Nonconformity of biofilm formation in vivo and in vitro based on Staphylococcus aureus accessory gene regulator status.
Ontology highlight
ABSTRACT:
SUBMITTER: Jordan SC
PROVIDER: S-EPMC8786897 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Staphylococcus aureus is a significant cause of chronic biofilm infections on medical implants. We investigated the biofilm regulatory cascade and discovered that the repressor of toxins (Rot) is part of this pathway. A USA300 community-associated methicillin-resistant S.?aureus strain deficient in Rot was unable to form a biofilm using multiple different assays, and we found rot mutants in other strain lineages were also biofilm deficient. By performing a global analysis of transcripts and protein production controlled by Rot, we observed that all the secreted protease genes were up-regulated in a rot mutant, and we hypothesized that this regulation could be responsible for the biofilm phenotype. To investigate this question, we determined that Rot bound to the protease promoters, and we observed that activity levels of these enzymes, in particular the cysteine proteases, were increased in a rot mutant. By inactivating these proteases, biofilm capacity was restored to the mutant, demonstrating they are responsible for the biofilm negative phenotype. Finally, we tested the rot mutant in a mouse catheter model of biofilm infection and observed a significant reduction in biofilm burden. Thus S.?aureus uses the transcription factor Rot to repress secreted protease levels in order to build a biofilm.
| S-EPMC4467170 | biostudies-literature
Project description:Biofilm formation in Staphylococcus aureus under in vitro growth conditions is generally promoted by high concentrations of sugar and/or salts. The addition of glucose to routinely used complex growth media triggered biofilm formation in S. aureus strain SA113. Deletion of ccpA, coding for the catabolite control protein A (CcpA), which regulates gene expression in response to the carbon source, abolished the capacity of SA113 to form a biofilm under static and flow conditions, while still allowing primary attachment to polystyrene surfaces. This suggested that CcpA mainly affects biofilm accumulation and intercellular aggregation. trans-Complementation of the mutant with the wild-type ccpA allele fully restored the biofilm formation. The biofilm produced by SA113 was susceptible to sodium metaperiodate, DNase I, and proteinase K treatment, indicating the presence of polysaccharide intercellular adhesin (PIA), protein factors, and extracellular DNA (eDNA). The investigation of several factors which were reported to influence biofilm formation in S. aureus (arlRS, mgrA, rbf, sarA, atl, ica, citZ, citB, and cidABC) showed that CcpA up-regulated the transcription of cidA, which was recently shown to contribute to eDNA production. Moreover, we showed that CcpA increased icaA expression and PIA production, presumably over the down-regulation of the tricarboxylic acid cycle genes citB and citZ.
| S-EPMC2346702 | biostudies-literature
Project description:Staphylococcus aureus generates biofilms during many chronic human infections, which contributes to its growth and persistence in the host. Multiple genes and pathways necessary for S. aureus biofilm production have been identified, but knowledge is incomplete, and little is known about spontaneous mutations that increase biofilm formation as infection progresses. Here, we performed in vitro selection of four S. aureus laboratory strains (ATCC 29213, JE2, N315, and Newman) to identify mutations associated with enhanced biofilm production. Biofilm formation increased in passaged isolates from all strains, exhibiting from 1.2- to 5-fold the capacity of parental lines. Whole-genome sequencing identified nonsynonymous mutations affecting 23 candidate genes and a genomic duplication encompassing sigB. Six candidate genes significantly impacted biofilm formation as isogenic transposon knockouts: three were previously reported to impact S. aureus biofilm formation (icaR, spdC, and codY), while the remaining three (manA, narH, and fruB) were newly implicated by this study. Plasmid-mediated genetic complementation of manA, narH, and fruB transposon mutants corrected biofilm deficiencies, with high-level expression of manA and fruB further enhancing biofilm formation over basal levels. This work recognizes genes not previously identified as contributing to biofilm formation in S. aureus and reveals genetic changes able to augment biofilm production by that organism.
| S-EPMC10016075 | biostudies-literature
Project description:Staphylococcus aureus is Gram-positive bacterium commonly associated with nosocomial infections. The development of biofilm exhibiting drug resistance especially in foreign body associated infections has enabled the bacterium to draw considerable attention. However, till date, consensus guidelines for in vitro biofilm quantitation and categorization criterion for the bacterial isolates based on biofilm-forming capacity are lacking. Therefore, it was intended to standardize in vitro biofilm formation by clinical isolates of S. aureus and then to classify them on the basis of their biofilm-forming capacity.A study was conducted for biofilm quantitation by tissue culture plate (TCP) assay employing 61 strains of S. aureus isolated from clinical samples during May 2015- December 2015 wherein several factors influencing the biofilm formation were optimized. Therefore, it was intended to propose a biofilm classification criteria based on the standard deviation multiples of the control differentiating them into non, low, medium, and high biofilm formers.Brain-heart infusion broth was found to be more effective in biofilm formation compared to trypticase soy broth. Heat fixation was more effective than chemical fixation. Although, individually, glucose, sucrose, and sodium chloride (NaCl) had no significant effect on biofilm formation, a statistically significant increase in absorbance was observed after using the supplement mix consisting of 222.2 mM glucose, 116.9 mM sucrose, and 1000 mM NaCl (P= 0.037).The present study puts forth a standardized in vitro TCP assay for biofilm biomass quantitation and categorization criteria for clinical isolates of S. aureus based on their biofilm-forming capacity. The proposed in vitro technique may be further evaluated for its usefulness in the management of persistent infections caused by the bacterium.
| S-EPMC5572203 | biostudies-literature
Project description:Biofilms are multicellular communities of microorganisms living as a quorum rather than as individual cells. The bacterial human pathogen Staphylococcus aureus uses oxygen as a terminal electron acceptor during respiration. Infected human tissues are hypoxic or anoxic. We recently reported that impaired respiration elicits a programmed cell lysis (PCL) phenomenon in S. aureus leading to the release of cellular polymers that are utilized to form biofilms. PCL is dependent upon the AtlA murein hydrolase and is regulated, in part, by the SrrAB two-component regulatory system (TCRS). In the current study, we report that the SaeRS TCRS also governs fermentative biofilm formation by positively influencing AtlA activity. The SaeRS-modulated factor fibronectin-binding protein A (FnBPA) also contributed to the fermentative biofilm formation phenotype. SaeRS-dependent biofilm formation occurred in response to changes in cellular respiratory status. Genetic evidence presented suggests that a high cellular titer of phosphorylated SaeR is required for biofilm formation. Epistasis analyses found that SaeRS and SrrAB influence biofilm formation independently of one another. Analyses using a mouse model of orthopedic implant-associated biofilm formation found that both SaeRS and SrrAB govern host colonization. Of these two TCRSs, SrrAB was the dominant system driving biofilm formation in vivo We propose a model wherein impaired cellular respiration stimulates SaeRS via an as yet undefined signal molecule(s), resulting in increasing expression of AtlA and FnBPA and biofilm formation.
| S-EPMC5520442 | biostudies-literature
Project description:The accessory gene regulator (agr) locus influences the expression of many virulence genes in the human pathogen Staphylococcus aureus. Four allelic groups of agr, which generally inhibit the regulatory activity of each other, have been identified within the species. Interference in virulence gene expression caused by different agr groups has been suggested to be a mechanism for isolating bacterial populations and a fundamental basis for subdividing the species. To test the hypothesis that the species is phylogenetically structured according to agr groups, we mapped agr groups onto a clone phylogeny inferred from partial sequences of 14 genes from 27 genetically diverse strains. Shimodaira-Hasegawa and parametric bootstrap tests rejected the hypotheses that the species is subdivided into three or five monophyletic agr groups but failed to reject the hypothesis that the species is subdivided into two groups that each consist of multiple clonal complexes and multiple agr groups. Additional evidence for agr recombination is found from clustered polymorphisms in complete agr sequences. However, agr recombination has not occurred frequently or randomly through time, because the topology and branch lengths of the clone phylogeny are reflected within each agr group. To account for these observations, we propose a new evolutionary model that involves a genetically polymorphic ancestral population of S. aureus that horizontally transferred agr groups between two subspecies groups near the time that these subspecies groups diverged.
| S-EPMC1317016 | biostudies-literature
Project description:Staphylococcus aureus is a human pathogen that forms biofilm on catheters and medical implants. The authors' earlier study established that 1,2,3,4,6-penta-O-galloyl-β-D-glucopyranose (PGG) inhibits biofilm formation by S. aureus by preventing the initial attachment of the cells to a solid surface and reducing the production of polysaccharide intercellular adhesin (PIA). Our cDNA microarray and MALDI-TOF mass spectrometric studies demonstrate that PGG treatment causes the expression of genes and proteins that are normally expressed under iron-limiting conditions. A chemical assay using ferrozine verifies that PGG is a strong iron chelator that depletes iron from the culture medium. This study finds that adding FeSO(4) to a medium that contains PGG restores the biofilm formation and the production of PIA by S. aureus SA113. The requirement of iron for biofilm formation by S. aureus SA113 can also be verified using a semi-defined medium, BM, that contains an iron chelating agent, 2, 2'-dipyridyl (2-DP). Similar to the effect of PGG, the addition of 2-DP to BM medium inhibits biofilm formation and adding FeSO(4) to BM medium that contains 2-DP restores biofilm formation. This study reveals an important mechanism of biofilm formation by S. aureus SA113.
| S-EPMC3313993 | biostudies-literature
Project description:Biofilm-based infection is a major healthcare burden. Methicillin-resistant Staphylococcus aureus (MRSA) is one of major organisms responsible for biofilm infection. Although biofilm is induced by a number of environmental signals, the molecule responsible for environmental sensing is not well delineated. Here we examined the role of ion transporters in biofilm formation and found that the sodium-glutamate transporter gltS played an important role in biofilm formation in MRSA. This was shown by gltS transposon mutant as well as its complementation. The lack of exogenous glutamate also enhanced biofilm formation in JE2 strain. The deficiency of exogenous glutamate intake accelerated endogenous glutamate/glutamine production, which led to the activation of the urea cycle. We also showed that urea cycle activation was critical for biofilm formation. In conclusion, we showed that gltS was a critical regulator of biofilm formation by controlling the intake of exogenous glutamate. An intervention to target glutamate intake may be a potential useful approach against biofilm.
| S-EPMC9684512 | biostudies-literature
Project description:BackgroundStaphylococcus aureus is a leading cause of human infections. The spread of antibiotic-resistant staphylococci has driven the search for novel strategies to supersede antibiotics use. Thus, targeting bacterial virulence rather than viability could be a possible alternative.ResultsThe influence of celastrol on staphyloxanthin (STX) biosynthesis, biofilm formation, antibiotic susceptibility and host pathogenesis in S. aureus has been investigated. Celastrol efficiently reduced STX biosynthesis in S. aureus. Liquid chromatography-mass spectrometry (LC-MS) and molecular docking revealed that celastrol inhibits STX biosynthesis through its effect on CrtM. Quantitative measurement of STX intermediates showed a significant pigment inhibition via interference of celastrol with CrtM and accumulation of its substrate, farnesyl diphosphate. Importantly, celastrol-treated S. aureus was more sensitive to environmental stresses and human blood killing than untreated bacteria. Similarly, inhibition of STX upon celastrol treatment rendered S. aureus more susceptible to membrane targeting antibiotics. In addition to its anti-pigment capability, celastrol exhibits significant anti-biofilm activity against S. aureus as indicated by crystal violet assay and microscopy. Celastrol-treated cells showed deficient exopolysaccharide production and cell hydrophobicity. Moreover, celastrol markedly synergized the action of conventional antibiotics against S. aureus and reduced bacterial pathogenesis in vivo using mice infection model. These findings were further validated using qRT-PCR, demonstrating that celastrol could alter the expression of STX biosynthesis genes as well as biofilm formation related genes and bacterial virulence.ConclusionsCelastrol is a novel anti-virulent agent against S. aureus suggesting, a prospective therapeutic role for celastrol as a multi-targeted anti-pathogenic agent.
| S-EPMC9011992 | biostudies-literature
Project description:The SaeRS two-component regulatory system of Staphylococcus aureus is known to affect the expression of many genes. The SaeS protein is the histidine kinase responsible for phosphorylation of the response regulator SaeR. In S. aureus Newman, the sae system is constitutively expressed due to a point mutation in saeS, relative to other S. aureus strains, which results in substitution of proline for leucine at amino acid 18. Strain Newman is unable to form a robust biofilm and we report here that the biofilm-deficient phenotype is due to the saeSP allele. Replacement of the Newman saeSP with saeSL, or deletion of saeRS, resulted in a biofilm-proficient phenotype. Newman culture supernatants were observed to inhibit biofilm formation by other S. aureus strains, but did not affect biofilm formation by S. epidermidis. Culture supernatants of Newman saeSL or Newman ΔsaeRS had no significant effect on biofilm formation. The inhibitory factor was inactivated by incubation with proteinase K, but survived heating, indicating that the inhibitory protein is heat-stable. The inhibitory protein was found to affect the attachment step in biofilm formation, but had no effect on preformed biofilms. Replacement of saeSL with saeSP in the biofilm-proficient S. aureus USA300 FPR3757 resulted in the loss of biofilm formation. Culture supernatants of USA300 FPR3757 saeSP, did not inhibit biofilm formation by other staphylococci, suggesting that the inhibitory factor is produced but not secreted in the mutant strain. A number of biochemical methods were utilized to isolate the inhibitory protein. Although a number of candidate proteins were identified, none were found to be the actual inhibitor. In an effort to reduce the number of potential inhibitory genes, RNA-Seq analyses were done with wild-type strain Newman and the saeSL and ΔsaeRS mutants. RNA-Seq results indicated that sae regulates many genes that may affect biofilm formation by Newman.
2015-04-30 | GSE65827 | GEO