ABSTRACT
The initial infection by the obligate intracellular bacillus Mycobacterium leprae evolves to leprosy in a small subset of the infected individuals. Transmission is believed to occur mainly by exposure to bacilli present in aerosols expelled by infected individuals with high bacillary load. Mycobacterium leprae-specific DNA has been detected in the blood of asymptomatic household contacts of leprosy patients years before active disease onset, suggesting that, following infection, the bacterium reaches the lymphatic drainage and the blood of at least some individuals. The lower temperature and availability of protected microenvironments may provide the initial conditions for the survival of the bacillus in the airways and skin. A subset of skin-resident macrophages and the Schwann cells of peripheral nerves, two M. leprae permissive cells, may protect M. leprae from effector cells in the initial phase of the infection. The interaction of M. leprae with these cells induces metabolic changes, including the formation of lipid droplets, that are associated with macrophage M2 phenotype and the production of mediators that facilitate the differentiation of specific T cells for M. leprae-expressed antigens to a memory regulatory phenotype. Here, we discuss the possible initials steps of M. leprae infection that may lead to active disease onset, mainly focusing on events prior to the manifestation of the established clinical forms of leprosy. We hypothesize that the progressive differentiation of T cells to the Tregs phenotype inhibits effector function against the bacillus, allowing an increase in the bacillary load and evolution of the infection to active disease. Epigenetic and metabolic mechanisms described in other chronic inflammatory diseases are evaluated for potential application to the understanding of leprosy pathogenesis. A potential role for post-exposure prophylaxis of leprosy in reducing M. leprae-induced anti-inflammatory mediators and, in consequence, Treg/T effector ratios is proposed.
ABSTRACT
Until there is an effective implementation of COVID-19 vaccination program, a robust testing strategy, along with prevention measures, will continue to be the most viable way to control disease spread. Such a strategy should rely on disparate diagnostic tests to prevent a slowdown in testing due to lack of materials and reagents imposed by supply chain problems, which happened at the beginning of the pandemic. In this study, we have established a single-tube test based on RT-LAMP that enables the visual detection of less than 100 viral genome copies of SARS-CoV-2 within 30 minutes. We benchmarked the assay against the gold standard test for COVID-19 diagnosis, qRT-PCR, using 177 nasopharyngeal RNA samples. For Ct ≤ 32, the RT-LAMP assay had a sensitivity of 100% and a specificity of 96.1%. Additionally, we set up a RNA extraction-free RT-LAMP test capable of detecting SARS-CoV-2 directly from saliva samples, albeit with lower sensitivity. The saliva was self-collected and the collection tube remained closed until inactivation, thereby ensuring the protection of the testing personnel. As expected, RNA extraction from saliva samples increased the sensitivity of the test. To lower the costs associated with RNA extraction, we performed this step using an alternative protocol that uses plasmid DNA extraction columns. We also produced the enzymes needed for the assay and established an in-house-made RT-LAMP test independent of specific distribution channels. Finally, we developed a new colorimetric method that allowed the detection of LAMP products by the visualization of an evident color shift, regardless of the reaction pH.