The association of detection method, season, and lactation stage on identification of fecal shedding in Mycobacterium avium ssp. paratuberculosis infectious dairy cows

Abstract

Mycobacterium avium ssp. paratuberculosis (MAP) is the causative organism of Johne's disease. Although fecal culture is considered the standard diagnostic test, the long incubation times, costs, and intermittent shedding of MAP hinder efficient screening programs based on culture results. The primary objectives of this study were to determine the detection ability of solid culture, broth culture, and real-time PCR (qPCR) for MAP in fecal samples and to assess how shedding patterns of MAP in feces vary with lactation stage and season. This knowledge could improve the use of these diagnostic assays in Johne's management programs. For this study, 51 MAP-infectious cows from 7 Atlantic Canadian dairy farms had fecal samples collected monthly over a 12-mo period. Samples were analyzed for MAP bacterial load via solid culture, broth culture, and qPCR. For all fecal samples, 46% [95% confidence interval (CI): 40 to 51%] were positive by solid culture, 55% (95% CI: 50 to 60%) by broth culture, and 78% (95% CI: 73 to 82%) by qPCR. Sensitivity of qPCR was numerically higher in the dry and postpartum lactation periods, and qPCR detection in summer and fall was 85% of that in winter and spring. Furthermore, culture-determined moderate or light shedding categories generally corresponded to qPCR cycle threshold values <35, but heavy shedding categories corresponded to qPCR values <29. Direct fecal qPCR is a MAP detection method that is quick and less costly than culture techniques, and it avoids the use of decontamination steps that could decrease numbers of bacteria in a sample below the detection limit. This study indicates that, for known MAP-positive cows, fecal qPCR had high sensitivity of MAP detection, thereby supporting the use of direct fecal qPCR as part of a Johne's herd control program.

Association of paratuberculosis sero-status with milk production and somatic cell counts across 5 lactations, using multilevel mixed models, in dairy cows

“…Serum ELISA measures humoral response to the Association of paratuberculosis sero-status with milk production and somatic cell counts across 5 lactations, using multilevel mixed models, in dairy cows presence of MAP. During early stages of infection, a cellular immune response is produced, whereas antibody production develops in later stages of infection, which translates to different test accuracy in different stages of infection (Stabel et al, 2014;Laurin et al, 2015Laurin et al, , 2017. Although serum ELISA specificity is considered to be greater than 97%, sensitivity can vary from 15 to 75% depending on the stage of the disease (Timms et al, 2011;Mitchell et al, 2015) and also on milk yield, lactation, herd prevalence, DIM, milk protein, and SCC (Eisenberg et al, 2015).…”

Estimation of the sensitivity and specificity of two serum ELISAs and one fecal qPCR for diagnosis of paratuberculosis in sub-clinically infected young-adult French sheep using latent class Bayesian modeling

“…It is less time consuming, especially for the detection of S-strains (sheep strains) of MAP that grow slowly in vitro compared to C-strains (cattle strains) [ 22 ]. There is also growing evidence that fecal qPCR might be at least as sensitive as, or even more sensitive than, fecal culture [ 23 , 24 ]. However, its analytical sensitivity depends on several factors, including sample quality, DNA extraction methods, DNA target and qPCR systems [ 25 , 26 ].…”

Knowledge gaps that hamper prevention and control ofMycobacterium aviumsubspeciesparatuberculosisinfection

“…Another reason for using ELISAs is that the extent of shedding correlates well with the ELISA titres (Dargatz et al., ), and hence, culling ELISA positives may be an effective and relatively low‐cost option for removing high shedders. Currently, faecal culture is often replaced with direct PCR on faeces (e.g., Laurin, Chaffer, McClure, McKenna, & Keefe, ; Plain et al., ).…”

“…Quantitative PCR (qPCR) for assessing the number of MAP bacteria in faeces has great promise in comparison with bacterial culture (Christopher‐Hennings et al., ; Khare et al., ; Kralik, Beran, & Pavlik, ; Laurin et al., ) (Table ). This technique has been validated on spiked faecal samples and proficiency panels, including faecal samples from naturally infected animals (e.g., Plain et al., ).…”

“…Currently, faecal culture is often replaced with direct PCR on faeces (e.g., Laurin, Chaffer, McClure, McKenna, & Keefe, 2015;Plain et al, 2014). JD control programmes have been implemented in most developed countries, with objectives based on the national economic situation of the cattle, sheep and goat industry and the herd-level prevalence of MAP infection (reviewed by Kennedy, 2011;Geraghty, Graham, Mullowney, & More, 2014).…”