Molecular Investigation and Characterization of Enterocytozoon bieneusi, Coxiella burnetii and Dientamoeba fragilis in Raw Milk and Cheese: A First Report for Dientamoeba fragilis.

purposeRaw milk and its products are increasingly popular among consumers recently. Milk already harbors a primary microbial flora during milking but can be further contaminated at any stage with pathogenic microorganisms. Although, the presence of C. burnetii in milk and other dairy products is frequently reported, E. bieneusi has been documented in a few studies, and D. fragilis has not been reported previously. Therefore, this study aimed to evaluate the contamination of raw milk from cattle, sheep, and goats, as well as cheese from cattle, with these pathogens to assess public health risk.

methodsA total of 150 samples from cattle, sheep, goat and homemade ready-to-eat cattle milk cheese were collected from farms, open-air markets, and delicatessen were examined using PCR analysis. Pathogen positive samples were sequenced for species identification and genotyping.

resultsThe overall prevalence of E. bieneusi, C. burnetii, and D. fragilis in the samples was 2.0%, 8.6%, and 6.0%, respectively. In cattle milk cheese samples, the prevalence was 3.3% for E. bieneusi, 10.0% for C. burnetii, and 3.3% for D. fragilis. The potentially zoonotic E. bieneusi BEB6 genotype was found in sheep milk and cattle milk cheese. D. fragilis isolates detected in cattle and sheep milk and cattle milk cheeses showed homology with genotype 1 from human, cattle and budgerigars in various countries including Türkiye.

conclusionThis is the first report of D. fragilis in milk and cattle milk cheese samples, as well as E. bieneusi in cattle milk cheese samples. These findings provide critical insights into the zoonotic transmission potential of these foods and contribute to a better understanding of epidemiology and public health risk.