Effects of povidone-iodine composite on the elimination of bacterial biofilm.

BACKGROUND: Povidone-iodine (PVP-I) is well known as an antiseptic and exhibits extensive activity against various pathogens. However, due to its uniquely unpleasant nature, it cannot be used locally to deactivate various sinonasal pathogens. Therefore, we developed a PVP-I composite that blocks the unpleasant odor of PVP-I for use as a local antiseptic in the sinonasal cavity and evaluated its effect on bacterial biofilm's formation and elimination in in vivo and in vitro models. METHODS: MTT, lactate dehydrogenase, and live/dead staining assay were performed to examine the cellular toxicity of PVP-I composites on the primary human nasal epithelial and RPMI 2650 cells. Crystal violet assay was performed to quantify bacterial biofilm after treating with various agents, including PVP-I and antibiotics. Hematoxylin-and-eosin staining, live/dead staining assay, and scanning electron microscopy were conducted to evaluate the effect of PVP-I on biofilm formation in a mice biofilm model. RESULTS: It was observed that the PVP-I composite did not have any significant toxic effect on the nasal epithelial cells. Furthermore, the PVP-I composite effectively inhibited the formation of bacterial biomass within a dose-dependent manner after 48 hours of incubation with Pseudomonas aeruginosa and Staphylococcus aureus. In mice, it effectively eliminated biofilm from the mucosa of the nasal cavity and maxillary sinus at the tested concentrations. CONCLUSION: The results of this study indicate that the PVP-I composite is a promising compound that could be used locally to prevent the formation of biofilms and to eliminate them from the sinonasal cavity.

Novel Cytoplasmic Bacteriocin Compounds Derived from Staphylococcus epidermidis Selectively Kill Staphylococcus aureus, Including Methicillin-Resistant Staphylococcus aureus (MRSA).

Staphylococcus aureus (S. aureus) is one of the well-known agents causing atopic dermatitis (AD) in susceptible individuals, and Staphylococcus epidermidis (S. epidermidis) produces class I thermostable bacteriocins that can selectively kill S. aureus, suggesting protective roles against AD. There is a large need for developing precise therapies only to target S. aureus and not to harm the beneficial microbiome. On the agar well diffusion assay, live planktonic S. epidermidis showed clear zones of inhibition of S. aureus growth, but heat-killed cells and cell-free supernatants did not show this. These results would lead us to hypothesize that cytoplasmic bacteriocin from S. epidermidis will be a promising agent to inhibit S. aureus growth. Therefore, we have extracted a novel thermolabile cytoplasmic bacteriocin from S. epidermidis using trichloroactic acid (TCA)/acetone precipitation method after cell lysis with a SDS-containing buffer. These bacteriocin selectively exhibited antimicrobial activity against S. aureus and methicillin-resistance Staphylococcus aureus (MRSA), presenting no active actions against S. epidermidis, E. coli, and Salmonella Typhimurium. The extracted cytoplasmic bacteriocin compounds revealed several diffuse bands of approximately 40-70 kDa by SDS-PAGE. These findings suggest that these cytoplasmic bacteriocin compounds would be a great potential means for S. aureus growth inhibition and topical AD treatment.

Mycobacterium tuberculosis Rv3463 induces mycobactericidal activity in macrophages by enhancing phagolysosomal fusion and exhibits therapeutic potential.

Macrophages are responsible for innate and adaptive immune response activation necessary for eliminating infections. Optimal activation of macrophages to phagocytize Mycobacterium tuberculosis is critical in anti-mycobacterial defense. Here, we identified a novel Rv3463 hypothetical protein that induces macrophage activation in Mtb culture filtrate. Recombinant Rv3463 activated mouse bone marrow-derived macrophages to induce the expression of surface molecules and secrete pro-inflammatory cytokines via the TLR2 and TLR4 pathways. Mitogen activated protein kinase, phospatidylinositol-4,5-bisphosphate 3-kinases, and the NF-κB signaling pathways are involved in Rv3463-mediated macrophage activation. Furthermore, Rv3463 induced bactericidal effects in Mtb-infected macrophages through phagosome maturation and phagolysosomal fusion enhanced by phospatidylinositol-4,5-bisphosphate 3-kinases and Ca2+ signaling pathways and exhibited therapeutic effects in a short-term Mtb-infection mouse model. Overexpression of Rv3463 in M. smegmatis caused rapid clearance of bacteria in macrophages and mice. Our study suggests that Rv3463 is a promising target for the development of post-exposure tuberculosis vaccines or adjunct immune-therapy.

Collagen-based sponge hastens wound healing via decrease of inflammatory cytokines.

The objective of this study was to compare and evaluate the efficacy of collagen-based sponge compared to commercial collagen sponge as a potent open wound-dressing material. In this study, 10 mm diameter skin incision was made on lateral side of rats. The wound was monitored regularly until day 12. Histopathology results revealed the faster re-epithelialization and lesser inflammatory cells, and also masson's trichrome staining showed that collagen fibrils were horizontal and interwoven in collagen-based sponge group. The expression of growth factors such as VEGF and TGF-ß1 was found to be upregulated in transcriptional and translational levels, suggesting the importance of collagen-based sponge as a potent wound-healing material. Furthermore, IL-6 and TNF-α in the wound tissue were significantly down-regulated in 2 and 6 days in collagen-based sponge group and anti-inflammatory cytokine IL-10 level was found to be upregulated throughout 12 days. These results cumulatively revealed that collagen-based sponge may serve as novel material for wound healing in the animal model.

Mycobacterium abscessus glycopeptidolipids inhibit macrophage apoptosis and bacterial spreading by targeting mitochondrial cyclophilin D.

Mycobacterium abscessus (MAB) is a species of nontuberculous mycobacteria (NTM) and a major causative pathogen of pulmonary diseases especially in patients with cystic fibrosis. MAB infection is notoriously difficult to treat because of its intrinsic or inducible resistance to most antibiotics. The rough (R) morphotype of MAB, lacking cell surface glycopeptidolipids (GPLs), is associated with more severe and persistent infection than the smooth (S) type; however, the mechanisms underlying the R type's virulence and the relation with GPLs remain unclear. In this study, we found that R-type MAB is much more proapoptotic than the S type, as a result of GPL-mediated inhibition of macrophage apoptosis. Polar GPLs inhibited an apoptotic response (induced by proapoptotic stimuli) by suppressing ROS production and the cytochrome c release and by preserving mitochondrial transmembrane potential. Furthermore, GPLs were found to be targeted to mitochondria and interacted with cyclophilin D; their acetylation was essential for this interaction. Finally, GPLs inhibited the intracellular growth and bacterial spreading of R-type MAB among macrophages via apoptosis inhibition. These findings suggest that GPLs limit MAB virulence by inhibiting apoptosis and the spread of bacteria and therefore provide a novel insight into the mechanism underlying virulence of MAB.

Mycobacterium avium MAV2054 protein induces macrophage apoptosis by targeting mitochondria and reduces intracellular bacterial growth.

Mycobacterium avium complex induces macrophage apoptosis. However, the M. avium components that inhibit or trigger apoptosis and their regulating mechanisms remain unclear. We recently identified the immunodominant MAV2054 protein by fractionating M. avium culture filtrate protein by multistep chromatography; this protein showed strong immuno-reactivity in M. avium complex pulmonary disease and in patients with tuberculosis. Here, we investigated the biological effects of MAV2054 on murine macrophages. Recombinant MAV2054 induced caspase-dependent macrophage apoptosis. Enhanced reactive oxygen species production and JNK activation were essential for MAV2054-mediated apoptosis and MAV2054-induced interleukin-6, tumour necrosis factor, and monocyte chemoattractant protein-1 production. MAV2054 was targeted to the mitochondrial compartment of macrophages treated with MAV2054 and infected with M. avium. Dissipation of the mitochondrial transmembrane potential (ΔΨm) and depletion of cytochrome c also occurred in MAV2054-treated macrophages. Apoptotic response, reactive oxygen species production, and ΔΨm collapse were significantly increased in bone marrow-derived macrophages infected with Mycobacterium smegmatis expressing MAV2054, compared to that in M. smegmatis control. Furthermore, MAV2054 expression suppressed intracellular growth of M. smegmatis and increased the survival rate of M. smegmatis-infected mice. Thus, MAV2054 induces apoptosis via a mitochondrial pathway in macrophages, which may be an innate cellular response to limit intracellular M. avium multiplication.

An evaluation of the use of immunoglobulin A antibody response against mycobacterial antigens for the diagnosis of Mycobacterium bovis infection in cattle.

Antibody responses are useful indicators of Mycobacterium bovis infection in cattle. Many studies have evaluated the ability of immunoglobulin G (IgG) to serodiagnose bovine tuberculosis (TB). In the current study, immunoglobulin A (IgA) and IgG responses against the MPB70 and MPB83 antigens of M. bovis, the 38 kDa phosphate-binding lipoprotein (PstS1) that is a well-known serodiagnostic M. tuberculosis antigen, and a newly identified protein, termed Rv1483c, were compared in M. bovis-infected and noninfected cattle as well as in field samples. The diagnostic utility of the IgA antibody to MPB70 and MPB83 for bovine TB was superior or comparable to that of the IgG antibody, and the sensitivity of serodiagnosis increased when the results of antigen binding by IgA and IgG were combined. The sensitivities of the IgG and IgA antibodies to the Rv1483c and PstS1 proteins were significantly lower than those to MPB70 and MPB83, and no diagnostic utility for Rv1483c was observed in field samples. Importantly, the IgA antibody reacted strongly to the MPB70 and MPB83 antigens and differentiated cattle with TB from healthy cattle in a multiantigen printed immunoassay. The results of this study support the feasibility of using IgA antibody against the MPB70 and MPB83 antigens to detect bovine TB. In addition, approaches using assays for both IgA and IgG antibodies may increase detection accuracy.

Association of p21-activated kinase-1 activity with aggressive tumor behavior and poor prognosis of head and neck cancer.

BACKGROUND: This study investigated the role of p21-activated kinase (PAK)-1 in progression of head and neck squamous cell carcinoma (HNSCC). METHODS: We examined PAK isoforms and explored whether PAK activation enhanced in vitro invasion of the HNSCC cell line. We analyzed the relationship between PAK1 expression and various clinicopathological features and investigated the effect of PAK1 overexpression on survival in 119 patients with HNSCC. RESULTS: PAK1 and PAK2 are predominantly expressed in HNSCC cells and patient tissues. Particularly, PAK1 makes the dominant contribution to increase in cell migration and invasion. There was a statistically significant correlation between PAK1 overexpression and aggressive cancer behavior. Moreover, PAK1 seemed to be a prognostic factor for overall and disease-specific survival in patients. Interestingly, enhancement of PAK1 expression was found in the invasive front of cancer. CONCLUSION: PAK1 is associated with the aggressive tumor behavior and poor prognosis of head and neck cancer.

Measurement of SUVs-maximum for normal region using VOI in PET/MRI and PET/CT.

The purpose of this research is to establish an overall data set associated with the VOI (Volume of Interest), which is available for simultaneous assessment of PET/MRI and PET/CT regardless of the use of contrast media. The participants as objects of this investigation are 26 healthy examinees in Korea, SUV (standardized-uptake-value)s-maximum evaluation for whole-body F-18 FDG (fluorodeoxyglucose) PET/MRI image using VOI of normal region has exhibited very significant difference to that for whole-body F-18 FDG PET/CT image (significant probability value (P) < 0.0001). However, there appeared high correlation between them in view of statistics (R-square (R) > 0.8). It is shown that one needs to decide SUVs-maximum for PET/MRI with the reduction of 25.0~26.4% from their evaluated value and needs to decide with the reduction of 28.8~29.4% in the same situation but with the use of contrast media. The use of SUVLBM-maximum (SUVLean Body Mass-maximum) is very advantageous in reading overall image of PET/CT and PET/MRI to medical doctors and researchers, if we consider its convenience and efficiency. We expect that this research enhances the level of the early stage accurate diagnosis with whole-body images of PET/MRI and PET/CT.

Lithium inhibits growth of intracellular Mycobacterium kansasii through enhancement of macrophage apoptosis.

Mycobacterium kansasii (Mk) is an emerging pathogen that causes a pulmonary disease similar to tuberculosis. Macrophage apoptosis contributes to innate host defense against mycobacterial infection. Recent studies have suggested that lithium significantly enhances the cytotoxic activity of death stimuli in many cell types. We examined the effect of lithium on the viability of host cells and intracellular Mk in infected macrophages. Lithium treatment resulted in a substantial reduction in the viability of intracellular Mk in macrophages. Macrophage cell death was significantly enhanced after adding lithium to Mk-infected cells but not after adding to uninfected macrophages. Lithium-enhanced cell death was due to an apoptotic response, as evidenced by augmented DNA fragmentation and caspase activation. Reactive oxygen species were essential for lithium-induced apoptosis. Intracellular scavenging by N-acetylcysteine abrogated the lithium-mediated decrease in intracellular Mk growth as well as apoptosis. These data suggest that lithium is associated with control of intracellular Mk growth through modulation of the apoptotic response in infected macrophages.

Characterization of Proinflammatory Responses and Innate Signaling Activation in Macrophages Infected with Mycobacterium scrofulaceum.

Mycobacterium scrofulaceum is an environmental and slow-growing atypical mycobacterium. Emerging evidence suggests that M. scrofulaceum infection is associated with cervical lymphadenitis in children and pulmonary or systemic infections in immunocompromised adults. However, the nature of host innate immune responses to M. scrofulaceum remains unclear. In this study, we examined the innate immune responses in murine bone marrow-derived macrophages (BMDMs) infected with different M. scrofulaceum strains including ATCC type strains and two clinically isolated strains (rough and smooth types). All three strains resulted in the production of proinflammatory cytokines in BMDMs mediated through toll-like receptor-2 and the adaptor MyD88. Activation of MAPKs (extracellular signal-regulated kinase 1/2, and p38, and c-Jun N-terminal kinase) and nuclear receptor (NF)-κB together with intracellular reactive oxygen species generation were required for the expression of proinflammatory cytokines in BMDMs. In addition, the rough morphotypes of M. scrofulaceum clinical strains induced higher levels of proinflammatory cytokines, MAPK and NF-κB activation, and ROS production than other strains. When mice were infected with different M. scrofulaceum strains, those infected with the rough strain showed the greatest hepatosplenomegaly, granulomatous lesions, and immune cell infiltration in the lungs. Notably, the bacterial load was higher in mice infected with rough colonies than in mice infected with ATCC or smooth strains. Collectively, these data indicate that rough M. scrofulaceum induces higher inflammatory responses and virulence than ATCC or smooth strains.

Mycobacterium tuberculosis Rv0652 stimulates production of tumour necrosis factor and monocytes chemoattractant protein-1 in macrophages through the Toll-like receptor 4 pathway.

Mycobacterial proteins interact with host macrophages and modulate their functions and cytokine gene expression profile. The protein Rv0652 is abundant in culture filtrates of Mycobacterium tuberculosis K-strain, which belongs to the Beijing family, compared with levels in the H37Rv and CDC1551 strains. Rv0652 induces strong antibody responses in patients with active tuberculosis. We investigated pro-inflammatory cytokine production induced by Rv0652 in murine macrophages and the roles of signalling pathways. In RAW264.7 cells and bone marrow-derived macrophages, recombinant Rv0652 induced predominantly tumour necrosis factor (TNF) and monocyte chemoattractant protein (MCP)-1 production, which was dependent on mitogen-activated protein kinases and nuclear factor-κB. Specific signalling pathway inhibitors revealed that the extracellular signal-regulated kinase 1/2 (ERK1/2), p38 and phosphatidylinositol 3-kinase (PI3K) pathways were essential for Rv0652-induced TNF production, whereas the ERK1/2 and PI3K pathways, but not the p38 pathway, were critical for MCP-1 production in macrophages. Rv0652-stimulated TNF and MCP-1 secretion by macrophages occurred in a Toll-like receptor 4-dependent and MyD88-dependent manner. In addition, Rv0652 significantly up-regulated the expression of the mannose receptor, CD80, CD86 and MHC class II molecules. These results suggest that Rv0652 can induce a protective immunity against M. tuberculosis through the macrophage activation.

Endoplasmic reticulum stress pathway-mediated apoptosis in macrophages contributes to the survival of Mycobacterium tuberculosis.

BACKGROUND: Apoptosis is thought to play a role in host defenses against intracellular pathogens, including Mycobacterium tuberculosis (Mtb), by preventing the release of intracellular components and the spread of mycobacterial infection. This study aims to investigate the role of endoplasmic reticulum (ER) stress mediated apoptosis in mycobacteria infected macrophages. METHODOLOGY/PRINCIPAL FINDINGS: Here, we demonstrate that ER stress-induced apoptosis is associated with Mtb H37Rv-induced cell death of Raw264.7 murine macrophages. We have shown that Mtb H37Rv induced apoptosis are involved in activation of caspase-12, which resides on the cytoplasmic district of the ER. Mtb infection increase levels of other ER stress indicators in a time-dependent manner. Phosphorylation of eIF2α was decreased gradually after Mtb H37Rv infection signifying that Mtb H37Rv infection may affect eIF2α phosphorylation in an attempt to survive within macrophages. Interestingly, the survival of mycobacteria in macrophages was enhanced by silencing CHOP expression. In contrast, survival rate of mycobacteria was reduced by phosphorylation of the eIF2α. Futhermore, the levels of ROS, NO or CHOP expression were significantly increased by live Mtb H37Rv compared to heat-killed Mtb H37Rv indicating that live Mtb H37Rv could induce ER stress response. CONCLUSION/SIGNIFICANCE: These findings indicate that eIF2α/CHOP pathway may influence intracellular survival of Mtb H37Rv in macrophages and only live Mtb H37Rv can induce ER stress response. The data support the ER stress pathway plays an important role in the pathogenesis and persistence of mycobacteria.

Targeting of Mycobacterium tuberculosis heparin-binding hemagglutinin to mitochondria in macrophages.

Mycobacterium tuberculosis heparin-binding hemagglutinin (HBHA), a virulence factor involved in extrapulmonary dissemination and a strong diagnostic antigen against tuberculosis, is both surface-associated and secreted. The role of HBHA in macrophages during M. tuberculosis infection, however, is less well known. Here, we show that recombinant HBHA produced by Mycobacterium smegmatis effectively induces apoptosis in murine macrophages. DNA fragmentation, nuclear condensation, caspase activation, and poly (ADP-ribose) polymerase cleavage were observed in apoptotic macrophages treated with HBHA. Enhanced reactive oxygen species (ROS) production and Bax activation were essential for HBHA-induced apoptosis, as evidenced by a restoration of the viability of macrophages pretreated with N-acetylcysteine, a potent ROS scavenger, or transfected with Bax siRNA. HBHA is targeted to the mitochondrial compartment of HBHA-treated and M. tuberculosis-infected macrophages. Dissipation of the mitochondrial transmembrane potential (ΔΨ(m)) and depletion of cytochrome c also occurred in both macrophages and isolated mitochondria treated with HBHA. Disruption of HBHA gene led to the restoration of ΔΨ(m) impairment in infected macrophages, resulting in reduced apoptosis. Taken together, our data suggest that HBHA may act as a strong pathogenic factor to cause apoptosis of professional phagocytes infected with M. tuberculosis.

Mycobacterial Heparin-binding Hemagglutinin Antigen Activates Inflammatory Responses through PI3-K/Akt, NF-κB, and MAPK Pathways.

BACKGROUND: Mycobacterium tuberculosis (Mtb) heparin binding hemagglutinin (HBHA) is an Ag known to evoke effective host immune responses during tuberculosis infection. However, the molecular basis of the host immune response to HBHA has not been fully characterized. In this study, we examined the molecular mechanisms by which HBHA can induce the expression of proinflammatory cytokines in macrophages. METHODS: HBHA-induced mRNA and protein levels of proinflammatory cytokines were determined in bone marrow-derived macrophages (BMDMs) using RT-PCR and ELISA analysis. The roles of intracellular signaling pathways for NF-κB, PI3-K/Akt, and MAPKs were investigated in macrophage proinflammatory responses after stimulation with HBHA. RESULTS: HBHA robustly activated the expression of mRNA and protein of both TNF-α and IL-6, and induced phosphorylation of NF-κB, Akt, and MAPKs in BMDMs. Both TNF-α and IL-6 production by HBHA was regulated by the NF-κB, PI3-K, and MAPK pathways. Furthermore, PI3-K activity was required for the HBHA-induced activation of ERK1/2 and p38 MAPK, but not JNK, pathways. CONCLUSION: These data suggest that mycobacterial HBHA significantly induces proinflammatory responses through crosstalk between the PI3-K and MAPK pathways in macrophages.

Endoplasmic reticulum stress response is involved in Mycobacterium tuberculosis protein ESAT-6-mediated apoptosis.

Mycobacterium tuberculosis (Mtb) infection leads to the induction of the apoptotic response, which is associated with bacilli killing. The early secreted mycobacterial antigen ESAT-6 of Mtb has been shown to induce apoptosis in human macrophages and epithelial cells. In the present study, we demonstrate that the stimulation of human epithelial A549 cells by ESAT-6 induces the endoplasmic reticulum (ER) stress response. We observed that ESAT-6 treatment increases intracellular Ca(2+) concentration, which results in ROS accumulation, and therefore induces the onset of ER stress-induced apoptosis. Our results uncover a novel apoptotic mechanism of ESAT-6 through ER stress responses in pathologic conditions such as tuberculosis.

Role of apoptosis-regulating signal kinase 1 in innate immune responses by Mycobacterium bovis bacillus Calmette-Guérin.

Mycobacterium bovis bacillus Calmette-Guérin (BCG) induces innate immune responses through Toll-like receptor (TLR) 2 and TLR4. We investigated the role of apoptosis-regulating signal kinase (ASK) 1 in reactive oxygen species (ROS)-mediated innate immune responses induced by BCG mycobacterial infection. In macrophages, M. bovis BCG stimulation resulted in rapid activation of mitogen-activated protein kinases (MAPKs), secretion of inflammatory cytokines, such as tumor necrosis factor (TNF)-alpha and interleukin (IL)-6, and ROS generation in a TLR2- and TLR4-dependent manner. M. bovis BCG-induced ROS production led to robust activation of ASK1 upstream of the c-jun-N-terminal kinase and p38 MAPK, but not extracellular-regulated kinase 1/2. Blocking ASK1 activity markedly attenuated M. bovis BCG-induced TNF-alpha and IL-6 production by macrophages. Both TLR2 and TLR4 were required for optimal activation of ASK1 in response to M. bovis BCG. Furthermore, we present evidence that TNF receptor-associated factor (TRAF) 6 activities were essential for ROS-mediated ASK1 activation by M. bovis BCG. Finally, ASK1 activities were required for effective control of intracellular mycobacterial survival. Thus, the results of this study suggest a novel role of the TLR-ROS-TRAF6-ASK1 axis in the innate immune response to mycobacteria as a signaling intermediate.

Differentially expressed genes in Mycobacterium tuberculosis H37Rv under mild acidic and hypoxic conditions.

The survival mechanism of dormant tubercle bacilli is unknown; however, accumulating evidence indicates that Mycobacterium tuberculosis can survive and persist in hypoxic and mildly acidic microenvironments. Such conditions are found in the acidic vacuoles of macrophages, which M. tuberculosis is known to target. We used DECAL (differential expression using customized amplification library) to identify the genes expressed under acidic and hypoxic conditions, following the cultivation of M. tuberculosis H37Rv at an acidic pH and/or under hypoxic or anoxic conditions in vitro. Of 960 clones analysed, 144 genes, consisting of 71 induced and 8 repressed genes, were identified by sequencing and divided into functional categories to characterize their cellular roles. In general, the genes induced under acidic and hypoxic conditions were involved in the biosynthesis of secondary metabolites (e.g. pks4), lipid metabolism, energy production (e.g. pckA) and cell wall biogenesis (e.g. Rv0696 and plcB). The combination of genes identified may explain the energy processing and energy storage of M. tuberculosis during latent infection. These findings not only enhance our understanding of the mechanism of dormancy, but they also may be useful in the design of therapeutic tools and vaccines for latent tuberculosis.

Identification of novel metronidazole-inducible genes in Mycobacterium smegmatis using a customized amplification library.

The incidence of antibiotic resistance in pathogenic bacteria is rising. Bacterial resistance may be a natural defense of organisms, or it may result from spontaneous mutations or the acquisition of exogenous resistance genes. We grew spontaneous metronidazole-resistant Mycobacterium smegmatis mutants on solid medium cultures and employed differential expression using a customized amplification library to analyze the global gene profiles of metronidazole-resistant mutants under hypoxic conditions. In total, 66 genes involved in metronidazole resistance were identified and functionally characterized using the gene role category of M. smegmatis. Overall, genes associated with cell wall synthesis, such as methyltransferase and glycosyltransferase, and genes encoding drug transporters were highly expressed. The genes may be involved in the natural drug resistance of mycobacteria by increasing mycobacterial cell wall permeability and the efflux pumps of active drugs. In addition, the genes may play a role in dormancy. The genes identified in this study may lead to a better understanding of the mechanisms of metronidazole resistance during dormancy.

In vitro and ex vivo activity of new derivatives of acetohydroxyacid synthase inhibitors against Mycobacterium tuberculosis and non-tuberculous mycobacteria.

Sulfometuron methyl (SM) is an inhibitor of acetohydroxyacid synthase (AHAS), the first common enzyme in the branched-chain amino acid biosynthetic pathway, and shows activity against Mycobacterium tuberculosis both in vitro and in vivo. To develop AHAS inhibitor derivatives with more potent activity, 100 sulfonylurea analogues were screened for antimycobacterial activity against M. tuberculosis and non-tuberculous mycobacteria (NTM), and then evaluated for intracellular activity using mouse macrophages. Three new compounds with antimycobacterial activity comparable with that of SM were identified. These compounds exhibit significant activity against intracellular M. tuberculosis (including the drug-resistant M. tuberculosis strains), and NTM Mycobacterium abscessus and Mycobacterium kansasii, respectively.