A new tool for cloning and gene expression in Streptococcus pneumoniae

Domingues, Susana; Aires, Andreia; Mohedano, María Luz; López, Paloma; Arraiano, Cecília M.

doi:10.1016/j.plasmid.2013.05.005

Cited by 8 publications

(7 citation statements)

References 36 publications

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“…Interestingly, infection with the strain expressing RNase R in trans even diminishes the survival rate. This is indicative of a direct role of RNase R in the pneumococcus pathogenicity, as, in this strain, RNase R levels are higher than in the wild type [37].…”

Section: Discussionmentioning

confidence: 73%

“…A reversion to higher expression levels was observed in the complemented RNase R strain. However, in this strain, the NanA transcriptional levels were increased relative to the wild type amount, which might be due to the augmented level of plasmid-expressed RNase R [37]. Together, these results are indicative of a role for RNase R in the modulation of this virulence factor.…”

Section: Transcriptional Expression Of Nana Is Affected By Rnase Rmentioning

confidence: 69%

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RNase R, a New Virulence Determinant of Streptococcus pneumoniae

et al. 2022

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Pneumococcal infections have increasingly high mortality rates despite the availability of vaccines and antibiotics. Therefore, the identification of new virulence determinants and the understanding of the molecular mechanisms behind pathogenesis have become of paramount importance in the search of new targets for drug development. The exoribonuclease RNase R has been involved in virulence in a growing number of pathogens. In this work, we used Galleria mellonella as an infection model to demonstrate that the presence of RNase R increases the pneumococcus virulence. Larvae infected with the RNase R mutant show an increased expression level of antimicrobial peptides. Furthermore, they have a lower bacterial load in the hemolymph in the later stages of infection, leading to a higher survival rate of the larvae. Interestingly, pneumococci expressing RNase R show a sudden drop in bacterial numbers immediately after infection, resembling the eclipse phase observed after intravenous inoculation in mice. Concomitantly, we observed a lower number of mutant bacteria inside larval hemocytes and a higher susceptibility to oxidative stress when compared to the wild type. Together, our results indicate that RNase R is involved in the ability of pneumococci to evade the host immune response, probably by interfering with internalization and/or replication inside the larval hemocytes.

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Section: Discussionmentioning

confidence: 73%

Section: Transcriptional Expression Of Nana Is Affected By Rnase Rmentioning

confidence: 69%

RNase R, a New Virulence Determinant of Streptococcus pneumoniae

et al. 2022

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“…Gene expression mutants were constructed using the medium-copy-number expression vector pIL253 (31). Both operons targeted by genetic engineering are absent from the genome of L. rhamnosus ATCC 53103, which could therefore be employed as the heterologous expression host.…”

Section: Methodsmentioning

confidence: 99%

Correlation of Lactobacillus rhamnosus Genotypes and Carbohydrate Utilization Signatures Determined by Phenotype Profiling

Ceapă

Lambert

Limpt

et al. 2015

Appl Environ Microbiol

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e Lactobacillus rhamnosus is a bacterial species commonly colonizing the gastrointestinal (GI) tract of humans and also frequently used in food products. While some strains have been studied extensively, physiological variability among isolates of the species found in healthy humans or their diet is largely unexplored. The aim of this study was to characterize the diversity of carbohydrate utilization capabilities of human isolates and food-derived strains of L. rhamnosus in relation to their niche of isolation and genotype. We investigated the genotypic and phenotypic diversity of 25 out of 65 L. rhamnosus strains from various niches, mainly human feces and fermented dairy products. Genetic fingerprinting of the strains by amplified fragment length polymorphism (AFLP) identified 11 distinct subgroups at 70% similarity and suggested niche enrichment within particular genetic clades. High-resolution carbohydrate utilization profiling (OmniLog) identified 14 carbon sources that could be used by all of the strains tested for growth, while the utilization of 58 carbon sources differed significantly between strains, enabling the stratification of L. rhamnosus strains into three metabolic clusters that partially correlate with the genotypic clades but appear uncorrelated with the strain's origin of isolation. Draft genome sequences of 8 strains were generated and employed in a gene-trait matching (GTM) analysis together with the publicly available genomes of L. rhamnosus GG (ATCC 53103) and HN001 for several carbohydrates that were distinct for the different metabolic clusters: L-rhamnose, cellobiose, L-sorbose, and ␣-methyl-Dglucoside. From the analysis, candidate genes were identified that correlate with L-sorbose and ␣-methyl-D-glucoside utilization, and the proposed function of these genes could be confirmed by heterologous expression in a strain lacking the genes. This study expands our insight into the phenotypic and genotypic diversity of the species L. rhamnosus and explores the relationships between specific carbohydrate utilization capacities and genotype and/or niche adaptation of this species.

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“…Transcriptomic analysis was performed to better understand how RNase R affects gene expression in S. pneumoniae clinical isolate TIGR4. We compared the transcriptome between Δ rnr mutant previously constructed [10, 12] and the WT by performing RNA-Seq of total RNA. We found 257 transcripts that had high expression values in either strain and more than 0.5 of log 2 FC between the two strains (Table S3 in Supplementary Material).…”

Section: Resultsmentioning

confidence: 99%

RNase R Controls Membrane Fatty Acid Composition inStreptococcus pneumoniae

Alipio

Bárria

Pobre

et al. 2023

Preprint

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Previous studies on RNase R have highlighted significant effects of this ribonuclease in several processes ofStreptococcus pneumoniaebiology. In this work we have studied the global impact of RNase R by comparing the transcriptional landscape of a deleted RNase R mutant to that of the wild-type strain, and this led us investigate specific targets affected by RNase R. RNA-Seq showed that RNase R deletion affects transcripts from several different biological processes. Of particular interest, elimination of RNase R results in overexpression of most of the genes encoding the components of type II fatty acid biosynthesis (FAS-II) cluster. We demonstrate that RNase R governs the turnover of most of genes from this pathway, affecting the outcome of the whole FAS-II cluster, and leading to an unbalanced membrane fatty acid composition. Our results show that the membrane of the deleted strain contains a higher proportion of unsaturated and long-chained fatty acids than the wild type strain. This leads to a higher fluidity of the Δrnrmutant membrane, which is probably related with the increased sensitivity to detergent observed in this strain. We demonstrate that RNase R expression is induced in cells challenged with H2O2, which is suggestive of a role for this ribonuclease on the regulation of membrane homeostasis under oxidative stress. Reprogramming of membrane fluidity is an adaptative cell response crucial for bacterial survival in constantly changing environmental conditions. The fact that RNase R controls the expression of several essential genes to the fatty acid synthesis unveils a new important function of this enzyme.

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A new tool for cloning and gene expression in Streptococcus pneumoniae

Cited by 8 publications

References 36 publications

RNase R, a New Virulence Determinant of Streptococcus pneumoniae

RNase R, a New Virulence Determinant of Streptococcus pneumoniae

Correlation of Lactobacillus rhamnosus Genotypes and Carbohydrate Utilization Signatures Determined by Phenotype Profiling

RNase R Controls Membrane Fatty Acid Composition inStreptococcus pneumoniae

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