Biology subjects

Pier, G.

Publications and source records attributed to Pier, G..

3 recordsLinked to original sources

bioRxiv2022

A high-throughput sequencing approach identifies immunotherapeutic targets for bacterial meningitis in neonates

BackgroundWorldwide, Escherichia coli is the leading cause of neonatal Gram-negative bacterial meningitis, but full understanding of the pathogenesis of this disease is not yet achieved. Moreover, to date, no vaccine is available against bacterial neonatal meningitis. MethodsHere, we used Transposon Sequencing of saturated banks of mutants (TnSeq) to evaluate E. coli K1 genetic fitness in murine neonatal meningitis. We identified E. coli K1 genes encoding for factors important for systemic dissemination and brain infection, and focused on products with a likely outer-membrane or extra-cellular localization, as these are potential vaccine candidates. We used in vitro and in vivo models to study the efficacy of active and passive immunization. ResultsWe selected for further study the conserved surface polysaccharide Poly-{beta}-(1-6)-N-Acetyl Glucosamine (PNAG), as a strong candidate for vaccine development. We found that PNAG was a virulence factor in our animal model. We showed that both passive and active immunization successfully prevented and/or treated meningitis caused by E. coli K1 in neonatal mice. We found an excellent opsonophagocytic killing activity of the antibodies to PNAG and in vitro these antibodies were also able to decrease binding, invasion and crossing of E. coli K1 through two blood brain barrier cell lines. Finally, to reinforce the potential of PNAG as a vaccine candidate in bacterial neonatal meningitis, we demonstrated that Group B Streptococcus, the main cause of neonatal meningitis in developed countries, also produced PNAG and that antibodies to PNAG could protect in vitro and in vivo against this major neonatal pathogen. InterpretationAltogether, these results indicate the utility of a high-throughput DNA sequencing method to identify potential immunotherapy targets for a pathogen, including in this study a potential broad-spectrum target for prevention of neonatal bacterial infections. FundingsANR Seq-N-Vaq, Charles Hood Foundation, Hearst Foundation. Groupe Pasteur Mutualite

microbiology↗

bioRxiv2021

A qnr-plasmid allows aminoglycosides to induce SOS in Escherichia coli

The plasmid-mediated quinolone resistance (PMQR) genes have been shown to promote high-level bacterial resistance to fluoroquinolone antibiotics, potentially leading to clinical treatment failures. In Escherichia coli, sub-inhibitory concentrations (sub-MIC) of the widely used fluoroquinolones are known to induce the SOS response. Interestingly, the expression of several PMQR qnr genes is controlled by the SOS master regulator. During the characterization of a small qnrD-plasmid carried in E. coli, we observed that the aminoglycosides become able to induce the SOS response in this species, thus leading to the transcription of qnrD. We found that induction of the SOS response is due to nitric oxide (NO) accumulation in presence of sub-MIC of aminoglycosides. We demonstrated that the NO accumulation is driven by two plasmid genes, ORF3 and ORF4, whose products act at two levels. ORF3 encode a FAD-binding oxidoreductase which helps NO synthesis, while ORF4 code for an FNR-type transcription factor, related to an O2-responsive regulator of hmp expression, able to repress the Hmp-mediated NO detoxification pathway of E. coli. Thus, this discovery, that other major classes of antibiotics may induce the SOS response could have worthwhile implications for antibiotic stewardship efforts in preventing the emergence of resistance.

microbiology↗

bioRxiv2016

Hyphal aggregation and surface attachment of Streptomyces is governed by extracellular poly-β-1,6-N-acetylglucosamine

Streptomycetes are multicellular filamentous microorganisms, which are major producers of antibiotics, anticancer drugs and industrial enzymes. When grown in submerged cultures, the preferred enzyme producer, Streptomyces lividans, forms dense mycelial aggregates or pellets, which requires the activity of the proteins encoded by the matAB and cslA-glxA. Here we show that matAB encodes the biosynthetic genes for the extracellular polymeric substance (EPS) poly-{beta}-1,6-N-acetylglucosamine or PNAG. Heterologous expression of matAB in actinomycetes that naturally lack these genes was sufficient for PNAG production and induction of mycelial aggregation. Also, overexpression of matAB in a non-pelleting cslA mutant restored pellet formation, which could effectively be antagonized by the PNAG-specific hydrolase, dispersin B. Extracellular accumulation of PNAG allowed Streptomyces to attach to hydrophilic surfaces, unlike attachment to hydrophobic surfaces, which involves a cellulase-degradable EPS produced by CslA. Altogether, our data support a model in which pellet formation depends on hydrophilic interactions mediated by PNAG and hydrophobic interactions involving the EPS produced by CslA. These new insights may be harnessed to improve growth and industrial exploitation of these highly versatile natural product and enzyme producers.

microbiology↗