Project description:Purpose: We recently reported that isogenic deletion of lysine decarboxylase (ΔcadA/SP_0916), an enzyme that catalyzes the biosynthesis of polyamine cadaverine in Streptococcus pneumoniae TIGR4 results in loss of capsular polysaccharide (CPS), which constitutes a novel mechanism of regulation of CPS. Here, we conducted RNA-Seq to elucidate molecular mechanisms of CPS regulation in polyamine synthesis impaired pneumococci. Result: Significantly differentially expressed genes in ΔcadA represent pneumococcal pathways involved in the biosynthesis of precursors for CPS and peptidoglycan. Conclusion: We establish a possible link and interchange between two cellular processes such as high energy demanding capsule production and oxidative stress responses in polyamine synthesis impaired pneumococci (ΔcadA).
Project description:In this study, we monitored the transcriptional changes of S. aureus by RNA-seq analysis to better understand the effect of benzyl isothiocyanate (BITC) on the virulence inhibition of S. aureus and determined the bacteriostatic effect of BITC at subinhibitory concentrations. Our results revealed that compared with the control group (SAC), the BITC-treated experimental group (SAQ_BITC) had 708 differentially expressed genes (DEGs), of which 333 genes were downregulated and the capsular polysaccharide (cp) was significantly downregulated. Furthermore, we screened five of the most virulent factors of S. aureus, including the capsular polysaccharide biosynthesis protein (cp5D), capsular polysaccharide synthesis enzyme (cp8F), thermonuclease (nuc), clumping factor (clf) and protein A (spa).
Project description:The primary mechanism by which pneumococcal capsular polysaccharide-based vaccines are believed to mediate protection is by induction of serotype-specific opsonic antibodies that facilitate bacterial killing by phagocytes (opsonophagocytosis). However, antibodies that are protective against experimental pneumococcal pneumonia in mice but do not promote opsonophagocytic killing in vitro have also been identified 1-3. Such non-opsonic antibodies are associated with bacterial clearance in vivo, but the mechanism by which this occurs is unknown. In this letter, we demonstrate that a protective, non-opsonic serotype 3 pneumococcal capsular polysaccharide-specific monoclonal antibody (MAb) enhances quorum sensing, which results in competence induction and fratricide of serotype 3 pneumococcus. Gene expression profile analysis revealed that the MAb together with the pneumococcal autoinducer, competence stimulating peptide 2 (CSP2), augments differential expression of competence (com) related bacteriocin-like peptide (blp) genes that are known to be involved in pneumococcal fratricide. Taken together, these findings reveal a previously unsuspected mechanism of antibody action, namely, enhancement of quorum sensing and bacterial fratricide. Given that this activity does not require phagocytes, antibodies that function accordingly may hold promise as adjuncts to current vaccines or as desired products of next generation pneumococcal vaccines. 6 samples
Project description:The bacterial cell envelope is a complex, multilayered structure that is not only essential to maintain cellular integrity, but also facilitates vital bacterial processes such as adaption, colonization and adhesion. Cell envelopes comprise a wide range of molecules, such as proteins and capsular polysaccharide (CPS), which collectively decorate the bacterial cell in a species- and strain specific manner. Here we characterized the 4 CPS gene cluster of L. plantarum and assessed the impact on CPS (combinatorial) gene deletions on surface polysaccharide composition.
Project description:The primary mechanism by which pneumococcal capsular polysaccharide-based vaccines are believed to mediate protection is by induction of serotype-specific opsonic antibodies that facilitate bacterial killing by phagocytes (opsonophagocytosis). However, antibodies that are protective against experimental pneumococcal pneumonia in mice but do not promote opsonophagocytic killing in vitro have also been identified 1-3. Such non-opsonic antibodies are associated with bacterial clearance in vivo, but the mechanism by which this occurs is unknown. In this letter, we demonstrate that a protective, non-opsonic serotype 3 pneumococcal capsular polysaccharide-specific monoclonal antibody (MAb) enhances quorum sensing, which results in competence induction and fratricide of serotype 3 pneumococcus. Gene expression profile analysis revealed that the MAb together with the pneumococcal autoinducer, competence stimulating peptide 2 (CSP2), augments differential expression of competence (com) related bacteriocin-like peptide (blp) genes that are known to be involved in pneumococcal fratricide. Taken together, these findings reveal a previously unsuspected mechanism of antibody action, namely, enhancement of quorum sensing and bacterial fratricide. Given that this activity does not require phagocytes, antibodies that function accordingly may hold promise as adjuncts to current vaccines or as desired products of next generation pneumococcal vaccines.
Project description:The bacterial cell envelope is a complex, multilayered structure that is not only essential to maintain cellular integrity, but also facilitates vital bacterial processes such as adaption, colonization and adhesion. Cell envelopes comprise a wide range of molecules, such as proteins and capsular polysaccharide (CPS), which collectively decorate the bacterial cell in a species- and strain specific manner. Here we characterized the 4 CPS gene cluster of L. plantarum and assessed the impact on CPS (combinatorial) gene deletions on surface polysaccharide composition. individual KOs of the cps clusters and the combination of all four together were compared with the WT expression in a loop design. The KO of 1-3 was directly compared to the WT (dye swapped)
Project description:Invasive pneumococcal disease is preceded by asymptomatic colonization of the human nasopharynx by Streptococcus pneumoniae. Progression from colonization to invasion is a watershed in the host-pathogen interaction, and exposes the pneumococcus to markedly different microenvironments. This in turn, requires alterations in gene expression profile to adapt to the new niche. One apparent adaptive mechanism is reversible phase variation between “transparent” and “opaque” colony opacity phenotypes. Transparent phase variants colonize the nasopharynx more efficiently than opaque variants of the same strain, while opaque variants exhibit higher systemic virulence. Previous studies have reported quantitative differences in surface components such as the capsule, teichoic acid and certain surface proteins between the two phenotypes, but the underlying regulatory mechanism is not understood. In the present study, we found no differences in expression of key surface proteins between opaque and transparent variants of S. pneumoniae strain D39, but opaque cells produced five-fold more capsular polysaccharide. Subsequent microarray and real-time RT-PCR analysis showed no differences in capsule gene expression, but several genes involved in uridine monophosphate (UMP) biosynthesis were up-regulated in the opaque phenotype. This correlated with significant increases in the intracellular concentrations of both UMP and UDP-glucose, which are essential precursors for capsule biosynthesis. Our data suggest a novel mechanism for pneumococcal capsule regulation, in which rate-limiting precursor pathways are modulated rather than the capsule biosynthetic genes themselves. Keywords: Phase variants
Project description:P. multocida is the causative agent of a wide range of diseases of animals, including fowl cholera in birds. Fowl cholera isolates of P. multocida generally express a capsular polysaccharide composed of hyaluronic acid. There have been reports of spontaneous capsule loss in P. multocida fowl cholera-causing strains but the mechanism by which this occurs has not been determined. In this study, we identified three independent strains that had spontaneously lost the ability to produce capsular polysaccharide. Quantitative RT-PCR showed that these strains had significantly reduced transcription of the capsule biosynthetic genes, but DNA sequence analysis identified no mutations within the cap biosynthetic locus. However, whole genome sequencing of paired capsulated and acapsular strains identified a single nucleotide polymorphism within fis that was present only in the acapsular strain. Sequencing of fis from two independently derived spontaneous acapsular strains showed that each contained a mutation within fis. Complementation of these strains with an intact copy of fis returned normal capsule expression to all strains. Therefore, expression of a functional Fis protein is absolutely required for normal capsule expression in P. multocida.DNA microarray studies comparing one of the acapsular pairs (AL114 to AL1115) identified approximately 30 genes as down-regulated in the mutant; including pfh_B2 which encodes the filamentous hemagglutinin, a known P. multocida virulence factor and the cross protective surface antigen plpE.