Detection of Coxiella burnetii DNA in sheep and goat milk and dairy products by droplet digital PCR in south Italy

Abstract

Coxiella burnetii is a Gram-negative obligate intracellular bacterium that is responsible for Q fever, a common zoonosis which is present virtually worldwide. This microorganism infects a wide range of wild and domestic mammals, but the main reservoirs are cattle, goats and sheep, which also represent sources of human infection. A potential route of transmission of this pathogen to humans is the consumption of C. burnetii-contaminated raw milk or dairy products derived from contaminated raw milk, although the role of these foods as possible infection sources is controversial. The aims of this study were (i) to apply two ddPCR based assays targeting the C. burnetii IS1111 and icd genes for the detection and quantification of C. burnetii DNA, and (ii) to evaluate the occurrence of C. burnetii DNA in raw milk and raw milk products from sheep and goats in Apulia and Basilicata regions of Southern Italy. Of 413 milk and cheese samples tested, 78 were positive for the presence of C. burnetii DNA (18.9%), specifically, 68 of 285 milk samples (23.9%) and 10 of 128 cheese samples (7.8%) The presence of both IS1111 and icd genes was detected in only 2 (2.6%) of the 78 positive samples, while the remaining 76 (97.4%) were positive only for IS1111. C. burnetii DNA was specifically detected by the ddPCR method, whereas no cross-amplification was observed with the DNA of other foodborne bacterial pathogens. The sensitivity of the ddPCR method was determined as 0.35 and 0.56 copies/μL for IS1111 and icd genes, respectively. The findings of this study demonstrate the presence of C. burnetii DNA in a significant proportion of raw milk and dairy products. Although there is no conclusive epidemiological evidence that C. burnetii infection occurs via food, the presence of this organism in raw milk and dairy products made of raw milk should be considered a potential hazard. ddPCR is a useful tool to investigate the quality and safety of food products due to its sensitivity and precision, and could be applied to routine testing.

Keywords: Cheese; Coxiella burnetii; Food safety; IS1111; Raw milk; icd.