; Coussement et al., 1981, Hill et alo, 1985). In most of the reports that implicated Eimeria spp. as a cause of clinical coccidiosis, the authors did not examine infected pigs for bacterial or viral pathogens. Therefore, the role of Eimeria spp. as a cause of diarrhea or illness in pigs remains unclear. Surveys conducted in the U.S.A. indicate that 60-90% of swine living on pastures or dirt lots are infected with Eimeria spp. (Vetterling, 1966a; Greiner 0304-4017/87/$03.50
© 1987 Elsevier Science Publishers B.V.
40 et al., 1982; Lindsay et al., 1984 ). One study that involved only sows indicated that 90% were infected with Eimeria spp. and that 61% of these sows were passing oocysts ofEimeria debliecki in their feces (Lindsay et al., 1984 ). Because of the prevalence of E. debliecki in sow feces, oocysts should be present in most farrowing houses and, therefore, available to infect nursing pigs. The present study was conducted to determine if an isolate of E. debliecki (Lindsay et al., 1985a) was capable of causing disease in neonatal nursing or weaned pigs. MATERIALS AND M E T H O D S
Oocysts Oocysts of E. debliecki were sporulated, cleaned of fecal debris and enumerated for inoculation as previously reported (Lindsay et al., 1985a). Oocysts were stored in 2.5% (w/v) potassium dichromate solution at 4°C for < 60 days before use.
Pregnant sows with known breeding dates were obtained from a producer in southwest Georgia, U.S.A. These sows had undergone gestation on dirt lots and were positive for Eimeria spp. upon fecal examination. Sows were housed separately in Rockefeller isolation rooms and were confined in farrowing crates that were on elevated plastisol-coated floors. Sows were allowed ad libitum consumption of water and feed containing a coccidiostat (Ampro125%, Merck & Co., Rahway, NJ, U.S.A.). The feces of all sows were examined by fecal flotation and were negative for coccidial oocysts by 1-6 days prior to farrowing. Three litters of nursing pigs and four groups of 4-week-old weaned pigs were used for experimental E. debliecki infections. Common production practices were used in the management of nursing and weaned pigs. All 4-week-old pigs were obtained from sows that had farrowed in Rockefeller isolation rooms and were managed as sows used for nursing pig studies. The pigs were weaned at 3 weeks of age, housed in the farrowing crate and fed feed that did not contain a coccidiostat and water ad libitum.
Experimental inoculation Three litters of six, 3-day-old nursing pigs were inoculated via a stomach tube with 8.0X105 (Litter A), 1.6X106 (Litter B), or 5.0X106 (Litter C) sporulated oocysts of E. debliecki. Four groups of six, 4-week-old weaned pigs were inoculated via a stomach tube with 8.0 × l0 s ( Group A ), 1.6 × 106 ( Group B), 5.0×10 e (Group C), or 1.0X107 (Group D) sporulated oocysts of E.
41
debliecki. Two to four pigs per experimental litter or group were not inoculated with E. debliecki and served as littermate controls for clinical signs o f disease. Examination of pigs Pigs were observed daily for clinicalsigns of coccidiosis.One inoculated pig from each litterof nursing pigs or each group of weaned pigs was killed and examined at necropsy on post-inoculation days (PID) 4, 5, 8, 10, 12 and 14. Tissue samples were taken from the duodenum, every 50 c m throughout the small intestine, cecum, spiral colon, lung, liver,kidney, spleen, mesenteric lymph node and stomach. These tissues were fixed in 10% neutral buffered formalin, embedded in paraffin, sectioned at 6/~m and stained with hematoxylin and eosin for histologicexaminations. Fecal samples were taken from the cecum and fecal flotationsusing Sheather's sugar solution were examined for coccidial oocysts and ova of Strongyloides ransomi. Measurements of 20 mature macrogamonts and 20 microgamonts were made from histological sections of a nursing and weaned pig inoculated with 5 × l0 s sporulated E. debliecki oocysts and examined PID 5. Measurements were made with a calibrated ocular micrometer and are expressed in micrometers with the mean followed by the range in parentheses. Non-fixed portions of the jejunum were examined for transmissible gastroenteritis virus, rotavirus and Escherichia coli bearing pilus antigens K88, K99 or 987P (determined by immunofluorescence tests). Additional non-fixed portions of jejunum were bacteriologically cultured on blood agar and incubated anaerobically to detect Clostridium perfringens and cultured in selenite broth and brilliant green sulfa agar and incubated aerobically to detect Salmonella sp. Littermate control pigs were examined only for clinical signs of disease, none were examined at necropsy. RESULTS
Nursing pigs None of the inoculated nursing pigs developed clinicalsigns of coccidiosisor died. At necrospy, milk curds were present in the stomachs and the feces were of normal consistency. Pathogenic bacteria and viruses were not isolatedfrom any of the nursing pigs. Littermate control pigs did not develop clinicalsigns of disease. Developmental stages of E. debliecki were observed in nursing pigs of Litter A examined PID 4 and 5, of Litter B examined PID 5 and 8, and of Litter C examined PID 4 and 5. The majority of developmental stages present in nursing pigs examined PID 4 were firstand second generation meronts, whereas
42
Fig. 1. Tissue section of jejunum from a nursing piglet experimentally infected with E. debliecki. Note the two macrogamonts (MG) and their close association with the lamina propria. Hematoxylin and eosin stain. Bar: 20/~m.
those examined PID 5 had mainly second generation meronts and sexual stages. Only sexual stages and oocysts were observed in nursing pigs examined PID 8. Twenty mature macrogamonts measured 19.3 X 14.2/~m (14-24 X 12-17) and 20 microgamonts measured 11.5 X 9.1 pm (8-15 X 8-11 ). None of the nursing pigs examined PID 10, 12, or 14 had developmental stages of E. debliecki in tissue sections. Highest concentrations of parasites were present in sections of the anterior jejunum. The duodenum of three pigs (Litter A, PID 4; Litter C, PID 4 and 5) had parasites, whereas none were observed in the ileum of any pig. Developmental stages of E. debliecki were located in enterocytes lining the distal two thirds of the villi. Parasites were located in the apical cytoplasm above the host cell nucleus and infected cells often appeared to bulge into the lamina propria ( Fig. 1 ) ; however, parasites were not observed in connective tissue cells of the lamina propria. Developmental stages were not observed in the cecum, colon or in extra-intestinal tissues from any nursing pig. Mild villous atrophy of the anterior jejunum, in association with large numbers of developing sexual stages, was observed in one pig (Litter C,PID 5) given 5.0 X 106 oocysts. No lesions of coccidiosis were observed in any other nursing pigs. Fecal flotations were negative for ova ofS. ransomi. Flotations from all nursing pigs examined PID 4 and 5 contained fully sporulated oocysts of E. debliecki, whereas those examined PID 8, 10, 12, and 14 contained unsporulated oocysts of E. debliecki.
43
Weaned pigs None of the inoculated weaned pigs developed clinical signs of coccidiosis or died. At necropsy, the stomachs contained ingested feed and the feces were of normal consistency. Pathogenic bacteria or viruses were not isolated from any weaned pigs. Littermate control pigs did not develop clinical signs of disease. Developmental stages ofE. debliecki were observed in weaned pigs of Groups A, B, C, and D examined PID 4 and 5 and in pigs of Groups A and D examined PID 8. The types of developmental stages observed in weaned pigs on PID 4, 5 and 8 were identical to those found in nursing pigs on these days. Twenty mature macrogamonts measured 18.4 X 13.6 ttm (16-22 X 10-16 ) and 20 microgamonts measured 10.2X7.8 ttm (8-12X6-10). None of the weaned pigs examined PID 10, 12, or 14 had developmental stages of E. debliecki in tissue sections. Highest concentrations of parasites were in the anterior jejunum. The duodenum of five pigs ( Group A, PID 8; Group B, PID 4 and 5; Group C, PID 5 and Group D, PID 4) had parasites, whereas none were observed in the ileum of any of the weaned pigs. The distribution of parasites along the villi and location within enterocytes was identical to that observed in nursing pigs. Developmental stages were not observed in the cecum, colon or in extra-intestinal tissues from any of the weaned pigs. Mild to moderate villous atrophy in association with large numbers of developing sexual stages was observed in sections of the anterior jejunum of pigs examined PID 5 from Groups B, C and D. No lesions of coccidiosis were observed in the other weaned pigs. Fecal flotations were negative for ova of S.ransomi. Flotations from all weaned pigs examined PID 8 and 10 contained unsporulated oocysts of E. debliecki, whereas all other flotations were negative for E. debliecki. DISCUSSION Biester and Murray (1929) reported that E. debliecki (probably a mixture of many Eimeria species) caused diarrhea and emaciation in experimentally infected weaned pigs. They also reported constipation as a sequel to infection and that the wall of the cecum and colon was grossly thickened at necropsy. Material in the cecum and colon often firmly adherent to the mucosal surface. Comparisons of the results of the present study to those of Biester and Murray are difficult because they did not state the age or the previous history of the pigs that they infected. Also, they did not state the numbers of oocysts used for experimental infections. In the present study, constipation and gross lesions were not observed. Boch and Wiesenhutter (1963) reported that daily inoculations (up to 28 days) of 4.0X 105-2.0X 106 oocysts (80% E. debliecki, 15% E. scabra and 5% E. polita) into 8-14-day-old nursing pigs caused diarrhea, emaciation and
44 reduction in weights. Diarrhea began by PID 7-16 and lasted for up to 20 days in one pig that died. They did not sacrifice any pigs to examine for microscopic lesions of coccidiosis, nor were attempts made to demonstrate the presence of any other enteropathogens. The presence of E. scabra in their inoculum may have influenced the results of their study, because it has been reported to cause coccidiosis in pigs under experimental ( Rommel, 1970) and natural conditions (Hill et al., 1985 ). The results of the present study and those of Vetterling (1966b) indicate that E. debUecki is not a pathogen of nursing or weaned pigs. Vetterling infected seven 2-week-old weaned pigs with 2.0 X l0 s oocysts of E. debliecki. These pigs did not develop clinical signs of disease, had no gross lesions and only mild microscopic lesions in the small intestine. Measurements of mature macrogamonts and microgamonts of E. debliecki in the present study are in close agreement with those reported by Vetterling (1966b). The location of developmental stages and their distribution in the small intestine of these pigs were also similar to those observed by Vetterling. We did, however, find developmental stages of E. debliecki in the duodenum of several pigs, a site that was not previously reported as being parasitized. Additionally, we have demonstrated that higher dosages of oocysts of E. debliecki do not cause clinical disease in nursing or weaned pigs. Coccidia vary in their inherent ability to produce disease and it would appear that E. debliecki is a non-pathogenic species. The numbers of oocysts of E. debliecki given to nursing pigs in our study are much higher than those needed to produce clinical signs of coccidiosis and deaths in pigs infected with I. suis (Stuart et al., 1980, 1982; Lindsay et al., 1985b). The results of the present study, in which known bacterial and viral pathogens of baby pigs were found not to be present, indicate that infections by E. debliecki alone will not cause disease in baby pigs and that E. debliecki is not a cause of neonatal or weaning diarrhea in pigs. The sporulated oocysts observed in nursing pigs on PID 4 and 5 came from orally inoculated oocysts given to the pigs at 3 days of age. Some oocysts normally do not excyst while passing through the digestive tract of a host and can be found in the feces after inoculation. However, it is unusual to find them for such a long period of time as observed in t h e present study. This may be due to young pigs having slow cecal transit time. This would also explain why nursing pigs had unsporulated oocysts in their cecal contents on PID 12 and 14, while weaned pigs were negative on these days. ACKNOWLEDGEMENTS The authors thank Dr. Lauerman for performing the bacteriological and virological examinations. This study was supported in part by a grant from the College of Veterinary Medicine (FAHDR-AL-V-145) to the authors. Pub-
45
lished as College of Veterinary Medicine publication 1830 and Alabama Agricultural Experiment Station Journal No. 5-861026.
REFERENCES Biester, H.E. and Murray, C., 1929. Studies in infectious enteritis of swine IV. Intestinal coccidiosis.J. Am. Vet. Med. Assoc., 75: 705-740. Boch, J. and Wiesenhutter, E., 1963. Beitrag Zur Klarung der Pathogenitat der Schweinekokzidien. Tierartztl. Vmsch,. 18: 223-235. Coussement, W., Horrens, A.M., Ducatelle, R., Berghen, P. and Geeraerts, J., 1981. Diarrhee bij zuigende biggen geassocieerd met Eimeria neodebliecki.Diergeneeskd. Tijdschr,. 40: 384-395. Greiner, E.C., Taylor, C, Frankenberger, W.B. and Belden, R.C, 1982. Coccidia of feralswine from Florida. J. Am. Vet. Med. Assoc., 181: 1275-1277. Hill,J.E.,Lomax, L.G., Lindsay, D.S. and Lynn, B.S,. 1985. Coccidiosis caused by Eimeria scabra in a finishing hog. J. Am. Vet. Med. Assoc., 186: 981-982. Lindsay, D.S., Current, W.L, Ernst J.V. and Stuart, B.P., 1983. Diagnosis of neonatal porcine coccidiosis caused by Isospora suis.Vet. Med. Small Anita. Clin., 78: 89-95. Lindsay, D.S., Ernst, J.V, Current, W.L., Stuart, B.P. and Stewart, T.B., 1984. Prevalence of oocysts of Isospora suis and Eimeria spp. from sows on farms with and without a history of neonatal coccidiosis. J. Am. Vet. Med. Assoc., 185: 419-421. Lindsay, D.S. Blagburn, B.L., Current, W.L. and Ernst, J.V., 1985a. Development of the swine coccidium Eimeria deblieckiDouwes, 1921 in mammalian cell cultures. J. Protozool., 32: 669-671. Lindsay, D.S, Current, W.L. and Taylor, J.R., 1985b. Effects of experimental Isospora suis infection on morbidity, mortality, and weight gain of nursing pigs. Am. J. Vet. Res., 46:1511-1512. Rommel, M., 1970. Verlauf der Eimeria scabra und E. politainfektion in vollempfanglichen Ferkeln und Lauferschweinen. Berl. Muench. Tieraerztal. Wochenschr., 83: 181-186. Rommel, M. and Ipczynski, V., 1967. Der Lebenszyklus des Schweinekozids Eirneriascabra (Henry, 1931 ). Berl. Muench. Tieraerztl. Wochenschr., 80: 65-70. Stuart, B.P. and Lindsay, D.S., 1986. Coccidiosis in swine. Vet. Clin. North Am. Food. Anita. Pract., 2: 455-468. Stuart, B.P., Lindsay, D.S. and Ernst, J.V., 1978. Coccidiosis as a cause of scours in baby pigs. Proc. Int. Syrup. Neonatal Diarrhea, 2: 371-382. Stuart, B.P., Lindsay, D.S., Ernst, J.V. and Gosser, H.S., 1980. Isosporasuis enteritisin pigs.Vet. Pathol., 17: 84-93. Stuart, B.P., Gosser, H.S., Allen, C.B. and BedeU, D.M., 1982. Coccidiosis in swine: dose and age response to Isospora suis.Can. J. Comp. Med,. 46: 317-320. Swanson, L.E. and Kates, K.C, 1940. Coccidiosis in a litterof pigs. Proc. Helminthol. Soc. Wash., 7: 29-30. Vetterling, J.M., 1966a. Prevalence of coccidia in swine from six localitiesin the United States. Cornell Vet., 56: 155-166. Vetterling,J.M., 1966b. Endogenous cycle of the swine coccidium Eimeria deblieckiDouwes, 1921. J. Protozool., 13: 290-300.