Development and validation of a real-time PCR assay for Cochlosoma anatis in turkeys

Abstract

Cochlosoma anatis is a flagellated protozoan parasite in the Trichomonadidae family that causes an enteric disease of turkeys and ducks known as cochlosomosis. Following the withdrawal of effective antiprotozoal medications from commercial poultry production, this organism has become of increased concern for the turkey industry. Historically, this organism has been detected by light microscopic evaluation of enteric mucosal scrapings. However, live trophozoites must be present in the sample for this method of detection because the parasite is differentiated from other protozoa by its characteristic movement. There is currently no real-time PCR (rtPCR) assay published for this organism; thus, we designed and validated a rtPCR assay. Ten-fold serial dilutions of gBlocks DNA representing the targeted region of the organism were evaluated to determine the limit of detection of the organism, which was ~50 copies per reaction. The assay was specific for C. anatis, with no cross-detection of 3 other poultry flagellates: Histomonas meleagridis, Tetratrichomonas gallinarum, and Trichomonas gallinae. Sensitivity and specificity compared to microscopic detection of live organisms were 100%. Coinfections of C. anatis with other pathogens have been reported. Thus, this assay can be added to syndromic panels for detection of enteric pathogens of turkeys.

Keywords

cochlosomosis protozoan turkeys

Get full access to this article

View all access options for this article.

References

Bermudez

. Miscellaneous and sporadic protozoal infections. In: Swayne

, et al, eds. Diseases of Poultry. 13th ed. Wiley, 2013:1182–1194.

Delgado-Viscogliosi

, et al Molecular phylogeny of parabasalids based on small subunit rRNA sequences, with emphasis on the Trichomonadinae subfamily. J Eukaryot Microbiol 2000;47:70–75.

Hampl

, et al Affiliation of Cochlosoma to trichomonads confirmed by phylogenetic analysis of the small-subunit rRNA gene and a new family concept of the order Trichomonadida. Int J Syst Evol Microbiol 2006;56(Pt 1):305–312.

Hussain

, et al Detection and quantification of Histomonas meleagridis by real-time PCR targeting single copy genes. Vet Parasitol 2015;212:382–388.

Lindsay

, et al Experimental Cochlosoma anatis infections in poultry. Vet Parasitol 1999;81:21–27.

Lowery

Evaluating the effects of Cochlosoma anatis infection on turkey poult gut health and performance. Master thesis. North Carolina State University, Raleigh, NC, 2023. https://repository.lib.ncsu.edu/handle/1840.20/41234

Lowery

, et al Cochlosoma anatis in poultry and its industry impact. Avian Dis 2024;68:169–174.

McElroy

, et al Molecular assay for the detection of Cochlosoma anatis in house flies and turkey specimens by polymerase chain reaction. Vet Parasitol 2005;127:165–168.

Sigrist

, et al A new duplex real-time PCR for simultaneous detection and differentiation of Tetratrichomonas gallinarum and Trichomonas gallinae. J Vet Diagn Invest 2022;34:631–637.