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The A. sendaiensis PA2 is a polyextremophile bacterium. In this study, we analyze the A. sendaiensis PA2 genome. The genome was assembled and annotated. The A. sendaiensis PA2 genome structure consists of a 2,956,928 bp long chromosome and 62.77% of G + C content. 3056 CDSs were predicted, and 2921 genes were assigned to a putative function. The ANIm and ANIb value resulted in 97.17% and 96.65%, the DDH value was 75.5%, and the value of TETRA (Z-score) was 0.98. Comparative genomic analyses indicated that three systems are enriched in A. sendaiensis PA2. This strain has phenotypic changes in cell wall during batch culture at 65 °C, pH 5.0 and without carbon and nitrogen source. The presence of unique genes of cell wall and sporulation subsystem could be related to the adaptation of A. sendaiensis PA2 to hostile conditions. [ABSTRACT FROM AUTHOR]

Abstract Genome‐wide screens have discovered a large set of essential genes in the opportunistic human pathogen Streptococcus pneumoniae. However, the functions of many essential genes are still unknown, hampering vaccine development and drug discovery. Based on results from transposon sequencing (Tn‐seq), we refined the list of essential genes in S. pneumoniae serotype 2 strain D39. Next, we created a knockdown library targeting 348 potentially essential genes by CRISPR interference (CRISPRi) and show a growth phenotype for 254 of them (73%). Using high‐content microscopy screening, we searched for essential genes of unknown function with clear phenotypes in cell morphology upon CRISPRi‐based depletion. We show that SPD_1416 and SPD_1417 (renamed to MurT and GatD, respectively) are essential for peptidoglycan synthesis, and that SPD_1198 and SPD_1197 (renamed to TarP and TarQ, respectively) are responsible for the polymerization of teichoic acid (TA) precursors. This knowledge enabled us to reconstruct the unique pneumococcal TA biosynthetic pathway. CRISPRi was also employed to unravel the role of the essential Clp‐proteolytic system in regulation of competence development, and we show that ClpX is the essential ATPase responsible for ClpP‐dependent repression of competence. The CRISPRi library provides a valuable tool for characterization of pneumococcal genes and pathways and revealed several promising antibiotic targets.

Proteins containing the NIF3 domain are highly conserved and are found in bacteria, eukaryotes, and archaea. YbgI is an Escherichia coli protein whose gene is conserved among bacteria. The structure of YbgI is known; however, the function of this protein in cells remains obscure. Our studies of E. coli cells with deleted ybgI gene suggest that YbgI is involved in formation of the bacterial cell wall. [ABSTRACT FROM AUTHOR]

Genome-wide screens have discovered a large set of essential genes in the opportunistic human pathogen Streptococcus pneumoniae. However, the functions of many essential genes are still unknown, hampering vaccine development and drug discovery. Based on results from transposon sequencing (Tn-seq), we refined the list of essential genes in S. pneumoniae serotype 2 strain D39. Next, we created a knockdown library targeting 348 potentially essential genes by CRISPR interference (CRISPRi) and show a growth phenotype for 254 of them (73%). Using high-content microscopy screening, we searched for essential genes of unknown function with clear phenotypes in cell morphology upon CRISPRi-based depletion. We show that SPD_1416 and SPD_1417 (renamed to MurT and GatD, respectively) are essential for peptidoglycan synthesis, and that SPD_1198 and SPD_1197 (renamed to TarP and TarQ, respectively) are responsible for the polymerization of teichoic acid (TA) precursors. This knowledge enabled us to reconstruct the unique pneumococcal TA biosynthetic pathway. CRISPRi was also employed to unravel the role of the essential Clp-proteolytic system in regulation of competence development, and we show that ClpX is the essential ATPase responsible for ClpP-dependent repression of competence. The CRISPRi library provides a valuable tool for characterization of pneumococcal genes and pathways and revealed several promising antibiotic targets. [ABSTRACT FROM AUTHOR]

As the response to unfavorable growth conditions, nonsporulating mycobacteria transform into the dormant state with the concomitant formation of the specialized dormant forms characterized by low metabolic activity and resistance to antibiotics. Such dormant cells can be reactivated under the influence of several factors including proteins of Rpf (Resuscitation promoting factor) family, which possess peptidoglycan hydrolase activity and were considered to belong to the group of the autocrine growth factors of the bacteria. Remarkable interest toward Rpf family is determined by its par-ticipation in resuscitation of the dormant forms of Mycobacterium tuberculosis, what in turn is the key element in resuscitation of the latent tuberculosis - an infectious disease that affects one third of the World's population. Experiments with Rpf mutant forms and with strains deleted in these proteins revealed a relationship between the enzymatic activity of this protein and its ability to resuscitate mycobacteria both in vitro and in vivo. This review discusses possible mechanisms of Rpf action including those related to possible participation of the products of mycobacterial Rpf-mediated cell wall hydrolysis (muropeptides) as signaling molecules. The unique ability of Rpf proteins to resuscitate the dormant forms of mycobacteria and to stimulate their proliferation would allow these proteins to occupy their niche in medicine-in diagnostics and in creation of antituberculosis subunit vaccines. [ABSTRACT FROM AUTHOR]

Rod-like bacteria maintain their cylindrical shapes with remarkable precision during growth. However, they are also capable to adapt their shapes to external forces and constraints, for example by growing into narrow or curved confinements. Despite being one of the simplest morphologies, we are still far from a full understanding of how shape is robustly regulated, and how bacteria obtain their near-perfect cylindrical shapes with excellent precision. However, recent experimental and theoretical findings suggest that cell-wall geometry and mechanical stress play important roles in regulating cell shape in rod-like bacteria. We review our current understanding of the cell wall architecture and the growth dynamics, and discuss possible candidates for regulatory cues of shape regulation in the absence or presence of external constraints. Finally, we suggest further future experimental and theoretical directions which may help to shed light on this fundamental problem. [ABSTRACT FROM AUTHOR]

Patrinia scabiosaefolia (PS) clinically had been prescribed in oriental ethnopharmacology, but its exact mechanism is unknown. In this study, we analyzed molecular perspective of antibiotic effects of PS. Escherichia coli O157:H7 was changed in four main mechanisms; (1) Cell wall biosynthesis decreased, (2) DNA transcription consists of folic acid metabolism interruption, (3) Protein synthesis and folding declined, (4) Bacterial cell motility increased. With these results, antibiotic effect was demonstrated at a molecular perspective of PS. Safe and effective drugs for antibiotic resistance will promote with the multi-target herbal medicine. [ABSTRACT FROM AUTHOR]

Fourteen-membered-ring macrolides have an antiinflammatory effect, in addition to their antibacterial effect, and are widely used at low dosages for long-term therapy for chronic inflammatory disease such as diffuse pan-bronchiolitis and chronic sinusitis. A macrolide-resistant coagulase-negative staphylococcal strain was obtained from the maxillary sinus of a patient with chronic sinusitis, who failed long-term macrolide therapy. The isolated strain was characterized as Staphylococcus capitis and had an MIC for erythromycin greater than 128 μg/ml. Morphological observation demonstrated that this macrolide-resistant S. capitis strain had a thicker cell wall than macrolide-sensitive S. capitis strains. Moreover, the strain was not carrying any other than the four genes that are known mainly to encode for macrolide resistance in S. aureus. Therefore, the strain had an unknown macrolide-resistance mechanism that might be related to cell wall thickening. [ABSTRACT FROM AUTHOR]

The cell wall of Gram-negative bacteria is essential for the integrity of the bacterial cell but also imposes a physical barrier to trans-envelope transport processes in which DNA and/or proteins are taken up or secreted by complex protein assemblies. The presence of genes encoding lytic transglycosylases in macromolecular transport systems (bacteriophage entry, type II secretion and type IV pilus synthesis, type III secretion, type IV secretion) suggests an important role for these specialised cell-wall-degrading enzymes. Such enzymes are capable of locally enlarging gaps in the peptidoglycan meshwork to allow the efficient assembly and anchoring of supramolecular transport complexes in the cell envelope. In this review, current knowledge on the role and distribution of these specialised murein-degrading enzymes in diverse macromolecular transport systems is summarised and discussed. [ABSTRACT FROM AUTHOR]

The innate immune system recognizes microorganisms through a series of pattern recognition receptors that are highly conserved in evolution. Peptidoglycan (PGN) is a unique and essential component of the cell wall of virtually all bacteria and is not present in eukaryotes, and thus is an excellent target for the innate immune system. Indeed, higher eukaryotes, including mammals, have several PGN recognition molecules, including CD14, Toll-like receptor 2, a family of peptidoglycan recognition proteins, Nod1 and Nod2, and PGN-lytic enzymes (lysozyme and amidases). These molecules induce host responses to microorganisms or have direct antimicrobial effects. [ABSTRACT FROM AUTHOR]

Various aspects of surface properties of the S-layer-carrying Bacillus stearothermophilus PV72 and of an S-layer-deficient mutant (strain PV72/T5) have been tested by adsorption assays on solid surfaces, electrostatic interaction chromatography and hydrophobic interaction chromatography. The adsorption assays have shown that cell adhesion of the S-layer-carrying strain was less influenced by environmental changes than it was with the S-layer-deficient mutant. Electrostatic interaction chromatography indicated that both strains have positively and negatively charged groups exposed on the cell surface but the S-layer-carrying strain reveals more positively charged groups than does the S-layer-deficient mutant. Hydrophobic interaction chromatography showed that both strains have a hydrophilic surface but that the hydrophilic properties are more pronounced with the strain lacking an S-layer. [ABSTRACT FROM AUTHOR]

The location of the covalently attached carbohydrate residue of the S-layer glycoprotein from Clostridium thermohydrosulfuricum L111-69 was determined by electron microscopical procedures after converting the hydroxyl groups of the carbohydrate chains into carboxyl groups by succinylation. The introduction of carboxyl groups was examined by labelling with polycationized ferritin (PCF; a net positively charged topographical marker for electron microscopy). Cyanogen bromide was used for activating vicinal hydroxyl groups of the carbohydrate chains which could then react with amino groups of amino carbonic acids or ferritin. The amount of covalently bound ferritin was determined by freeze-etching and UV-measurement. Both, succinylation experiments and covalent attachment of ferritin confirmed that at least a considerable portion of the carbohydrate residue must be located on the S-layer surface. [ABSTRACT FROM AUTHOR]

Labelling experiments using a positively charged topographical marker for electron microscopy, polycationized ferritin, showed that the S-layers of two closely related clostridia Clostridium thermohydrosulfuricum L111-69 and C. thermosaccharolyticum D120-70 do not exhibit a net negative charge, as usually observed for bacterial cell surfaces. Chemical modification of reactive sites confirmed that amino and carboxyl groups are exposed on the S-layer surface of both strains. Amino-specific, bifunctional agents crosslinked both S-layer lattices. Studies with carbodiimides revealed that only the S-layer surface of C. thermohydrosulfuricum L111-69 had amino and carboxyl groups closely enough aligned to permit electrostatic interactions between the constituent protomers. The regular structure of this S-layer lattice was lost upon converting the carboxyl groups into neutral groups by amidation. Disintegration of both S-layer lattices occurred upon N-acetylation or N-succinylation of the free amino groups. Adhesion experiments showed that in neutral and weakly alkaline environment whole cells of C. thermosaccharolyticum D120-70 exhibited a stronger tendency to bind to charged surfaces than whole cells of C. thermohydrosulfuricum L111-69, but showed a lower tendency to bind to hydrophobic materials. [ABSTRACT FROM AUTHOR]

The structures, amino acid- and neutral sugar compositions of the crystalline surface layers (S-layers) of four selected strains each of Bacillus stearothermophilus and Desulfotomaculum nigrificans were compared. Among the four strains of each species a remarkable diversity in the molecular weights of the S-layer subunits and in the geometry and constants of the S-layer lattices was apparent. The crystalline arrays included hexagonal (p6), square (p4) and oblique (p2) lattices. In vitro self-assembly of isolated S-layer subunits (or S-layer fragments) led to the formation of flat sheets or open-ended cylindrical assembly products. The amino acid composition of the S-layers exhibited great similarities and was predominantly acidic. With the exception of the S-layers of two strains of B. stearothermophilus (where only traces of neutral sugars could be detected), all other S-layer proteins seemed to be glycosylated. Among these strains significant differences in the amount and composition of the glycan portions were found. Based on this diversity interesting questions may be asked about the biological significance of the carbohydrate units of glycoproteins in prokaryotic organisms. [ABSTRACT FROM AUTHOR]

Determination of the primary structure of the peptidoglycan of 15 strains of Oerskovia showed that three different peptidoglycan types occur. Oerskovia xanthineolytica strains contain the l-Lys- d-Ser-β- d-Asp type, whereas Oerskovia turbata strains show the new peptidoglycan types l-Lys- l-Thr-β- d-Asp or l-Lys- l-Thr-ψ- d-Glu, respectively. Research on the cytochromes of Oerskovia revealed the presence of a, b and c types. O. turbata can be clearly distinguished from O. xanthineolytica by the occurrence of cytochrome a in cells, isolated from the stationary phase. The following conclusions were made: O. turbata and O. xanthineolytica can be clearly separated on the basis of different peptidoglycan types and cytochrome patterns. This distinction is in perfect correlation with the classical separation method of O. turbata and O. xanthineolytica on the basis of xanthine degradation. l-Lys- d-Ser-β- d-Asp peptidoglycan type does not only occur in O. xanthineolytica but also in some coryneform bacteria such as Corynebacterium manihot (Fiedler et al. 1970), Cellulomonas cartae (Stackebrandt et al. 1978; Stackebrandt and Kandler 1980), Brevibacterium fermentans and Nocardia cellulans. [ABSTRACT FROM AUTHOR]

We induced granulomatous enterocolitis in rabbits by injecting them with muramyl dipeptide (MDP), a subunit of the peptidoglycan polymers that endow the bacterial cell wall with structural rigidity, emulsified with Freund's incomplete adjuvant (FIA). Injections of 0.1 ml of a water-in-oil emulsion of MDP and FIA were given submucosally at six sites in the rectum and colon, 10 cm proximal to the anus, using a flexible endoscope. Four rabbits each were sacrificed 1, 2, and 4 weeks after a single injection of the emulsion. Another 4 rabbits each were injected six times at 1- and 2-week intervals, and were sacrificed 1 and 2 weeks after the last injection of the emulsion, respectively. In all 20 rabbits, injected with the MDP emulsion, histological findings of the colon consisted of cellular infiltrations of plasma cells and lymphocytes, granulomatous lesions, and granulomas, although the findings differed in degree. Cellular infiltration in hyperplastic villi and denuded epithelia of the small intestine were seen in 2 of 8 rabbits repeated that received MDP emulsion injections. The histological changes in this animal model may be useful for studying the pathogenesis of inflammatory bowel disease in humans. [ABSTRACT FROM AUTHOR]

The qualitative and quantitative composition of purified cell walls of Halococcus morrhuae CCM 859 was determined. Glucose, mannose, galactose; glucuronic and galacturonic acids; glucosamine, galactosamine, gulosaminuronic acid; acetate, glycine and sulfate are found as major constituents. The amino sugars are N-acetylated. It was not possible to fractionate the cell wall in chemically different polymers. Evidence is presented that the major cell wall polymer of this strain is a complex heteroglycan which seems, like the peptidoglycan of most bacteria, to be responsible for the rigidity and stability of the cell wall. In addition it could be proved that this heteroglycan is sulfated and therefore differs considerably from previously described bacterial cell wall polymers. [ABSTRACT FROM AUTHOR]