Antibiotic Resistance In Gram-positive Bacteria Research Articles

Genome-encoded ABCF factors implicated in intrinsic antibiotic resistance in Gram-positive bacteria: VmlR2, Ard1 and CplR.

Genome-encoded antibiotic resistance (ARE) ATP-binding cassette (ABC) proteins of the F subfamily (ARE-ABCFs) mediate intrinsic resistance in diverse Gram-positive bacteria. The diversity of chromosomally-encoded ARE-ABCFs is far from being fully experimentally explored. Here we characterise phylogenetically diverse genome-encoded ABCFs from Actinomycetia (Ard1 from Streptomyces capreolus, producer of the nucleoside antibiotic A201A), Bacilli (VmlR2 from soil bacterium Neobacillus vireti) and Clostridia (CplR from Clostridium perfringens, Clostridium sporogenes and Clostridioides difficile). We demonstrate that Ard1 is a narrow spectrum ARE-ABCF that specifically mediates self-resistance against nucleoside antibiotics. The single-particle cryo-EM structure of a VmlR2-ribosome complex allows us to rationalise the resistance spectrum of this ARE-ABCF that is equipped with an unusually long antibiotic resistance determinant (ARD) subdomain. We show that CplR contributes to intrinsic pleuromutilin, lincosamide and streptogramin A resistance in Clostridioides, and demonstrate that C. difficile CplR (CDIF630_02847) synergises with the transposon-encoded 23S ribosomal RNA methyltransferase Erm to grant high levels of antibiotic resistance to the C. difficile 630 clinical isolate. Finally, assisted by uORF4u,our novel tool for detection of upstream open reading frames, we dissect the translational attenuation mechanism that controls the induction of cplR expression upon an antibiotic challenge.

The transmembrane segment of TagH is required for wall teichoic acid transport under heat stress in Bacillus subtilis.

Wall teichoic acids (WTAs) are anionic polymers that are covalently linked to peptidoglycan and play important roles in cell shape determination, cell division, autolysis, pathogenesis and antibiotic resistance in Gram-positive bacteria. In Bacillus subtilis, WTA is synthesized in the cytoplasm and translocated by an ABC transporter, TagGH. In this study, we found that the transmembrane segment of TagH is required for WTA transport under high temperatures. Cells expressing TagH302-FL (a construct fused to the 6×FLAG tag after the transmembrane segment, which lacks the C-terminal extracellular domain) grew normally at high temperatures, similar to those expressing the full-length TagH-FL fusion. In contrast, cells expressing TagH275-FL, which lacks both the transmembrane segment and the extracellular domain, exhibited a temperature-sensitive phenotype at temperatures above 49 °C and a growth defect at 50 °C. Interestingly, this growth defect was dissolved by an additional incubation at 37 °C. A similar temperature-sensitive phenotype was observed in cells expressing an N-terminal 6×FLAG tag fusion of TagH275. Immunofluorescence microscopy (IFM) indicated that TagG and TagH are localized on the cytoplasmic membrane in a patch-like manner. In addition, the C-terminal-truncated forms, TagH275-FL and TagH302-FL, were localized in similar patch-like patterns at 37 °C; only foci for TagH275-FL were remarkably reduced at high temperatures. Moreover, cell surface decoration with WTA was considerably reduced in cells harbouring TagH275-FL at high temperature, supporting the results of IFM observation. These results suggest that the transmembrane segment of TagH plays an important role in WTA export at high temperatures.

Les bactéries à Gram positives multirésistantes : probabilités de résistance ? Que craindre ?

Antibiotic resistance in Gram-positive bacteria already has a significant impact on morbidity and mortality. The situation is particularly alarming in the case of methicillin-resistant Staphylococcus aureus and glycopeptide-resistant Enterococcus faecium, which have shown an ability to spread to new ecological niches and to generate new clones with both increased drug resistance and increased virulence. The potential for genetic exchanges between these two species raises the specter of highly resistant and virulent S. aureus strains.

Reversal of antibiotic resistance in Gram-positive bacteria by the antihistaminic azelastine

Antibiotic resistance represents a serious problem that complicates microbial infection. The use of 'helper compounds' capable of enhancing the antimicrobial activity of antibiotics is being investigated. Azelastine, a new generation antihistaminic, possesses certain antibacterial activity and is capable of inducing alteration in the bacterial membrane permeability. Hence, we hypothesized that it could reverse resistance to antibiotics. Azelastine significantly increased the antibacterial activity of eight antibiotics belonging to five different classes (β-lactams, macrolides, fluoroquinolones, aminoglycosides and tetracyclines) against nine Gram-positive clinical isolates: five Staphylococcus aureus, two Staphylococcus epidermidis and two Enterococcus faecium, seven of which were multi-drug resistant, reversing their resistance to the tested antibiotics. The synergistic effects of azelastine with the studied antibiotics increased with raising the pH from 5 to 8. Antibiotics did not affect the ability of azelastine to alter the permeability of a liposomal artificial membrane model, an effect thought to be critical for the interaction with antibiotics. The findings of this study present azelastine as a potential 'helper compound' that could reverse the resistance of multi-drug resistant Gram-positive clinical isolates to antibiotics.

Calcium-dependent potentiation of the pro-inflammatory functions of human neutrophils by tigecycline in vitro

Tigecycline is the prototype of the recently introduced, intravenously administered glycylcycline class of antibiotics, developed in response to the increasing problem of antibiotic resistance in Gram-positive bacteria, especially Staphylococcus aureus, as well as Gram-negative bacteria and anaerobes. However, relatively little is known about the immunomodulatory potential of tigecycline, specifically its interactions with human neutrophils. In the current study we investigated the effects of tigecycline at therapeutically relevant concentrations and greater (0.625-10 mg/L) on alterations in cytosolic Ca(2+) concentrations, generation of antimicrobial reactive oxygen species (ROS) and release of granule proteases [elastase, matrix metalloproteinase-8 (MMP-8) and matrix metalloproteinase-9 (MMP-9)] by human blood neutrophils activated with the chemoattractant N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP; 1 μM). Cytosolic Ca(2+) concentrations were measured using fura-2/AM-based spectrofluorimetry and radiometric procedures, generation of ROS by oxygen consumption and myeloperoxidase-mediated auto-iodination, and protease release by ELISA procedures. Exposure of the cells to fMLP resulted in activation of the generation of ROS, as well as release of the granule proteases, all of which were significantly increased by pre-incubation of the cells with tigecycline in a dose-dependent manner. Tigecycline-mediated enhancement of these neutrophil functions was associated with elevations in the concentrations of cytosolic Ca(2+), which appeared to result from the Ca(2+) ionophore activity of tigecycline. Tigecycline, by functioning as a Ca(2+) ionophore, and independent of antimicrobial activity, potentiates the pro-inflammatory functions of human neutrophils in vitro.

Streptogramins: an answer to antibiotic resistance in gram-positive bacteria

The past 15 years have seen the emergence of grampositive microorganisms as major nosocomial pathogens, and a corresponding increase in their antibiotic resistance. Penicillin-resistant pneumococci, meticillinresistant staphylococci, and vancomycin-resistant enterococci have been reported worldwide. 1 Gold HS Moellering Jr, RC Antimicrobial-drug resistance. N Engl J Med. 1996; 335: 1445-1453 Crossref PubMed Scopus (375) Google Scholar In addition, certain species, such as Enterococcus faecium and glycopeptide-intermediate Staphylococcus aureus (GISA), have become resistant to so many antibiotics that they have been described as “untreatable” pathogens. 2 Leclercq R Courvalin P Resistance to glycopeptides in enterococci. Clin Infect Dis. 1997; 24: 545-554 Crossref PubMed Scopus (198) Google Scholar , 3 Hiramatsu K Hanaki H Ino T Yabuta K Oguri T Tenover FC Methicillin-resistant Staphylococcus aureus clinical strain with reduced vancomycin susceptibility. J Antimicrob Chemother. 1997; 40: 135-136 Crossref PubMed Scopus (1646) Google Scholar However, the clinical consequences of such resistance should not be exaggerated. Extensive spread of resistant enterococci seems to be limited to the USA or to particular hospital ecosystems, and the number of publications on GISA greatly exceeds that of clinical isolates. Nevertheless, where present, these strains cause infections impossible to treat, and they would pose a substantial threat worldwide if they were to be disseminated.