Submitted to: American Association of Veterinary Parasitologists Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: 7/1/2005
Publication Date: 8/1/2005
Citation: Harmon, A.F., Zarlenga, D.S., Hildreth, M.B. 2005. Field use of a multiplex pcr for the differnetiation of strongylenematodes in cattle. American Association of Veterinary Parasitologists Paper No. 19. [Abstract]. Interpretive Summary:
Technical Abstract: Despite significant losses from strongyle parasites of cattle, there is still no practical method for differentiating eggs from the five prevalent genera in the U.S (Cooperia, Ostertagia, Trichostrongylus, Haemonchus, and Oesophagostomum) obtained from fecal samples. A multiplex PCR assay has recently been described to differentiate the eggs at the genus level. This assay was designed to detect DNA extracted from fecal eggs and was validated using DNA from larva and eggs from experimentally-infected animals; however, the test has yet to be applied to field samples from naturally-infected animals. There was also a need to develop a more rapid method for isolating DNA from the eggs. To this end, we first infected two calves with Ostertagia ostertagi as a source of strongyle eggs. These eggs were used to evaluate different lysis methods using three different commercial DNA extraction kits. Eggs were also used to develop an iodine based system of preservation to prevent larval development but still permit PCR quality DNA to be extracted from eggs that have been stored at room temperature. With an optimized DNA extraction procedure, we began testing the assay on herds in South Dakota. Samples from 4 herds from Eastern South Dakota were obtained, and the assay was performed on these. Cooperia and Ostertagia were detected in all herds, and one herd also contained Haemonchus. As quality control, every tenth sample is sequenced, and the larval identification was performed.