Quantitation of viable Coxiella burnetii in milk products using a liquid medium-based MPN-PCR assay

This Technical Research Communication describes a new method by which thermally treated Coxiella in milk products may be grown in a liquid growth medium and quantitated using an MPN-PCR assay. Coxiella is generally not used in studies on thermal and non-thermal processing of milk due to the need for specialized and highly laborious techniques such as animal assays and tissue culture for determining viability. Recently, a liquid growth medium (ACCM-2) and modified atmosphere were used to grow Coxiella from pure cultures, infected mouse tissues, and clinical samples, however, the ability to grow Coxiella from a food such as milk has not been shown. The potential ability to enrich Coxiella directly from contaminated milk presents a new avenue for conducting pasteurization research in which the viability of heat-treated or injured cultures could be more easily determined through direct enrichment of Coxiella in ACCM-2. ACCM-2 medium allowed enrichment of Coxiella from bovine whole milk and cream, whole goat, and whole camel milks but not whole water buffalo milk. Enrichment was possible from whole bovine milk containing as few as 6 Coxiella ge/ml of milk. The applicability of this ACCM-2 enrichment method was shown when using an MPN-PCR assay to quantitate the number of viable Coxiella remaining in whole bovine milk after 64 °C thermal treatment for up to 10 min.