Progress 10/01/99 to 09/30/04
Outputs
Major accomplishments of this project included: An experimental infection model of Haemonchus contortus infection in sheep was developed that has been used to identify parasite resistant and parasite susceptible animals and to investigate breed related resistance to parasitism. Katahdin sheep appear to have increased resistance to gastrointestinal helminths because parasite resistant hair sheep were included in their breeding. Katahdin lambs were evaluated for their resistance to gastrointestinal nematodiasis, but did not seem to demonstrate increased levels of resistance. We have tested several alternative methods of nematode control in sheep and goats including diatomaceous earth, copper boluses, quebracho tannins and herbal dewormers. Copper boluses appear to have some value for parasite control and will be investigated further. Diatomaceous earth, quebracho tannin and the herbal dewormers did not seem to have any effect on numbers of nematode eggs in feces. In vitro
studies were conducted to investigate the effects of ponazuril on the important Apicomplexan parasite Neospora and Toxoplasma, that cause abortion in small ruminants and cattle, respectively. This drug was found to be highly effective against the organisms.
Impacts
Evaluation of proposed alternatives to commercial drug use for treatment of gastrointestinal worms in sheep and goats is of critical importance to producers in much of the eastern and central U.S. because of the dramatic increase in drug resistance in the parasites. Although laboratory research or research conducted in other areas may suggest possible alternatives, these must be tested locally to determine their utility under specific climatic and management conditions. Work carried out in Virginia and collaborating states will have a direct impact on the ability of producers to raise small ruminants profitably.
Publications
- Vanimisetti, H. B., S. P. Greiner, A. M. Zajac, and D. R. Notter. 2002. Between-breed variation in response to Haemonchus contortus infection in sheep. J. Anim. Sci. 80(Suppl):1.
- Mitchell S.M., Zajac A.M., Davis W.L., Lindsay D.S. 2003. Mode of action of ponazuril against Toxoplasma gondii tachyzoites. J. Eukaryotic Micro. 50(Suppl):689-90.
- Vannimisetti H.B., Greiner, S.P, Zajac A.M., Notter D.R. 2004. Performance of hair sheep composite breeds: Resistance of lambs to Haemonchus contortus. J. Anim. Sci. 82:595-604.
- Vanimisetti H.B., Andrew S.L., Zajac A.M., Notter D.R. 2004. Inheritance of fecal egg count and packed cell volume and their relationship with production traits in sheep infected with Haemonchus contortus. J. Anim. Sci. 82:1602-1611.
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Progress 10/01/02 to 09/30/03
Outputs
In the previous year, a study was done investigating the effect of supplemental feeding of the plant tannin, quebracho, on an experimental infection of Haemonchus contortus in goats. This year, a second study was completed in which quebracho tannin was provided as a supplement to grazing goats exposed to natural infection with H. contortus. In neither situation was any beneficial effect of quebracho observed. The effect of the aniprotozoal agent, ponazuril, was also tested against Toxoplasma gondii in vitro. Ponazuril caused a 94 percent reduction in tachyzoite multiplication at 5.0 micrograms/ml and 99 percent at 1.0 microgram/ml Under the conditions of the study, ponazuril was coccidiostatic. Ultrastructural observations of treated parasites indicate that ponazuril interferes with parasite cytokenisis.
Impacts
Evaluation of proposed alternatives to commercial drug use for treatment of gastrointestinal worms in sheep and goats is of critical importance to producers in much of the eastern and central U.S. because of the dramatic increase in drug resistance in the parasites. Although laboratory research or research conducted in other areas may suggest possible alternatives, these must be tested locally to determine their utility under specific climatic and management conditions. Work carried out in Virginia and collaborating states will have a direct impact on the ability of producers to raise small ruminants profitably.
Publications
- Notter, D.R., Andrew, S.A., Zajac, A.M. 2003. Responses of hair sheep, wool sheep and hair X wool crossbreds to a single fixed dose of infective larvae of Haemonchus contortus. Journal of Small Ruminant Research. 47:221-225.
- Lindsay, D.S., Neifer, R., Hildreth, M. 2002. Porcine enteritis associated with Eimeria spinosa Henry, 1931 infection. Journal of Parasitology 88:1262-1263
- Mitchell, S., Zajac, A., Davis, W., Lindsay D. 2003. Activity of ponasuril against Toxoplasma gondii in African Green Monkey kidney cells. Annual meeting of the Southeastern Society of Parasitologists.
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Progress 10/01/01 to 09/30/02
Outputs
We examined the effect of feeding plant tannin on trichostrongylid parasites of goats. This is part of a W-102, a multistate project that examines alternative methods of trichostrongylid parasite control in small ruminants where parasite resistance has become an important limiting factor to successful production of sheep and goats. In an experiment conducted in Virginia in collaboration with Virginia State Univerisity and the USDA-ARS research station in Beaver, WV 32 young male and female goats were divided into 2 groups. Each group was housed in confinement and fed either a protein supplement or protein supplement containing quebracho plant extract. Fecal egg counts and blood packed cell volumes were followed for 8 weeks. No difference was seen between treated and untreated animals in the parasitologic parameters measured nor were any differences in weight gains detected, although additional studies are planned to examine the effects of feeding of higher levels of
the quebracho extract.
Impacts
This research will lead to the development of alternative means of parasite control in sheep and goats in the U.S. Current deworming drugs available often have little impact because of high levels of drug resistance. Use of plants containing high levels of tannins could be a valuable means of control if appropriate levels and methods of application are determined for U.S. conditions.
Publications
- Zajac, A., Wildeus, S., and Turner, K. 2002. Effect of Quebracho Extract on Experimental Haemonchus contortus Infection in Goats. Presentatation at the annual meeting of the American Association of Veterinary Parasitologists, Nashville TN, July. abstract
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Progress 10/01/00 to 09/30/01
Outputs
1 - Control of Parasitic Diseases - Equine protozoal myeloencephalitis (EPM) is the most important protozoal disease of horses in North America. Our study examined the efficacy of pyrantel tartrate in inhibiting merozoite production of Sarcocystis neurona in cell cultures and in preventing clinical disease in interferon-0 (IFN-0) gene knockout (BALB/c-Ifngtm1ts) mice. Pyrantel tartrate had little activity in cell culture. A dose of 25 ug/ml caused a 34 percent reduction in merozoite production. No activity was seen against sporocyst-induced S. neurona infections in mice treated prophylacticly with 4 mg pyrantel tartrate per mouse per day in the drinking water. We have developed a new test, Sarcocystis neurona agglutination test (SAT), to detect S. neurona antibodies in horses and other animals. We have used our SAT to examine the prevalence of antibodies to S. neurona in wildlife and determined that the prevalence of antibodies in raccoons is very similar to that of
horses. Our new test may aid in understanding the epidemiology of EPM and in identifying new intermediate hosts. 2 - Define Roles in Parasite Control - We have examined the roles of inducible and endothelial nitric oxide synthase in resistance to sporocyst induced S. neurona infections in genetically defined mice. We found that mice lacking the gene for inducible nitric oxide synthase (iNOS) and mice lacking the gene for endothelial nitric oxide synthase (eNOS) were resistant to clinical infection and had no lesions in their brains at necropsy. In contrast, clinical disease and deaths occurred in mice with no gene for interferon-0 (IFN-0). Results of this study indicate that iNOS and eNOS are not major mediators of resistance to S. neurona infections. Results of this study suggests that IFN-0 mediated immunity to S. neurona may be mediated by non-nitric oxide dependent mechanisms. Neospora hughesi is a recently described apicomplexan parasite that has been associated with several
cases of equine protozoal myeloencephalitis. Genes of the two dense granule proteins of N. hughesi (isolate Nh-A1) and four different isolates of N. caninum were isolated via PCR and their DNA sequences were determined. Results of our study indicate that diagnostic tests based on differences in dense granule sequences and antigenicity may have potential to differentiate between N. hughesi and N. caninum. Such diagnostic tests would be valuable tools to aid in our understanding of the epidemiology of these parasites. Additionally, dense granule proteins are immunogenic and they may have potential as use in recombinant vaccines against neosporosis. We have expressed N. caninum dense granule 1 (NcGRA7) and surface reactive sequence 2 (SRS2) in the RB51 vaccine strain of Brucella abortus. We are currently evaluating these candidate vaccines in a gerbil model of neosporosis. We hope to vaccinate cattle against both abortion-causing agents using this approach.
Impacts
Results of experiments on breed variation in resistance to gastrointestinal parasites of sheep will help producers breed for resistance rather than relying solely on chemical means of control. Development of a new diagnostic test for the agent causing EPM in horses will permit better understanding of the distribution and transmission of this important equine parasite.
Publications
- Lindsay, D.S., A.C. Rosypal, J.A. Spencer, M.A. Cheadle, A.M. Zajac, C. Rupprecht, J.P. Dubey, and B.L. Blagburn. 2001. Prevalence of agglutinating antibodies to Sarcocystis neurona in raccoons, Procyon lotor, from the United States. Vet. Parasitol. (in press).
- Walsh, C.P., R. Vemulapalli, N. Sriranganathan, A.M. Zajac, M.C. Jenkins, and D. S. Lindsay. 2001. Molecular comparison of the dense granule proteins GRA6 and GRA7 of Neospora hughesi and Neospora caninum Int. J. Parasitol. 31:253-258.
- Dubey, J.P., D. Fritz, D.S. Lindsay, S.K. Shen, O.C.H. Kwok, and K.C. Thompson. 2001. Diclazuril preventive therapy of gamma interferon knockout mice fed Sarcocystis neurona sporocysts. Vet. Parasitol. 94:257-263.
- Dubey, J.P., D.E. Mattson, C.A. Speer, A. N. Hamir, D.S. Lindsay, B.M. Rosenthal, O.C.H. Kwock, R.J. Baker, D.M. Mulroony, S.J. Tornquist, and T. C. Gerros. 2001. Characteristics of a recent isolate of Sarcocystis neurona (SN7) from a horse and loss of pathogenicity of the SN7 and SN6 stains after passage in cell culture. Vet. Parasitol. 95:155-166.
- Dubey, J.P., D.S. Lindsay, C. E. Kerber, N. Kasai, H. F. T. Pena, O.C.H. Kwok, S.K. Shen, and B.M. Rosenthal. 2001. First isolation of Sarcocystis neurona from the South American opossum, Didelphis albiventris, from Brazil . Vet. Parasitol. 95:295-304.
- Dubey, J.P., D.S. Lindsay, O.C.H. Kwok and S.K. Shen. 2001. The gamma interferon knockout mouse model of Sarcocystis neurona: Comparison of infectivity of merozoites and sporocysts and routes of inoculation. J. Parasitol. 2001
- Lindsay, D.S. and J. P. Dubey. 2001. Direct agglutination test for the detection of antibodies to Sarcocystis neurona in experimentally infected animals. Vet. Parasitol.95:179-186.
- Lindsay, D.S., and J. P. Dubey. 2001. Determination of the activity of pyrantel tartrate against Sarcocystis neurona. Vet. Parasitol. 97:141-144.
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Progress 10/01/99 to 09/30/00
Outputs
The efficacy of a DNA vaccine to Neospora caninum dense granule GRA7 was investigated and found to partially protect mice against encephalitis. Dogs were infected with Neospora caninum and oocyst production and serological responses monitored. Oocysts were shed for a short time by puppies. Antibody was not detected during the period of oocyst shedding, but parasite specific antibody was detected subsequently. Biological and genetic differences between Neospora hughesi and Neospora caninum were identified and supported separation of the organisms into distinct species. In an effort to establish an equine model of Sarcocystis neurona infection, parasites were inoculated into the subarachnoid space of horses. None of the horses developed clinical disease, although all animals developed serum and CSF antibody to the S. neurona. In other work with Sarcocystis neurona, gamma interferon knockout mice were shown to be susceptible to infection with the parasite. The effects of
diatomaceous earth (DE) on developing strongylid larvae were also examined in an in vitro system to determine if this product would help control important parasitic infections in ruminants. Trichostrongylid eggs (primarily Haemonchus contortus) were isolated from sheep feces and added to 96 well microtitre plates with nutritive medium. Varying concentrations of DE were added to wells and survival and development to the third larval stage were monitored. Reduction in the number of L3 developing from eggs was apparent only at very high levels of DE. Preliminary experiments to develop an effective vaccine for the infection were also completed.
Impacts
Results of research conducted in the last year improve the understanding of the parasite Neospora caninum, a leading cause of bovine abortion in the U.S. These results provided information on the role of dogs as a final host for the parasite. This knowledge is critical in designing effective control programs to prevent infection of cattle.
Publications
- Zajac A M., Gipson TA. 2000. Multiple anthelmintic resistance in a goat herd. Vet. Parasitol. 87:163-172.
- Lindsay DS, Upton SJ, Owens DS, Morgan UM, Mead JR, Blagburn BL. 2000. Cryptosporidium andersoni n. sp. (Apicomplexa: Cryptosporiidae) from Cattle, Bos taurus. J. Euk. Microbiol. 47: 91-95.
- Dubey JP, and Lindsay DS. 2000. High susceptibility of gerbils to oral infection with Neospora caninum oocysts. Parasitol. Res 86:165-168.
- Lindsay DS, and Dubey JP. 2000. Determination of the activity of diclazuril against Sarcocystis neurona and Sarcocystis falcatula in cell cultures. J. Parasitol. 86:164-166.
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Progress 10/01/98 to 09/30/99
Outputs
The extent of the susceptibility of Katahdin sheep to nematode parasitism was examined. Fecal egg counts, Packed Cell Volumes and body weight were monitored in 17 Katahdin lambs and 19 Spanish goat kids during the grazing season. Spanish goats were used as a sentinel groups because previous work showed that Spanish kids were susceptible to Haemonchus contortus infection and developed clinical disease. Lambs and kids were severely affected by parasitism and coccidiosis after weaning and 2 lambs and 4 kids died. Following treatment, levels of parasitism in both groups of animals remained low, probably because of dry conditions. In spring 1999, levels of infection in a parasite resistant sheep breed, the Barbados Blackbelly, were compared with levels in Katahdin sheep. Five lambs of each breed were followed for 4 months and euthanized for determination of total worm burdens. Fecal egg counts of Katahdin lambs were significantly higher than those of Blackbelly lambs
before euthanasia. A study establishing parameters for nonpathogenic experimental infection of sheep with Haemonchus contortus was concluded. The protocol allows ewes and lambs to be tested for susceptibility to the parasite without loss of growth or production. Adult ewes are dewormed and 3 days later receive 20,000 third stage H. contortus larvae. Lambs are similarly treated but receive 10,000 third stage larvae. Ewes are monitored by fecal egg count and PCV every other week from weeks 3 to 13 after infection. Lambs are monitored weekly from week 3 to 8 after infection. After 2 years of testing during spring and fall lambing, no sheep developed clinical haemonchosis and differences have been great enough to allow evaluation of variation in susceptibility. For the fourth year, levels of parasitism in steers in a low input grazing management system were compared to animals in a traditional grazing system. In the conventional system, 24 steers are winter, spring and summer grazed on
fescue with a late summer move to fescue-red clover. In the low input sustainable system, 24 steers graze tall fescue-alfalfa in autumn and winter. The remainder of the system is divided into crop paddocks used for grazing and/or harvesting. No significant effects of management system on parasite levels have been found. Studies were conducted on the transmission of the flagellate parasite Cochlosoma anatis in turkeys and the role of Cochlosoma in turkey enteritis. Oral inoculation of turkeys with trophozoites consistently reproduced infections in recipient turkeys. Trophozoites were most numerous in the jejunum and ileum but could be observed in the duodenum, ceca, colon, and feces, of turkeys. When naive turkeys were placed on contaminated litter vacated by excreting turkeys only 1 of 12 turkeys became infected. When naive turkeys were placed in boxes with birds currently excreting trophozoites 7 of 8 turkeys became infected. Trophozoites did not survive exposure to water or
freezing. Bobwhite quail and chickens could also be infected and serve as a source of infection for turkeys.
Impacts
Experiments on experimental Haemonchus infection and differences in breed resistance in small ruminants can be used in formulating recommendations to producers on ways to use selection for parasite resistance in breeding programs. Results of studies on the protozoan Cochlosoma are also of use to producers in understanding the role of the parasite in the important turkey poult enteritis complex.
Publications
- Lindsay D., Larsen C., Zajac A., Pierson F. 1999. Experimental Cochlosoma anatis infections in poultry. Veterinary Parasitology. 81:21-27.
- Zajac, A., Gipson, T. 1998 .Variation in Resistance to Trichostrongyle Parasites in Goats. 43rd Annual Meeting, American Association of Veterinary Parasitologists, Baltimore, Maryland
- Lindsay, D., Larsen, C.T., Zajac, A., Pierson, F.W. 1998. Experimental Cochlosoma anatis infections in poultry. 43rd Annual Meeting, American Association of Veterinary Parasitologists, Baltimore, Maryland.
- Zajac, A., Wildeus, S. 1999. Gastrointestinal parasitism of Katahdin lambs and Spanish kids in eastern Virgninia. Annual Meeting of the Southern Association of Agricultural Scientists, Memphis, Tennessee.
- Notter, D., Andrew, S., Zajac, A. 1999. Response of Wool, Hair and Wool X Hair Crossbred Lambs to a Fixed Dose of Infective Larvae of H. contortus. Annual Meeting of the Southern Association of Agricultural Scientists, Memphis, Tennessee.
- Lindsay, D.S., C. P. Walsh, and A. M. Zajac. 1999. Potential for transmission of Toxoplasma gondii in goats milk. 6th International Workshops on Opportunistic Protists and Society of Protozoologists. (Raleigh, NC. May).
- Walsh, C.P., A.M. Zajac, C.T. Larsen, and D.S. Lindsay. 1999. Cochlosoma anatis: Ultrastructure and attachment. 71st Northeastern Conference on Avian Diseases. (Blacksburg, VA, June 18)
- Lindsay, D.S., C.P. Walsh, A.M. Zajac, C.T. Larsen, and F.W. Pierson. 1999. Cochlosoma anatis: Biology and host specificity. 71st Northeastern Conference on Avian Diseases. (Blacksburg, VA, June 18)
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Progress 10/01/97 to 09/30/98
Outputs
Variation in breed resistance of goats to GI trichostrongylids was examined in collaboration with Virginia State University. Monitoring of fecal egg counts and Packed Cell Volumes of both adult periparturient does and market weight kids indicated that Myotonic goats show greater resistance to Haemonchus contortus infection than Spanish and Nubian breeds Larval culture of composite fecal samples showed that H. contortus is was the most common trichostrongyle species throughout the year. Market weight kids (4-7 of each breed) were euthanized and worm burdens in the abomasum and small intestine evaluated. Pygmy and Myotonic goats had significantly lower worm burdens than Nubian and Spanish goats. The most common parasite genera in goats of all breeds was Trichostrongylus (samples collected in May). H. contortus was also common. Variation in breed resistance was less apparent in periparturient does, which may indicate that breed differences in younger animals are
associated with the rate of development of immunity. Genetic resistance to parasites in sheep is also being examined. An experimental infection model of H. contortus using the Beltsville parasite strain has been developed. Adult ewes are dewormed and 3 days later receive 20,000 3rd stage H. contortus larvae. Lambs are similarly treated but receive 10,000 larvae. Ewes and lambs are monitored by fecal egg count and PCV. A further study is being conducted at Virginia State University comparing responses to parasites in Katahdin sheep (believed to be parasite resistant) pastured with goats. Because levels of drug resistance are increasing in the US it is important to appreciate whether susceptibility to drugs reappears in field situations. In 1995 and 1996 goats in the VSU herd had parasites resistant to benzimidazoles, macrolides and levamisole. In 1997 it appeared that levamisole had regained its efficacy after use was discontinued in 1995. This spring, however, resistance to levamisole
had reappeared after using the drug again in 1997. Human toxoplasmosis has been acquired by the consumption of unpasteurized goats milk. A preliminary study was conducted to determine how long T. gondii tachyzoites could survive in goat milk. Unpasteurized and pasteurized goat milk were spiked with tachyzoites and incubated at 37 or 4 C for various times. T. gondii. tachyzoites did not survive for over 6 hours when incubated in goat milk at 37 C. Tachyzoites incubated at 4 C survived for 4-7 days. These results indicate that refrigerated unpasteurized goat milk could be a potential source of T. gondii infection. Studies are also being conducted on the transmission of the flagellate parasite Cochlosoma anatis in turkeys and the role of this organism in causing turkey enteritis. Results suggest that the organism is directly transmitted between birds and that young birds are more likely to develop clinical disease.
Impacts
(N/A)
Publications
- No publications reported this period
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Progress 10/01/96 to 09/30/97
Outputs
To compensate for the effects of annual climatic variation and detect effects which may be cumulative over several years, infection with trichostrongylid nematode parasites in untreated cattle was followed for a second grazing season to compare the impact of a low-input sustainable management program with a conventional grazing program. No deworming treatments were given to the cattle in either group at spring turnout. Monitoring of fecal egg counts was begun in December, 1996 and continued through September 1997. Blood samples were also collected monthly for determination of serum pepsinogen levels. In the 2 previous years an early peak in fecal egg counts was seen in late winter. In the current year this increase was less pronounced. As in the previous years, animals in the low-input group had lower fecal egg counts at the start of the grazing season. However, clinical disease requiring treatment developed in both groups of steers in June. Levels of parasitism
following treatment remained low through the remainder of the grazing season. Results of monitoring for a third consecutive grazing season continue to indicate that the low-input system has had no annual or cumulative beneficial effect on levels of gastrointestinal nematodiasis.
Impacts
(N/A)
Publications
- No publications reported this period
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Progress 10/01/95 to 09/30/96
Outputs
Infection with trichostrongylid nematode parasites in untreated cattle was followed for a second grazing season to compare the impact of a low-input sustainable management program with a conventional grazing program. No deworming treatments were given to the cattle in either group at spring turnout. Monitoring of fecal egg counts was begun in the winter and continued for 11 months. As in the previous year, animals in the low-input group had lower fecal egg counts at the start of the grazing season. However, egg counts in the two groups of cattle reached similar levels in the summer and did not differ markedly for the remainder of the period. Weight gains were also similar between the 2 groups. Results from the previous and current grazing seasons do not indicate that the low-input system has an economically significant impact on levels of gastrointestinal nematodiasis.
Impacts
(N/A)
Publications
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Progress 01/01/94 to 09/30/95
Outputs
Infection with trichostrongylid nematode parasites was followed in two groups ofcattle (4 replicates of each group). Fecal parasite egg counts were determined in 24 commercial stocker cattle raised conventionally on corn and fescue pasture and were compared with those of 24 cattle raised in a low input sustainable agriculture (LISA) program involving extensive pasture and crop rotations. No anthelmintic treatment was undertaken at the beginning of the grazing season to determine whether differences in the grazing system alone would be adequate for parasite control. However, both groups of animals developed evidence of clinical parasitism over the course of the grazing season and required anthelmintic treatment.
Impacts
(N/A)
Publications
- NO PUBLICATIONS REPORTED THIS PERIOD.
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