MicroRNAs (miRNAs) are short non-coding RNAs that regulate the expression of protein coding genes of viruses and eukaryotes at the post-transcriptional level. The eukaryotic genes regulated by miRNAs include those whose products are critical for biological processes such as cell proliferation, metabolic pathways, immune response, and development. It is now increasingly recognized that modulation of miRNAs associated with biological processes is one of the strategies adopted by bacterial pathogens to survive inside host cells. In this review, we present an overview of the recent findings on alterations of miRNAs in the host cells by facultative intracellular bacterial pathogens. In addition, we discuss how the altered miRNAs help in the survival of these pathogens in the intracellular environment.

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Mycobacterium bovis BCG is widely used as a vaccine against tuberculosis due to M. tuberculosis (Mtb), which kills millions of people each year. BCG variably protects children, but not adults against tuberculosis. BCG evades phagosome maturation, autophagy, and reduces MHC-II expression of antigen-presenting cells (APCs) affecting T-cell activation. To bypass these defects, an autophagy-inducing, TLR-2 activating C5 peptide from Mtb-derived CFP-10 protein was overexpressed in BCG in combination with Ag85B. Recombinant BCG 85C5 induced a robust MHC-II-dependent antigen presentation to CD4 T cells in vitro, and elicited stronger T H 1 cytokines (IL-12, IL-1β, and TNFα) from APCs of C57Bl/6 mice increasing phosphorylation of p38MAPK and ERK. BCG 85C5 also enhanced MHC-II surface expression of MΦs by inhibiting MARCH1 ubiquitin ligase that degrades MHC-II. BCG 85C5 infected APCs from MyD88 or TLR-2 knockout mice showed decreased antigen presentation. Furthermore, BCG 85C5 induced LC3-dependent autophagy in macrophages increasing antigen presentation. Consistent with in vitro effects, BCG 85C5 markedly expanded both effector and central memory T cells in C57Bl/6 mice protecting them against both primary aerosol infection with Mtb and reinfection, but was less effective among TLR-2 knockout mice. Thus, BCG 85C5 induces stronger and longer lasting immunity, and is better than BCG against tuberculosis of mice.

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The organic hydroperoxide stress resistance regulator (OhrR) is a MarR type of transcriptional regulator that primarily regulates the expression of organic hydroperoxide reductase (Ohr) in bacteria. In mycobacteria, the genes encoding these proteins exist in only a few species, which include the fast-growing organism Mycobacterium smegmatis. To delineate the roles of Ohr and OhrR in defense against oxidative stress in M. smegmatis, strains lacking the expression of these proteins were constructed by deleting the ohrR and ohr genes, independently and together, through homologous recombination. The OhrR mutant strain (MS⌬ohrR) showed severalfold upregulation of Ohr expression, which could be observed at both the transcript and protein levels. Similar upregulation of Ohr expression was also noticed in an M. smegmatis wild-type strain (MSWt) induced with cumene hydroperoxide (CHP) and t-butyl hydroperoxide (t-BHP). The elevated Ohr expression in MS⌬ohrR correlated with heightened resistance to oxidative stress due to CHP and t-BHP and to inhibitory effects due to the antituberculosis drug isoniazid (INH). Further, this mutant strain exhibited significantly enhanced survival in the intracellular compartments of macrophages. In contrast, the strains lacking either Ohr alone (MS⌬ohr) or both Ohr and OhrR (MS⌬ohr-ohrR) displayed limited or no resistance to hydroperoxides and INH. Additionally, these strains showed no significant differences in intracellular survival from the wild type. Electrophoretic mobility shift assays (EMSAs) revealed that the overexpressed and purified OhrR interacts with the ohr-ohrR intergenic region with a greater affinity and this interaction is contingent upon the redox state of the OhrR. These findings suggest that Ohr-OhrR is an important peroxide stress response system in M. smegmatis.A ll living organisms generate reactive oxygen species (ROS) as a by-product of metabolism. Accidental collision of O 2 with flavoproteins of metabolic pathways initially produces superoxides (O 2 Ϫ ) (1). These O 2 Ϫ are subsequently converted to other products, like hydrogen peroxide (H 2 O 2 ), hydroxyl radicals (OH), and others, by enzymatic and nonenzymatic mechanisms (2). In addition to this inadvertent generation of O 2 Ϫ during metabolism, phagocytic cells of eukaryotes produce O 2 Ϫ through a special pathway for killing invading pathogens (3, 4). Human phagocytes also produce nitric oxide (NO), through inducible nitric oxide synthase, and this reacts with O 2 Ϫ to generate highly toxic peroxynitrites (OOONO) (5). Production of this compound by phagocytes is considered an important and effective way of combating the invading pathogens (6). Excess ROS can oxidize macromolecules, like nucleic acids, protein, carbohydrates, and lipids, which affect cellular processes of all types (7).Two families of enzymes are associated with detoxification of alkyl and organic hydroperoxides in bacteria. The first one belongs to the peroxiredoxin family and is called alkyl hydroperoxide reductase (AhpC) (8, 9). It exis...

Summary Bacterial antioxidants play a critical role in the detoxification of endogenously and host derived oxidative radicals during host-pathogen interactions. Recently, the osmotically induced bacterial protein C (OsmC) is included in the antioxidant category of enzymes as it shows structural and functional relationships with organic hydroperoxide reductase (Ohr) enzyme. A copy of the gene encoding OsmC is conserved across mycobacterial species, including Mycobacterium tuberculosis (Rv2923c) and M. smegmatis (MSMEG2421), but its role in protecting these species against oxidative stress is unknown. To determine the role of OsmC in mycobacterial oxidative stress, we overexpressed and purified OsmCs of M. tuberculosis and M. smegmatis and assessed their ability to reduce peroxide substrates like hydrogen peroxide (H2O2), cumene hydroperoxide (CHP) and t-butyl hydroperoxide (t-BHP) in Ferrous Ion Oxidation in Xylenol (FOX) assay. This revealed that OsmCs from both species were capable of reducing both inorganic (H2O2) and organic (CHP and t-BHP) peroxides. Further, an M. smegmatis mutant (MSΔosmC) deficient in OsmC exhibited reduced reduction of CHP and t-BHP than the parental wild type strain, indicating that OsmC protein contributes significantly for the total peroxide reductase activity of mycobacteria. The MSΔosmC strain was also sensitive to organic hydroperoxides, which could be reversed by complementing with a plasmid borne osmC. Plasmid borne osmC also increased the resistance of M. smegmatis wild type strain to isoniazid (INH) but at a relatively lower level than ahpC, an organic hydroperoxide reductase. These results suggest that OsmC plays an important role in peroxide metabolism and protecting mycobacteria against oxidative stress.

Mycoplasma genitalium is an important sexually transmitted pathogen that affects both men and women. In genital-mucosal tissues, it initiates colonization of epithelial cells by attaching itself to host cells via several identified bacterial ligands and host cell surface receptors. We have previously shown that a mutant form of M. genitalium lacking methionine sulfoxide reductase A (MsrA), an antioxidant enzyme which converts oxidized methionine (Met(O)) into methionine (Met), shows decreased viability in infected animals. To gain more insights into the mechanisms by which MsrA controls M. genitalium virulence, we compared the wild-type M. genitalium strain (G37) with an msrA mutant (MS5) strain for their ability to interact with target cervical epithelial cell lines (HeLa and C33A) and THP-1 monocytic cells. Infection of epithelial cell lines with both strains revealed that MS5 was less cytotoxic to HeLa and C33A cell lines than the G37 strain. Also, the MS5 strain was more susceptible to phagocytosis by THP-1 cells than wild type strain (G37). Further, MS5 was less able to induce aggregation and differentiation in THP-1 cells than the wild type strain, as determined by carboxyfluorescein diacetate succinimidyl ester (CFSE) labeling of the cells, followed by counting of cells attached to the culture dish using image analysis. Finally, MS5 was observed to induce less proinflammatory cytokine TNF-α by THP-1 cells than wild type G37 strain. These results indicate that MsrA affects the virulence properties of M. genitalium by modulating its interaction with host cells.

Background: Patient delay can contribute to a poor outcome in the management of head and neck cancers (HNC). The main objective of the present study was to investigate the factors associated with patient delay in our population. Materials and Methods: Patients with cancers of the head and neck attending a regional cancer center of North East India were consecutively interviewed during the period from June 2014 to November 2014. The participation of patients was voluntary. The questionnaire included information on age, gender, residential status, educational qualification, monthly family income, any family history of cancer, and history of prior awareness on cancer from television (TV) program and awareness program. Results: Of 311 (n) patients, with an age range of 14-88 years (mean 55.4 years), 81.7% were males and 18.3% females (M:F=4.4). The overall median delay was 90 days (range=7 days-365 days), in illiterate patients the median delay was 90 days and 60 days in literate patients (P=0.002), the median delay in patients who had watched cancer awareness program on TV was 60 days and in patients who were unaware about cancer information from TV program had a median delay of 90 days (p=0.00021) and delay of <10 weeks was seen in 139 (44.6%) patients, a delay of 10-20 weeks in 98 (31.5%) patients, and a delay of 20-30 weeks in 63 (20.2%) patients. Conclusions: Education and awareness had a significant impact in reduction of median patient delay in our HNC cases.