Conversion of Mycobacterium smegmatis to a pathogenic phenotype via passage of epithelial cells during macrophage infection

Abstract

Mycobacteria encounter many different cells during infection within their hosts. Although alveolar epithelial cells play an essential role in host defense as the first cells to be challenged upon contact with mycobacteria, they may contribute to the acquisition of mycobacterial virulence by increasing the expression of virulence or adaptation factors prior to being ingested by macrophages on the side of pathogens. From this aspect, the enhanced virulence of nonpathogenic Mycobacterium smegmatis (MSM) passed through human alveolar A549 epithelial cells (A-MSM) was compared to the direct infection of MSM (D-MSM) in THP-1 macrophages and mouse models. The intracellular growth rate and cytotoxicity of A-MSM were significantly increased in THP-1 macrophages. In addition, compared to D-MSM, A-MSM induced relatively greater interleukin (IL)-1β, IL-6, IL-8, IL-12, TNF-α, MIP-1α, and MCP-1 in THP-1 macrophages. As a next step, a more persistent A-MSM infection was observed in a murine infection model with the development of granulomatous inflammation. Finally, 58 genes induced specifically in A-MSM were partially identified by differential expression using a customized amplification library. These gene expressions were simultaneously maintained in THP-1 infection but no changes were observed in D-MSM. Bioinformatic analysis revealed that these genes are involved mainly in bacterial metabolism including energy production and conversion, carbohydrate, amino acid, and lipid transport, and metabolisms. Conclusively, alveolar epithelial cells promoted the conversion of MSM to the virulent phenotype prior to encountering macrophages by activating the genes required for intracellular survival and presenting its pathogenicity.

Increased survival and proliferation of the epidemic strain Mycobacterium abscessus subsp. massiliense CRM0019 in alveolar epithelial cells

“…Because of the evidence that mycobacteria become more infective after the first intracellular infection [ 61 , 62 ], we evaluated whether this holds for M. abscessus obtained from macrophages. The aforementioned results showed that M. abscessus were highly internalized by macrophages and ended up in a mature lysosomal compartment without, however, evidence of intracellular death (Figs.…”

“…Although second infection occurred to a lesser degree when compared to initial infection, only the clinical strain CRM0019 grew in A549 cells and macrophages in a second infection, which may explain the long persistence of the observed in vivo infection [ 32 ]. Evidence suggests that intracellular passage through epithelial cells changes the mycobacterial phenotype to a more pathogenic one during infection of macrophages, epithelial or endothelial cells [ 61 , 77 , 78 ]. In our study, mycobacteria obtained from macrophages were not more infective but were still able to survive in either macrophages or alveolar epithelial cells.…”

“…Besides being shielded from phagocytic cells, mycobacteria inside AEC could acquire a more infective phenotype. Indeed, it has been shown that passage of mycobacteria through alveolar epithelial cells promotes the conversion of non-virulent M. smegmatis to a virulent phenotype before they infect macrophages [ 61 ]. Likewise, a recent study also showed that M. abscessus obtained from amoeba are more infective than those growing extracellularly [ 62 ].…”

Quantifying intracellular Mycobacterium tuberculosis: An essential issue for in vitro assays

“…Also was found the rapid stirring on ice in a overhead stirrer to avoid agglutination, followed by passage through a 0.8 μm membrane to guarantee a predominance of bacteria just in controls by Ziehl-Neelsen staining (Shin, Han, Manning, & Collins, 2007). Lastly, there is also gentle sonication over short periods of time, for example, at 30 W for 5 min (Chapeton-Montes, Plaza, Barrero, & Patarroyo, 2007;Kim et al, 2011).…”

Host-Mycobacterium avium subsp. paratuberculosis interactome reveals a novel iron assimilation mechanism linked to nitric oxide stress during early infection

“…Epithelium processing and interaction with bovine mammary epithelial cells (MAC-T), a surrogate for the intestinal epithelium, results in enhanced phagocytosis during secondary infection [ 23 ]. Changes to invasion phenotype due to epithelial processing are also seen with M. smegmatis , a saprophytic mycobacteria that is nonpathogenic [ 24 ]. M. smegmatis exposed to A549 epithelial cells had a significant increase in intracellular growth during secondary infection in THP-1 macrophages [ 24 ].…”