Detection of Mycobacterium avium subspecies paratuberculosis in environmental samples by faecal culture and real-time PCR in relation to apparent within-herd prevalence as determined by individual faecal culture

Environmental samples are a cost-saving and easy-to-use approach to diagnose paratuberculosis at the herd level. Detailed knowledge concerning its uncertainties in herds with a low prevalence of Mycobacterium avium spp. paratuberculosis (MAP) is required to design sampling strategies in control programmes. This study aimed to calculate a threshold level of the apparent within-herd prevalence (WHPapp) as determined by individual mycobacterial cultivation (faecal culture; FC) of all cows thus allowing the detection of a herd as MAP-positive at a certain probability level (P-d). Out of 200 environmental samples taken twice from five predefined locations in a barn, 25 were positive by FC and 60 were positive by a quantitative real-time PCR method (qPCR). A logistic regression model was used to calculate the WHPapp threshold of detection. For 50% P-d, a WHPapp threshold of 2.9% was calculated for the combination of three samples (milking area, main cow alleyways, holding pen) tested simultaneously both by FC and qPCR. The threshold increased to 6.2% for 90% P-d. Repeated environmental sampling did not reduce the WHPapp threshold. Depending on the particular needs for prevalence estimation or in control programmes (single or repeated sampling) the provided WHPapp thresholds at different P-d will enable decisions to be made about various sampling strategies.