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United States Department of Agriculture

Agricultural Research Service

Research Project: IMMUNOLOGY AND INTERVENTION STRATEGIES FOR JOHNE'S DISEASE Title: Optimization of methods for the detection of Mycobacterium avium subsp. paratuberculosis in milk and colostrum of naturally infected dairy cows

Authors
item Bradner, Laura -
item Robbe-Austerman, Suelee -
item Beitz, D. -
item Stabel, Judith

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: December 1, 2011
Publication Date: February 4, 2012
Citation: Bradner, L., Robbe-Austerman, S., Beitz, D., Stabel, J.R. 2012. Optimization of methods for the detection of Mycobacterium avium subsp. paratuberculosis in milk and colostrum of naturally infected dairy cows [abstract]. 11th International Colloquium on Paratuberculosis. p. 183.

Technical Abstract: Mycobacterium avium subsp. paratuberculosis (MAP) is primarily shed into the feces but it has also been isolated from the milk and colostrum of cows. Because of this, there exists concern about transfer of the organism from dam to calf and about the prevalence of MAP in the milk supply. The prevalence of MAP in milk has is not well defined because of the complexity of the milk matrix. The objective of this study is to optimize the decontamination of whole raw milk for the isolation of viable MAP and compare recovery rates in liquid and solid culture mediums. The efficacy of two liquid culture mediums, TREK-ESP and BD Bactec 12B were compared for recovery thresholds, speed of recovery, incidence of contamination and reproducibility of results. Milk collected from a non-infected cow was spiked with MAP (102 to 108 cfu/ml). Variables investigated included concentration of hexadecylpyridinium chloride (HPC) as the primary decontaminant, temperature of decontamination, centrifugation speed, and time of incubation. It was found that neither length of exposure to HPC or concentration of HPC had significant effects on the recovery of MAP from milk. Because of known lethal effects of HPC on MAP, the most efficient decontamination parameters with the highest recovery rates of MAP were 0.75% HPC exposed for 5 hr at room temperature. Since increased temperature (>25°C) enhanced growth of microbial contaminants, room temperature was the optimal choice. In comparing the two liquid culture mediums, Bactec 12B was superior in recovery thresholds and speed of recovery of viable MAP. TREK-ESP culture demonstrated an increased incidence in false positive and false negative results that were not observed in Bactec 12B medium. Future decontamination studies will evaluate the efficacy of N-acetyl-L-cysteine (NALC)-sodium hydroxide. Optimized methods will be used to assess the frequency and level of MAP shed into milk during a complete lactation period of naturally infected dams.

Last Modified: 9/22/2014
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