Immunomagnetic bead separation coupled with bead beating and real-time PCR was found to be a very effective procedure for the isolation, separation, and detection of Mycobacterium avium subsp. paratuberculosis from milk and/or fecal samples from cattle and American bison. Samples were spiked with M. avium subsp. paratuberculosis organisms, which bound to immunomagnetic beads and were subsequently lysed by bead beating; then protein and cellular contaminants were removed by phenol-chloroform-isopropanol extraction prior to DNA precipitation. DNA purified by this sequence of procedures was then analyzed by conventional and real-time IS900-based PCR in order to detect M. avium subsp. paratuberculosis in feces and milk. By use of this simple and rapid technique, 10 or fewer M. avium subsp. paratuberculosis organisms were consistently detected in milk (2-ml) and fecal (200-mg) samples, making this sensitive procedure very useful and costeffective for the diagnosis of clinical and subclinical Johne's disease (paratuberculosis) compared to bacteriological culture, which is constrained by time, labor, and expense under diagnostic laboratory conditions.
Background: The scientific evidence of the health risks associated with the consumption of raw milk has been known for a long time. However, less clear is the impact of acquiring infectious diseases from raw milk consumption in the United States (US) due to incomplete reporting of cases and the complex factors associated with the sale and consumption of raw milk. Investigations of this current study focused on human brucellosis, one of the infectious diseases commonly acquired through the consumption of raw milk and milk products, and which continues to be a public health threat worldwide.Methodology: A qualitative systematic review of the sources of opinions that contribute to the increased trend of raw milk sales and consumption in the US was conducted.Results: Interestingly, opinions about the sale of raw milk and/or the benefits arising from its consumption varied by US region, with the proportion of messages supporting raw milk consumption being highest in the Northeast compared to other US regions. Several evidence gaps and factors that possibly contribute to the increased prevalence of raw milk-acquired brucellosis were identified including inadequate monitoring of the raw milk sales process and lack of approved diagnostic methods for validating the safety of raw milk for human consumption. Conclusions:The unavailability of data specifying brucellosis cases acquired from raw milk consumption have precluded the direct association between raw milk and increased brucellosis prevalence in the United States. Nevertheless, the evidence gaps identified in this study demonstrate the need for intensified surveillance of raw-milk acquired infectious diseases including human brucellosis; establishment of safety and quality control measures for the process of selling raw milk; and design of an effective strategy for the prevention of raw milk-acquired infectious diseases including brucellosis. Overall, for the first time, this study has not only shown the gaps in evidence that require future investigations, but also, variations in the perception of raw milk consumption that may impact disease acquisition in different US regions.
The phagocytosis and destruction of pathogens in lysosomes constitute central elements of innate immune defense. Here, we show that Brucella, the causative agent of brucellosis, the most prevalent bacterial zoonosis globally, subverts this immune defense pathway by activating regulated IRE1α-dependent decay (RIDD) of Bloc1s1 mRNA encoding BLOS1, a protein that promotes endosome–lysosome fusion. RIDD-deficient cells and mice harboring a RIDD-incompetent variant of IRE1α were resistant to infection. Inactivation of the Bloc1s1 gene impaired the ability to assemble BLOC-1-related complex (BORC), resulting in differential recruitment of BORC-related lysosome trafficking components, perinuclear trafficking of Brucella-containing vacuoles (BCVs), and enhanced susceptibility to infection. The RIDD-resistant Bloc1s1 variant maintains the integrity of BORC and a higher-level association of BORC-related components that promote centrifugal lysosome trafficking, resulting in enhanced BCV peripheral trafficking and lysosomal destruction, and resistance to infection. These findings demonstrate that host RIDD activity on BLOS1 regulates Brucella intracellular parasitism by disrupting BORC-directed lysosomal trafficking. Notably, coronavirus murine hepatitis virus also subverted the RIDD–BLOS1 axis to promote intracellular replication. Our work establishes BLOS1 as a novel immune defense factor whose activity is hijacked by diverse pathogens.
This study investigated the interaction of genetic, environmental, and psychological factors in predicting depression and anxiety symptoms in medical students. First-year medical students (n = 141) completed measures of anxiety, depression, hope, and spiritual meaning at three time points during the first year. Buccal samples were utilized to genotype each individual at the s/l variant in the promoter region of the serotonin transporter gene (5-HTTLPR). A subset (n = 91) completed a measure of recent stressors. Mean levels of depression increased during year one of medical school. Presence of the s/s genotype in the promoter region of the 5-HTT gene was associated with greater increases in depression, but only in association with higher numbers of recent stressors. Spiritual meaning and hope were found to counteract genetic susceptibility to stress-related depressive symptoms in the more vulnerable s/s group.
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