Abstract
Mycobacterium avium subspecies paratuberculosis (MAP) are detectable viable in milk and other dairy products. The molecular mechanisms allowing the adaptation of MAP in these products are still poorly understood. To obtain information about respective adaptation of MAP in milk, we differentially analyzed the proteomes of MAP cultivated for 48 h in either milk at 37 °C or 4 °C or Middlebrook 7H9 broth as a control. From a total of 2197 MAP proteins identified, 242 proteins were at least fivefold higher in abundance in milk. MAP responded to the nutritional shortage in milk with upregulation of 32% of proteins with function in metabolism and 17% in fatty acid metabolism/synthesis. Additionally, MAP upregulated clusters of 19% proteins with roles in stress responses and immune evasion, 19% in transcription/translation, and 13% in bacterial cell wall synthesis. Dut, MmpL4_1, and RecA were only detected in MAP incubated in milk, pointing to very important roles of these proteins for MAP coping with a stressful environment. Dut is essential and plays an exclusive role for growth, MmpL4_1 for virulence through secretion of specific lipids, and RecA for SOS response of mycobacteria. Further, 35 candidates with stable expression in all conditions were detected, which could serve as targets for detection. Data are available via ProteomeXchange with identifier PXD027444.
Highlights
Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of paratuberculosis or Johne’s disease, a chronic granulomatous enteritis of cattle and small ruminants [1,2]
To analyze if Mycobacterium avium ssp. paratuberculosis (MAP) changes its protein expression through incubation in milk, we performed a differential proteomics experiment using quantitative liquid chromatography–tandem mass spectrometry (LC–MS/MS) analysis of MAP incubated in Middlebrook 7H9 (M7H9) standard cell culture medium and milk at 37 ◦C and at 4 ◦C
The analysis revealed that 32% of the proteins were assigned to the function metabolism (Figure 3, blue), 19% to each stress response/immune evasion/antibiotic resistance (Figure 3, green) and to transcription/translation (Figure 3, pink), 17% to fatty acid metabolism/synthesis (Figure 3, gray), and 13% to cell wall synthesis/cell division (Figure 3, yellow)
Summary
Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of paratuberculosis or Johne’s disease, a chronic granulomatous enteritis of cattle and small ruminants [1,2]. Viable MAP were found in pasteurized milk, cheese, and even in dried milk products such as infant formula [12,13,14]. This might have been caused by post-processing contamination, but there is evidence of MAP being able to survive the pasteurization process [15,16,17,18,19]. The mechanisms behind this tenacity, are not fully understood and have been vigorously discussed over the last years [8,20]. Theories include clumping, intracellular localization of MAP in milk, and formation of heat-resistant forms such as endospores [16,21,22]
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