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<jats:title>Abstract</jats:title><jats:p>The growth of mycobacterial cells requires successful coordination between elongation and septation. However, it is not clear which factors mediate this coordination. Here, we studied the function and post‐translational modification of an essential division factor, SepIVA, in <jats:italic>Mycobacterium smegmatis</jats:italic>. We find that SepIVA is arginine methylated, and that alteration of its methylation sites affects both septation and polar elongation of <jats:italic>Msmeg.</jats:italic> Furthermore, we show that SepIVA regulates the localization of MurG and that this regulation may impact polar elongation. Finally, we map SepIVA's two regulatory functions to different ends of the protein: the N‐terminus regulates elongation while the C‐terminus regulates division. These results establish SepIVA as a regulator of both elongation and division and characterize a physiological role for protein arginine methylation sites for the first time in mycobacteria.</jats:p>

<jats:title>Abstract</jats:title><jats:p>Bacteria deal with an unpredictable, and often hostile, environment by being unpredictable themselves. This article will link some contributions made by variable DNA topology and nucleoid‐associated proteins to the generation of stochasticity in bacterial gene expression and describe how the associated mechanistic insights can elucidate the means by which diversity in antibody and neuronal cell development might be produced in humans and other higher organisms. The focus here will not be on mutation; instead, the article will address epigenetic effects on gene expression brought about by the modulation of topoisomerase activity in both prokaryotes and eukaryotes.</jats:p>

<jats:title>Abstract</jats:title><jats:p>The infection course of <jats:italic>Mycobacterium tuberculosis</jats:italic> is highly dynamic and comprises sequential stages that require damaging and crossing of several membranes to enable the translocation of the bacteria into the cytosol or their escape from the host. Many important breakthroughs such as the restriction of mycobacteria by the autophagy pathway and the recruitment of sophisticated host repair machineries to the <jats:italic>Mycobacterium</jats:italic>‐containing vacuole have been gained in the <jats:italic>Dictyostelium discoideum</jats:italic>/<jats:italic>M. marinum</jats:italic> system. Despite the availability of well‐established light and advanced electron microscopy techniques in this system, a correlative approach integrating both methods with near‐native ultrastructural preservation is currently lacking. This is most likely due to the low ability of <jats:italic>D. discoideum</jats:italic> to adhere to surfaces, which results in cell loss even after fixation. To address this problem, we improved the adhesion of cells and developed a straightforward and convenient workflow for 3D‐correlative light and electron microscopy. This approach includes high‐pressure freezing, which is an excellent technique for preserving membranes. Thus, our method allows to monitor the ultrastructural aspects of vacuole escape which is of central importance for the survival and dissemination of bacterial pathogens.</jats:p>

<jats:title>Abstract</jats:title><jats:p><jats:italic>Salmonella</jats:italic> Typhi, the invasive serovar of <jats:italic>S. enterica</jats:italic> subspecies <jats:italic>enterica</jats:italic>, causes typhoid fever in healthy human hosts. The emergence of antibiotic‐resistant strains has consistently challenged the successful treatment of typhoid fever with conventional antibiotics. Antimicrobial resistance (AMR) in <jats:italic>Salmonella</jats:italic> is acquired either by mutations in the genomic DNA or by acquiring extrachromosomal DNA via horizontal gene transfer. In addition, <jats:italic>Salmonella</jats:italic> can form a subpopulation of antibiotic persistent (AP) cells that can survive at high concentrations of antibiotics. These have reduced the effectiveness of the first and second lines of antibiotics used to treat <jats:italic>Salmonella</jats:italic> infection. The recurrent and chronic carriage of <jats:italic>S.</jats:italic> Typhi in human hosts further complicates the treatment process, as a remarkable shift in the immune response from pro‐inflammatory Th1 to anti‐inflammatory Th2 is observed. Recent studies have also highlighted the overlap between AP, persistent infection (PI) and AMR. These incidents have revealed several areas of research. In this review, we have put forward a timeline for the evolution of antibiotic resistance in <jats:italic>Salmonella</jats:italic> and discussed the different mechanisms of the same availed by the pathogen at the genotypic and phenotypic levels. Further, we have presented a detailed discussion on <jats:italic>Salmonella</jats:italic> antibiotic persistence (AP), PI, the host and bacterial virulence factors that can influence PI, and how both AP and PI can lead to AMR.</jats:p>

<jats:title>Abstract</jats:title><jats:p>Free‐living organisms frequently encounter unfavorable abiotic environmental factors. Those who adapt and cope with sudden changes in the external environment survive. Desiccation is one of the most common and frequently encountered stresses in nature. On the contrary, ionizing radiations are limited to high local concentrations of naturally occurring radioactive materials and related anthropogenic activities. Yet, resistance to high doses of ionizing radiation is evident across the tree of life. The evolution of desiccation resistance has been linked to the evolution of ionizing radiation resistance, although, evidence to support the idea that the evolution of desiccation tolerance is a necessary precursor to ionizing radiation resistance is lacking. Moreover, the presence of radioresistance in hyperthermophiles suggests multiple paths lead to radiation resistance. In this minireview, we focus on the molecular aspects of damage dynamics and damage response pathways comprising protective and restorative functions with a definitive survival advantage, to explore the serendipitous genesis of ionizing radiation resistance.</jats:p>

<jats:title>Abstract</jats:title><jats:p><jats:italic>Brucella abortus</jats:italic> is a facultative, intracellular, zoonotic pathogen that resides inside macrophages during infection. This is a specialized niche where <jats:italic>B. abortus</jats:italic> encounters various stresses as it navigates through the macrophage. In order to survive this harsh environment, <jats:italic>B. abortus</jats:italic> utilizes post‐transcriptional regulation of gene expression through the use of small regulatory RNAs (sRNAs). Here, we characterize a <jats:italic>Brucella</jats:italic> sRNAs called MavR (for <jats:italic>M</jats:italic>urF‐ <jats:italic>a</jats:italic>nd <jats:italic>v</jats:italic>irulence‐regulating s<jats:italic>R</jats:italic>NA), and we demonstrate that MavR is required for the full virulence of <jats:italic>B. abortus</jats:italic> in macrophages and in a mouse model of chronic infection. Transcriptomic and proteomic studies revealed that a major regulatory target of MavR is MurF. MurF is an essential protein that catalyzes the final cytoplasmic step in peptidoglycan (PG) synthesis; however, we did not detect any differences in the amount or chemical composition of PG in the Δ<jats:italic>mavR</jats:italic> mutant. A 6‐nucleotide regulatory seed region within MavR was identified, and mutation of this seed region resulted in dysregulation of MurF production, as well as significant attenuation of infection in a mouse model. Overall, the present study underscores the importance of sRNA regulation in the physiology and virulence of <jats:italic>Brucella</jats:italic>.</jats:p>

<jats:title>Abstract</jats:title><jats:p>The nosocomial bacterium <jats:italic>Acinetobacter baumannii</jats:italic> is protected from antibiotic treatment by acquiring antibiotic resistances and by forming biofilms. Cell attachment, one of the first steps in biofilm formation, is normally induced by environmental metabolites. We hypothesized that vanillic acid (VA), the oxidized form of vanillin and a widely available metabolite, may play a role in <jats:italic>A. baumannii</jats:italic> cell attachment. We first discovered that <jats:italic>A. baumannii</jats:italic> actively breaks down VA through the evolutionarily conserved <jats:italic>vanABKP</jats:italic> genes. These genes are under the control of the repressor VanR, which we show binds directly to VanR binding sites within the <jats:italic>vanABKP</jats:italic> genes bidirectional promoter. VA in turn counteracts VanR inhibition. We identified a VanR binding site and searched for it throughout the genome, especially in pili encoding promoter genes. We found a VanR binding site in the pilus encoding <jats:italic>csu</jats:italic> operon promoter and showed that VanR binds specifically to it. As expected, a strain lacking VanR overproduces Csu pili and makes robust biofilms. Our study uncovers the role that VA plays in facilitating the attachment of <jats:italic>A. baumannii</jats:italic> cells to surfaces, a crucial step in biofilm formation. These findings provide valuable insights into a previously obscure catabolic pathway with significant clinical implications.</jats:p>

<jats:title>Abstract</jats:title><jats:p>Pathogenic <jats:italic>Rhodococcus equi</jats:italic> release the virulence‐associated protein A (VapA) within macrophage phagosomes. VapA permeabilizes phagosome and lysosome membranes and reduces acidification of both compartments. Using biophysical techniques, we found that VapA interacts with model membranes in four steps: (i) binding, change of mechanical properties, (ii) formation of specific membrane domains, (iii) permeabilization within the domains, and (iv) pH‐specific transformation of domains. Biosensor data revealed that VapA binds to membranes in one step at pH 6.5 and in two steps at pH 4.5 and decreases membrane fluidity. The integration of VapA into lipid monolayers was only significant at lateral pressures &lt;20 mN m<jats:sup>−1</jats:sup> indicating preferential incorporation into membrane regions with reduced integrity. Atomic force microscopy of lipid mono‐ and bilayers showed that VapA increased the surface heterogeneity of liquid disordered domains. Furthermore, VapA led to the formation of a new microstructured domain type and, at pH 4.5, to the formation of 5 nm high domains. VapA binding, its integration and lipid domain formation depended on lipid composition, pH, protein concentration and lateral membrane pressure. VapA‐mediated permeabilization is clearly distinct from that caused by classical microbial pore formers and is a key contribution to the multiplication of <jats:italic>Rhodococcus equi</jats:italic> in phagosomes.</jats:p>

<jats:title>Abstract</jats:title><jats:p>The diarrheal disease cholera is caused by the versatile and responsive bacterium <jats:italic>Vibrio cholerae</jats:italic>, which is capable of adapting to environmental changes. Among others, the alternative sigma factor RpoS activates response pathways, including regulation of motility‐ and chemotaxis‐related genes under nutrient‐poor conditions in <jats:italic>V. cholerae</jats:italic>. Although RpoS has been well characterised, links between RpoS and other regulatory networks remain unclear. In this study, we identified the ArcAB two‐component system to control <jats:italic>rpoS</jats:italic> transcription and RpoS protein stability in <jats:italic>V. cholerae</jats:italic>. In a manner similar to that seen in <jats:italic>Escherichia coli</jats:italic>, the ArcB kinase not only activates the response regulator ArcA but also RssB, the anti‐sigma factor of RpoS. Our results demonstrated that, in <jats:italic>V. cholerae</jats:italic>, RssB is phosphorylated by ArcB, which subsequently activates RpoS proteolysis. Furthermore, ArcA acts as a repressor of <jats:italic>rpoS</jats:italic> transcription. Additionally, we determined that the cysteine residue at position 180 of ArcB is crucial for signal recognition and activity. Thus, our findings provide evidence linking RpoS response to the anoxic redox control system ArcAB in <jats:italic>V. cholerae</jats:italic>.</jats:p>

<jats:title>Abstract</jats:title><jats:p>In the human pathogen <jats:italic>Staphylococcus aureus</jats:italic>, branched‐chain fatty acids (BCFAs) are the most abundant fatty acids in membrane phospholipids. Strains deficient for BCFAs synthesis experience auxotrophy in laboratory culture and attenuated virulence during infection. Furthermore, the membrane of <jats:italic>S. aureus</jats:italic> is among the main targets for antibiotic therapy. Therefore, determining the mechanisms involved in BCFAs synthesis is critical to manage <jats:italic>S. aureus</jats:italic> infections. Here, we report that the overexpression of SAUSA300_2542 (annotated to encode an acyl‐CoA synthetase) restores BCFAs synthesis in strains lacking the canonical biosynthetic pathway catalyzed by the branched‐chain α‐keto acid dehydrogenase (BKDH) complex. We demonstrate that the acyl‐CoA synthetase activity of MbcS activates branched‐chain carboxylic acids (BCCAs), and is required by <jats:italic>S. aureus</jats:italic> to utilize the isoleucine derivative 2‐methylbutyraldehyde to restore BCFAs synthesis in <jats:italic>S. aureus</jats:italic>. Based on the ability of some staphylococci to convert branched‐chain aldehydes into their respective BCCAs and our findings demonstrating that branched‐chain aldehydes are in fact BCFAs precursors, we propose that MbcS promotes the scavenging of exogenous BCCAs and mediates BCFA synthesis via a de novo alternative pathway.</jats:p>