Dataset Information

Autotransporters Drive Biofilm Formation and Autoaggregation in the Diderm Firmicute Veillonella parvula.

ABSTRACT: The Negativicutes are a clade of the Firmicutes that have retained the ancestral diderm character and possess an outer membrane. One of the best studied Negativicutes, Veillonella parvula, is an anaerobic commensal and opportunistic pathogen inhabiting complex human microbial communities, including the gut and the dental plaque microbiota. Whereas the adhesion and biofilm capacities of V. parvula are expected to be crucial for its maintenance and development in these environments, studies of V. parvula adhesion have been hindered by the lack of efficient genetic tools to perform functional analyses in this bacterium. Here, we took advantage of a recently described naturally transformable V. parvula isolate, SKV38, and adapted tools developed for the closely related Clostridia spp. to perform random transposon and targeted mutagenesis to identify V. parvula genes involved in biofilm formation. We show that type V secreted autotransporters, typically found in diderm bacteria, are the main determinants of V. parvula autoaggregation and biofilm formation and compete with each other for binding either to cells or to surfaces, with strong consequences for V. parvula biofilm formation capacity. The identified trimeric autotransporters have an original structure compared to classical autotransporters identified in Proteobacteria, with an additional C-terminal domain. We also show that inactivation of the gene coding for a poorly characterized metal-dependent phosphohydrolase HD domain protein conserved in the Firmicutes and their closely related diderm phyla inhibits autotransporter-mediated biofilm formation. This study paves the way for further molecular characterization of V. parvula interactions with other bacteria and the host within complex microbiota environments.IMPORTANCE Veillonella parvula is an anaerobic commensal and opportunistic pathogen whose ability to adhere to surfaces or other bacteria and form biofilms is critical for it to inhabit complex human microbial communities such as the gut and oral microbiota. Although the adhesive capacity of V. parvula has been previously described, very little is known about the underlying molecular mechanisms due to a lack of genetically amenable Veillonella strains. In this study, we took advantage of a naturally transformable V. parvula isolate and newly adapted genetic tools to identify surface-exposed adhesins called autotransporters as the main molecular determinants of adhesion in this bacterium. This work therefore provides new insights on an important aspect of the V. parvula lifestyle, opening new possibilities for mechanistic studies of the contribution of biofilm formation to the biology of this major commensal of the oral-digestive tract.

SUBMITTER: Bechon N

PROVIDER: S-EPMC7549365 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Autotransporters Drive Biofilm Formation and Autoaggregation in the Diderm Firmicute Veillonella parvula.

Béchon Nathalie N Jiménez-Fernández Alicia A Witwinowski Jerzy J Bierque Emilie E Taib Najwa N Cokelaer Thomas T Ma Laurence L Ghigo Jean-Marc JM Gribaldo Simonetta S Beloin Christophe C

Journal of bacteriology 20201008 21

The Negativicutes are a clade of the Firmicutes that have retained the ancestral diderm character and possess an outer membrane. One of the best studied Negativicutes, Veillonella parvula, is an anaerobic commensal and opportunistic pathogen inhabiting complex human microbial communities, including the gut and the dental plaque microbiota. Whereas the adhesion and biofilm capacities of V. parvula are expected to be crucial for its maintenance and development in ...[more]

PMID: 32817093

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Project description:Veillonella parvula is a biofilm-forming commensal found in the lungs, vagina, mouth, and gastro-intestinal tract of humans, yet it may develop into an opportunistic pathogen. Furthermore, the presence of Veillonella has been associated with the development of a healthy immune system in infants. Veillonella belongs to the Negativicutes, a diverse clade of bacteria that represent an evolutionary enigma: they phylogenetically belong to Gram-positive (monoderm) Firmicutes yet maintain an outer membrane (OM) with lipopolysaccharide similar to classic Gram-negative (diderm) bacteria. The OMs of Negativicutes have unique characteristics including the replacement of Braun's lipoprotein by OmpM for tethering the OM to the peptidoglycan. Through phylogenomic analysis, we have recently provided bioinformatic annotation of the Negativicutes diderm cell envelope. We showed that it is a unique type of envelope that was present in the ancestor of present-day Firmicutes and lost multiple times independently in this phylum, giving rise to the monoderm architecture; however, little experimental data is presently available for any Negativicutes cell envelope. Here, we performed the first experimental proteomic characterization of the cell envelope of a diderm Firmicute, producing an OM proteome of V. parvula. We initially conducted a thorough bioinformatics analysis of all 1,844 predicted proteins from V. parvula DSM 2008's genome using 12 different localization prediction programs. These results were complemented by protein extraction with surface exposed (SE) protein tags and by subcellular fractionation, both of which were analyzed by liquid chromatography tandem mass spectrometry. The merging of proteomics and bioinformatics results allowed identification of 78 OM proteins. These include a number of receptors for TonB-dependent transport, the main component of the BAM system for OM protein biogenesis (BamA), the Lpt system component LptD, which is responsible for insertion of LPS into the OM, and several copies of the major OmpM protein. The annotation of V. parvula's OM proteome markedly extends previous inferences on the nature of the cell envelope of Negativicutes, including the experimental evidence of a BAM/TAM system for OM protein biogenesis and of a complete Lpt system for LPS transport to the OM. It also provides important information on the role of OM components in the lifestyle of Veillonella, such as a possible gene cluster for O-antigen synthesis and a large number of adhesins. Finally, many OM hypothetical proteins were identified, which are priority targets for further characterization.

Project description:Veillonella parvula is a biofilm-forming commensal found in the lungs, vagina, mouth, and gastro-intestinal tract of humans, yet it may also develop into an opportunistic pathogen. Furthermore the presence of Veillonella has been associated with the development of a healthy immune system in infants. Veillonella belongs to the Negativicutes, a diverse clade of bacteria that represent an evolutionary enigma: they are phylogenetically placed within Gram-positive (monoderm) Firmicutes yet maintain an outer membrane (OM) with lipopolysaccharide similar to classic Gram-negative (diderm) bacteria. The OMs of Negativicutes have unique characteristics, such as the replacement of Braun’s lipoprotein by OmpM for anchoring the outer membrane to the peptidoglycan. Through phylogenomic analysis, we have recently provided the first bioinformatic annotation of the Negativicute diderm cell envelope. We showed that it is a unique type of envelope that was present in the ancestor of present-day Firmicutes and lost multiple times independently in this phylum, giving rise to the monoderm architecture. However, little experimental data is presently available for any Negativicute cell envelope. Here, we have performed the first experimental proteomic characterization of the cell envelope of these atypical diderm Firmicutes, producing an OM proteome of Veillonella parvula. We initially conducted a thorough bioinformatics analysis of all 1844 predicted proteins from Veillonella parvula DSM 2008’s genome using seven different localization prediction programs. These results were then complemented by protein extraction with surface exposed protein tags and subcellular fractionation, which were then sequenced by liquid chromatography tandem mass spectrometry. The merging of proteomics and bioinformatics results allowed identification of 76 OM proteins. Their annotation markedly extends previous inferences on the nature of the cell envelope of Negativicutes, and provides important information on the role of OM systems in the lifestyle of Veillonella.

Dataset Information

Autotransporters Drive Biofilm Formation and Autoaggregation in the Diderm Firmicute Veillonella parvula.

Publications

Autotransporters Drive Biofilm Formation and Autoaggregation in the Diderm Firmicute Veillonella parvula.

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