A Murine Model of Mycobacterium kansasii Infection Reproducing Necrotic Lung Pathology Reveals Considerable Heterogeneity in Virulence of Clinical Isolates.

Among non-tuberculous mycobacteria, Mycobacterium kansasii is one of the most pathogenic, able to cause pulmonary disease indistinguishable from tuberculosis in immunocompetent susceptible adults. The lack of animal models that reproduce human-like lung disease, associated with the necrotic lung pathology, impairs studies of M. kansasii virulence and pathogenicity. In this study, we examined the ability of the C57BL/6 mice, intratracheally infected with highly virulent M. kansasii strains, to produce a chronic infection and necrotic lung pathology. As a first approach, we evaluated ten M. kansasii strains isolated from Brazilian patients with pulmonary disease and the reference strain M. kansasii ATCC 12478 for virulence-associated features in macrophages infected in vitro; five of these strains differing in virulence were selected for in vivo analysis. Highly virulent isolates induced progressive lung disease in mice, forming large encapsulated caseous granulomas in later stages (120-150 days post-infection), while the low-virulent strain was cleared from the lungs by day 40. Two strains demonstrated increased virulence, causing premature death in the infected animals. These data demonstrate that C57BL/6 mice are an excellent candidate to investigate the virulence of M. kansasii isolates. We observed considerable heterogeneity in the virulence profile of these strains, in which the presence of highly virulent strains allowed us to establish a clinically relevant animal model. Comparing public genomic data between Brazilian isolates and isolates from other geographic regions worldwide demonstrated that at least some of the highly pathogenic strains isolated in Brazil display remarkable genomic similarities with the ATCC strain 12478 isolated in the United States 70 years ago (less than 100 SNPs of difference), as well as with some recent European clinical isolates. These data suggest that few pathogenic clones have been widely spread within M. kansasii population around the world.

Exposition to Biological Control Agent Trichoderma stromaticum Increases the Development of Cancer in Mice Injected With Murine Melanoma.

Biological control agents (BCA) are an alternative to chemical pesticides and an emerging strategy to safely eliminate plant pathogens. Trichoderma spp. are the most common fungi used as BCAs. They produce spores that are released into the air and can potentially interact with immune system of mammals. We previously showed that Trichoderma affects expression of genes encoding pattern recognition receptors (PRRs) and cytokines in mice. PRRs are involved in the recognition of microorganisms and can lead to pro-tumoral signaling. Here, we evaluated if mice injected with low doses of murine melanoma exhibited increased development of lung tumor when treated with conidia of T. stromaticum. Mice treated with T. stromaticum and inoculated with B16-F10 melanoma cells exhibited significant increase in tumor uptake (p = 0.006) and increased number of visible nodules in the lungs (p = 0.015). We also analyzed mRNA expression levels of genes encoding PRRs in lung of mice exposed to T. stromaticum and demonstrated that mice treated with T. stromaticum conidia exhibited lower expression levels of Clec7a and increased expression of Tlr4 (toll like receptor 4) compared to non-treated controls. The expression levels of Clec7a and Tlr2 were increased in mice treated with T. stromaticum and inoculated with murine melanoma compared to controls only inoculated with melanoma. Our results demonstrate that intranasal exposition to T. stromaticum increases tumor in the B16-F10 model, which may raise concerns regarding the safety of its use in agriculture.

Evaluation of the effects of Quercetin and Kaempherol on the surface of MT-2 cells visualized by atomic force microscopy.

This study investigated the anti-viral effects of the polyphenolic compounds Quercetin and Kaempherol on the release of HTLV-1 from the surface of MT-2 cells. Atomic force microscopy (AFM) was used to scan the surface of the MT-2 cells. MT-2 cells were fixed with 100% methanol on round glass lamina or cleaved mica and dried under UV light and laminar flow. The images were captured on a Multimode equipment monitored by a NanoScope IIId controller from Veeco Instruments Inc operated in tapping mode and equipped with phase-imaging hardware. The images demonstrated viral budding structures 131 ± 57 nm in size, indicating profuse viral budding. Interestingly, cell-free viruses and budding structures visualized on the surface of cells were less common when MT-2 was incubated with Quercetin, and no particles were seen on the surface of cells incubated with Kaempherol. In summary, these data indicate that HTLV-1 is budding constantly from the MT-2 cell surface and that polyphenolic compounds were able to reduce this viral release. Biological samples were analyzed with crude cell preparations just after cultivation in the presence of Quercetin and Kaempherol, showing that the AFM technique is a rapid and powerful tool for analysis of antiviral activity of new biological compounds.