PCR Testing & Sheath Wash

Sheath wash PCR testing principals, specimen collection and submission

PCR TEST FOR BULL SHEATH WASH

Due to the constant improvement in the accuracy of tests done by human and animal laboratories, PathCare Vetlab made a decision to offer a PCR test to detect the presence of Tritrichomonas foetus and Campylobacter Fetus in bull sheath washes and scrapings. One of the most rapidly developing diagnostic fields is in molecular biology and specifically the PCR (polymerase chain reaction) test. This test relies on detection of nucleic acids found in living organisms. The nucleic acids DNA (deoxyribonucleic acid) and RNA (ribonucleic acid) contain the genetic information that is unique to a species (e.g. human, animal, plant, parasite, bacterium and virus) and an individual member of the species. PCR is a technique that permits the analysis of any short segment of DNA (or RNA) by reproducing (amplifying) selected sections of the genetic material. These sequences are compared to the nucleotide segments from a known source and can be used to identify the origin of the specific genetic material. The PathCare PCR laboratory has developed tests to identify Tritrichomonas foetus and Campylobacter fetus subspecies venerealis and C. fetus subspecies fetus in sheath wash samples from bulls. The tests that were developed are validated against known ATCC (American Type Culture Collection) strains of the organisms. . The advantage of this test is that PCR is specific for the organisms and an accurate result is obtained.

BACKGROUND OF REPRODUCTIVE DISEASES IN BULLS

Trichomonosis is a venereal disease of cattle and once established in a herd, very difficult to eradicate. It is a problem in beef cattle and causes repeat breeding, abortions and a high percentage of unbred cows. Bulls are the main source of infection and animals over 4 years of age are long-term carriers of the organism. They show no clinical signs. In cows (and heifers) inflammation of the reproduction tract varies from vaginal discharge to pyometra. In pregnant cows placentitis leads to early embryonal death (1 – 16 weeks). There is a poor uterine immunity in cows and they become susceptible to re-infection within 3-4 months.

Bovine genital campylobacteriosis (vibriosis) is caused by Campylobacter fetus subspecies venerealis. It causes poor conception, increased return to service, reduced calving rates, occasional abortions and permanent infertility. The disease is spread by asymptomatic infected bulls. Many infected cows “self-cure” in 3-4 months. Some cows develop permanent infertility due to chronic changes in the reproductive tract. Campylobacter fetus subspecies fetus is an inhabitant of the intestine of cattle and may cause sporadic abortions in cattle and sheep.

SAMPLE REQUIREMENTS

Sheath wash samples from bulls is collected and transported in phosphate buffered saline (PBS) (Ordered from the Vetlab) .We recommend that the samples be kept on ice after collection and for transportation to the laboratory to ensure an optimal PCR test result. Samples should be submitted to the laboratory as soon as possible after collection. Sensitivity of the tests may be influenced by contamination by sand, faecal and plant material as well as blood and urine. Sensitivity is decreased with delay in sample submission. The PBS should not be older than three (3) months. Please take note of the expiry date and store in a fridge until required.

GUIDELINES FOR BULL CERTIFICATION

T. foetus and C. fetus infection must be absent in samples for three (3) consecutive PCR tests on preputial scrapings and or washes collected at 7-10 days intervals. A bull should be allowed at least 2 weeks sexual rest before implementing the testing procedure. The reason for the three (3) consecutive negative tests, is that the organisms are not persistently present in the preputium. A positive bull may produce negative results.

References

Prof. Gert Pretorius and Ms. Jaclyn Gerber (2012) (PathCare PCR Department). Personal communication

www.medterms.com “Definition of Polymerase Chain Reaction” Peter Nosworthy (2009), Veterinary Officer, PIRSA. “Bovine genital campylobacteriosis”

Frank Schulze, Audrey Bagon, Wolfgang Muller and Helmut Hotzel (2006) J. Clin. Microbiol. Vol. 44 no. 6 2019-2024 “Identification of Campylobacter fetus Subspecies by Phenotypic Differentiation and PCR”

Dietmar Holm (2013), Dept of Production Animal Studies, Faculty ofVeterinary Science, University of Pretoria. “Choosing the correct Trichomonosis diagnostic protocol”

Dr. Lucia Lange , Mr. Johann Muller Dr. Anita Schewegman(2016-2017) (PathCare PCR Department and Vet Lab)