1. Conclusions  Better parasitology screening, records, and protocols need to be developed for shelters.  Results were similar to 2003 ISU Maddie’s Summer Scholars results for common gastrointestinal parasites.  Primers and probes were not specific to C. parvum and C. felis (18S ribosomal sequence that primers and probes were designed from is conserved).  Inadvertent contamination could have caused all samples to appear positive for Cryptosporidium. References 1.Anderson, Bruce, PhD. “AVS 471” University of Idaho College of Agriculture and Life Sciences (2005). 2.Bowman et al. "Gastro-intestinal Parasites of Cats." International Veterinary Information Service (2003): 1-33. 2. Berkley, Rachel et al. “The incidence of parasitism in Midwest shelter owned companion animals.” Iowa State University Summer Scholars (2003). 3. Burrows, Colin F., BVetMed, PhD, MRCVS. "Cryptosporidium." Perspectives (1994): 53 – 54. 4. Gookin, J. L. et al. “Single-tube nested PCR for detection of Tritrichomonas foetus in feline feces.” Journal of Clinical Microbiology 40 (2002): 4126 – 4130. 5. Lindsay, David S., PhD and Anne M. Zajac, DVM, PhD. "Cryptosporidium Infections in Cats and Dogs." Compendium (2004): 864 – 874. 6. Mtambo, MMA et al. "Cryptosporidium infection in cats: Prevalence of infection in domestic and feral cats in Glasgow area." Vet Rec 129 (1991): 502 – 504. 7. Scorza, A. V. et al. "Polymerase Chain Reaction for the Detection of Cryptosporidium Spp. In Cat Feces." Journal of Parasitology 89 (2003): 423 – 426. 8. Wu, Z. et al. "Specific PCR primers for Cryptosporidium parvum with extra high sensitivity." Molecular and Cellular Probes 14 (2000): 33 – 39. Acknowledgements Special thanks to: Amanda Ramer, Dr. Michael Wannemuehler, Dr. Albert Jergens, Andrea Dorn, Iowa State University CVM Clinical Pathology Department, Dr. Daryl Nydam, Ames Animal Shelter, Animal Rescue League of Iowa, Boone Area Humane Society, Dubuque Humane Society, and Iowa City Animal Care and Adoption Center. Prevalence of Cryptosporidium Parvum or Cryptosporidium Felis and Common Gastrointestinal Parasites in the Iowa Shelter Cat Population Sharon Brown; Laurie Meythaler-Mullins; Julie Ann Jarvinen, DVM, PhD; Douglas Jones, VMD, PhD; Christine Petersen, DVM, PhD; Claudia J Baldwin, DVM, MS College of Veterinary Medicine, Iowa State University, Ames, IA Abstract It has been reported that approximately 40% of cats in animal shelters are infected with one or more specie of gastrointestinal parasite 3. For this reason, parasite diagnosis and control is a major area of concern among shelter cat populations. Common gastrointestinal parasites are fairly easy to detect with zinc sulfate and sugar solution fecal floatations, and parasites can be managed effectively with proper deworming protocols 2 and shelter management. Cryptosporidium parvum (C. parvum) and Cryptosporidium felis (C. felis), microscopic gastrointestinal coccidian protozoans, are of rising concern to shelters, because Cryptosporidium is often misdiagnosed in companion animals and has zoonotic potential 4. The most common symptom of Cryptosporidium infection and most gastrointestinal diseases is watery diarrhea, but infection can also be accompanied by dehydration, weight loss, fever, and vomiting. Besides the fact that C. parvum and C. felis are difficult to diagnose, they are also concerning, because infected cats are not always symptomatic 6, but fully capable of spreading the parasite to others. Cats that are stressed, pregnant, or immunosuppressed are most likely to display symptoms. Our study found results similar to others in terms of common gastrointestinal parasite prevalence, but our results specific for Cryptosporidium were inconclusive. We found that 37% of shelter cats were infected with at least one specie of gastrointestinal parasite. More specifically, based on the 100 fecal samples we collected from 5 different Iowa shelters, we found the Iowa shelter cat population to have the following prevalence of parasites: 1% Ancyclostoma sp., 2% Trichuris sp., 16% Ascarid sp., 6% Cestode sp., 17% Coccidia sp., and 4% Giardia sp. 100% of our samples showed the presence of C. parvum or C. felis when analyzed with real-time PCR (polymerase-chain reaction). Although it is theoretically possible, it is unlikely that all samples were legitimately positive. More likely, the primers and probe used were not specific to C. parvum and C. felis or our samples became contaminated during the DNA extraction process. Even very small amounts of contamination at any point in the study could have effected our results, because we determined with serial dilutions that the primers and probe used in real-time PCR analysis were accurate at a DNA concentration of approximately 1 attogram (10 -18 grams) of DNA per microliter. We also found that the primers and probe used amplified pure T. foetus DNA, suggesting that the main reason all samples reported positive results during real-time PCR analysis was that the primers and probe were not specific to C. parvum and C. felis. The results of our study may be helpful to animal shelters in establishing effective deworming protocols, as well as help to build upon what is currently known about diagnostic assays for C. parvum and C. felis. Introduction Parasite diagnosis and control is a major area of concern among shelter cat populations. Gastrointestinal parasites – Ancyclostoma sp., Trichuris sp., Ascarid sp., Cestode sp., Coccidia sp., and Giardia sp. – can not only cause prolonged infection in cats, but can be spread easily from an infected cat to other cats and people 1. Proper shelter management, parasite screening, and deworming protocols greatly reduce the problem. Unlike these well known parasites that are relatively easy to detect, Cryptosporidium is more difficult to detect by routine methods. Cryptosporidium parvum (C. parvum) and Cryptosporidium felis (C. felis), however, are known to infect cats and cause symptoms of watery diarrhea, dehydration, weight loss, fever, and vomiting 5. Cryptosporidium is of rising concern, because pervious studies have suggested up to 38.8% of cats in the United States are infected with some specie of Cryptosporidium 5. Adding to the problem, not all infected cats display symptoms, thus there is a potential for cats to spread the disease without ever showing symptoms themselves 7. Infected cats that are stressed, pregnant, or immunosuppressed are most likely to display symptoms 4. Methods  100 fresh fecal samples were collected from cats at 5 different Iowa animal shelters.  Fecal matter from both healthy cats and cats showing signs of gastrointestinal parasite infection of all breeds and backgrounds was sampled.  Samples were transported on ice to ISU and then analyzed by sugar solution and zinc sulfate fecal floatation methods.  DNA was extracted from 200-mg samples of frozen feces using the QIAamp DNA stool minikit.  Real-time PCR (polymerase-chain reaction) was used to identify the presence of C. parvum or C. felis in each sample. Results  37% of cats tested were positive for at least one species of gastrointestinal parasite.  The total of 46 positive gastrointestinal parasite findings.  Prevalence of Ancyclostoma sp. was found to be 1%.  Prevalence of Trichuris sp. was found to be 2%.  Prevalence of Ascarid sp. was found to be 16%.  Prevalence of Cestode sp. was found to be 6%.  Prevalence of Coccidia sp. was found to be 17%.  Prevalence of Giardia sp. was found to be 4%.  63% of cats sampled showed gastrointestinal symptoms (diarrhea) and/or were infected with gastrointestinal parasites.  19% were symptomatic and tested positive for common parasites.  25% were symptomatic, but tested negative for common parasites.  19% were asymptomatic, but tested positive for common parasites.  Percentage of parasite positive cats from shelters ranged from 26.1 to 55.2%  Primers and probes could detect DNA in samples at a concentration of approximately 1 attogram.  100% of the 100 fecal samples collected and tested showed the presence of C. parvum or C. felis. Serial dilutions of purified C. parvum DNA sample (PCR) Percentage of positive infections caused by given GI parasite (n = 46 positive findings) Sugar solution fecal floatation positive for Cryptosporidium (Spherical oocysts of C. parvum) 1 Statement of Problem C. parvum and C. felis are difficult to detect in small animals and often misdiagnosed 6. Many animal shelter do not regularly screen for gastrointestinal parasites and lack appropriate parasitology records. Cryptosporidium species have an outer protective capsule that allow them to survive outside the body for long periods of time. Transmission of Cryptosporidium is hard to eliminate within shelters, because it is resistant to chlorine-based disinfectants, and because it can easily be carried from an infected animal to a non-infected animal on shelter employees and equiptment 4. Both C. parvum and C. felis have a zoonotic potential. Veterinary Summer Scholar Research Program, The College of Veterinary Medicine, Iowa State University Supported by the Maddie’s Fund, The Pet Rescue Foundation Specific Aims 1. Assess the prevalence of C. parvum and C. felis in the Iowa shelter cat population. 2. Determine the prevalence of common gastrointestinal parasites in the Iowa shelter cat population. 3. Evaluate current shelter parasitology records and make treatment protocol recommendations based on our findings of common gastrointestinal parasites. Positive curves for samples 1-38 and 96-100 (PCR) Percentage of cats infected (n = 100)Range of positive findings: Ancyclostoma sp. 0 to 3%; Trichuris sp. 0 to 18%; Ascarid sp. 0 to 41%; Cestode sp. 0 to 17%; Coccidia sp. 9 to 24%; Giardia sp. 0 to 19%