scholarly journals Interconnections between Sigma B, agr, and Proteolytic Activity in Staphylococcus aureus Biofilm Maturation

2009 ◽  
Vol 77 (4) ◽  
pp. 1623-1635 ◽  
Author(s):  
Katherine J. Lauderdale ◽  
Blaise R. Boles ◽  
Ambrose L. Cheung ◽  
Alexander R. Horswill
Keyword(s):  
Staphylococcus Aureus ◽  
Hydrolase Activity ◽  
Extracellular Proteases ◽  
Sigma B ◽  
Genes Encoding ◽  
Usa300 Strain ◽  
Sigma Factor B ◽  
Murein Hydrolase ◽  
Host Tissues

ABSTRACT Staphylococcus aureus is a proficient biofilm former on host tissues and medical implants. We mutagenized S. aureus strain SH1000 to identify loci essential for ica-independent mechanisms of biofilm maturation and identified multiple insertions in the rsbUVW-sigB operon. Following construction and characterization of a sigB deletion, we determined that the biofilm phenotype was due to a lack of sigma factor B (SigB) activity. The phenotype was conserved in a sigB mutant of USA300 strain LAC, a well-studied community-associated methicillin-resistant S. aureus isolate. We determined that agr RNAIII levels were elevated in the sigB mutants, and high levels of RNAIII expression are known to have antibiofilm effects. By introducing an agr mutation into the SH1000 or LAC sigB deletion strain, S. aureus regained biofilm capacity, indicating that the biofilm phenotype was agr dependent. Protease activity is linked to agr activity and ica-independent biofilm formation, and we observed that the protease inhibitors phenylmethylsulfonyl fluoride and α-macroglobulin could reverse the sigB biofilm defect. Similarly, inactivating genes encoding both the aureolysin and Spl extracellular proteases in the sigB mutant restored biofilm capacity. Due to the growing link between murein hydrolase activity and biofilm maturation, autolysin zymography was performed, which revealed an altered profile in the sigB mutant; again, the phenotype could be repaired through protease inactivation. These findings indicate that the lack of SigB activity results in increased RNAIII expression, thus elevating extracellular protease levels and altering the murein hydrolase activity profile. Altogether, our observations demonstrate that SigB is an essential regulator of S. aureus biofilm maturation.

scholarly journals Transcription of the Staphylococcus aureus cid and lrg Murein Hydrolase Regulators Is Affected by Sigma Factor B

2004 ◽  
Vol 186 (10) ◽  
pp. 3029-3037 ◽  
Author(s):  
Kelly C. Rice ◽  
Toni Patton ◽  
Soo-Jin Yang ◽  
Alexis Dumoulin ◽  
Markus Bischoff ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Northern Blot ◽  
Exponential Growth ◽  
Sigma Factor ◽  
Positive Impact ◽  
Consensus Sequence ◽  
Hydrolase Activity ◽  
Factor B ◽  
Sigma Factor B ◽  
Murein Hydrolase

ABSTRACT The Staphylococcus aureus lrg and cid loci are homologous operons that have been shown to regulate murein hydrolase activity and affect sensitivity to penicillin. Although the mode of action of these operons has not been demonstrated, a model based on the similarities of the lrgA and cidA gene products to the bacteriophage holin family of proteins has been proposed. In this study, the transcription organization and regulation of these operons were examined by Northern blot analyses. Unexpectedly, cidB and a gene located immediately downstream, designated cidC, were found to be cotranscribed on a 2.7-kb transcript. Maximal cidBC transcription occurred during early exponential growth, and high-level transcription of cidBC was dependent on the rsbU-mediated activation of the alternative sigma factor B (σB). In contrast, lrgAB transcription in stationary phase was negatively regulated by σB. Although cidABC transcription was not detected by Northern blot analysis, reverse transcriptase PCR revealed that these genes are also cotranscribed as a single RNA message in early exponential growth. Primer extension analysis revealed the presence of two cidBC transcription start sites, but no apparent σB-dependent promoter consensus sequence was identified in these regions. The rsbU gene was also shown to have a positive impact on murein hydrolase activity but a negligible effect on sensitivity to penicillin-induced killing. These results suggest that the lrgAB and cidBC genes may be part of the S. aureus σB-controlled stress regulon.

scholarly journals Characterization of the Accessory Sec System of Staphylococcus aureus

2008 ◽  
Vol 190 (18) ◽  
pp. 6188-6196 ◽  
Author(s):  
Ian R. Siboo ◽  
Donald O. Chaffin ◽  
Craig E. Rubens ◽  
Paul M. Sullam
Keyword(s):  
Staphylococcus Aureus ◽  
Surface Expression ◽  
Complete Loss ◽  
Transport Pathway ◽  
Gram Positive Bacteria ◽  
Difference Gel Electrophoresis ◽  
Genes Encoding ◽  
Sec System ◽  
Specialized System

ABSTRACT The SraP adhesin of Staphylococcus aureus is a member of a highly conserved family of serine-rich surface glycoproteins of gram-positive bacteria. For streptococci, export of the SraP homologs requires a specialized transport pathway (the accessory Sec system). Compared to streptococci, however, SraP is predicted to differ in its signal peptide and glycosylation, which may affect its dependence on a specialized system for transport. In addition, two genes (asp4 and asp5) essential for export in Streptococcus gordonii are missing in S. aureus. Thus, the selectivity of the accessory Sec system in S. aureus may also differ compared to streptococci. To address these issues, the five genes encoding the putative accessory Sec system (secY2, secA2, and asp1-3) were disrupted individually in S. aureus ISP479C, and the resultant mutants were examined for SraP export. Disruption of secA2 resulted in the near complete loss of SraP surface expression. Similar results were seen with disruption of secY2 and asp1, asp2, or asp3. To assess whether the accessory Sec system transported other substrates, we compared secreted proteomes of ISP479C and a secA2 isogenic mutant, by two-dimensional fluorescence difference gel electrophoresis. Although two consistent differences in proteome content were noted between the strains, neither protein appeared to be a likely substrate for accessory Sec export. Thus, the accessory Sec system of S. aureus is required for the export of SraP, and it appears to be dedicated to the transport of this substrate exclusively.

scholarly journals Characterization of a CC1153 PVL-producing community-acquired methicillin-resistant Staphylococcus aureus from a dog bite wound

2017 ◽  
Vol 11 (06) ◽  
pp. 513-516 ◽  
Author(s):  
Edet Ekpenyong Udo ◽  
Samar Saed Boswihi ◽  
Tsonyu Ivanov Dimitrov ◽  
Bobby Noronha ◽  
Bindu Mathew ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Companion Animals ◽  
Dog Bite ◽  
Bite Wound ◽  
Methicillin Resistant ◽  
Wound Culture ◽  
Genes Encoding ◽  
Disease Hospital

The isolation of a rare community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) strain from an infected dog bite wound is here reported. A 27–year old man presented with a deep open wound on his right hand caused by his dog’s bite at the outpatient clinic of Infectious Disease Hospital (IDH), Kuwait. A wound swab was obtained for bacteriological culture and susceptibility testing. The wound culture yielded pure heavy growth of an MRSA isolate, designated IDH70, which was susceptible to vancomycin, teicoplanin, erythromycin, clindamycin, trimethoprim, fusidic acid and rifampicin. The patient was successfully treated with a combination of rifampicin and cotrimoxazole twice daily for 10 days. Molecular characterization revealed that IDH70 was positive for genes encoding Panton-Valentine leucocidin. IDH70 also carried the SCCmec V genetic element, belonged to coagulase type XIIIa, spa type t903, and was assigned to clonal complex 1153 and sequence type ST1153 (ST1153-V-t903). The report highlights the increasing burden of CA-MRSA in the community and the risk of its acquisition from bites of companion animals.

scholarly journals Epidemiology and microbiological characterization of clinical isolates of Staphylococcus aureus in a single healthcare region of the UK, 2015

2016 ◽  
Vol 145 (2) ◽  
pp. 386-396 ◽  
Author(s):  
C. HORNER ◽  
L. UTSI ◽  
L. COOLE ◽  
M. DENTON
Keyword(s):  
Staphylococcus Aureus ◽  
Methicillin Resistance ◽  
Current Data ◽  
Clinical Isolates ◽  
Dilution Method ◽  
Genes Encoding ◽  
Mupirocin Resistance ◽  
The Uk ◽  
Resistance Rates

SUMMARYWe investigated the epidemiology and characterization of isolates of Staphylococcus aureus within the Yorkshire and Humber (YH) region in the UK. In July 2015, each laboratory within YH (n = 14) was assigned two consecutive days during which all clinical isolates of S. aureus were collected. Isolates were tested for antibiotic susceptibilities and the presence of genes encoding methicillin resistance (mecA and mecC), Panton–Valentine leukocidin (PVL) (lukS-PV), and efflux-mediated chlorhexidine resistance (qacA); isolates were also characterized by spa-types. Minimum inhibitory concentrations (MICs) to chlorhexidine were determined by the broth dilution method. Of 520 isolates collected, 6·2% were methicillin-resistant S. aureus (MRSA, all mecA-positive) and mupirocin resistance was low [0·8%, 95% confidence interval (CI) 0·3–2·0] and only found in MRSA. Carriage of the qacA gene was identified in 1·7% (95% CI 0·8–3·3) of isolates and 3·5% (95% CI 2·2–5·4) had a chlorhexidine MIC of 4 mg/l. The PVL gene was infrequent (3·7%, 95% CI 2·4–5·6). Genotyping identified 234 spa-types that mapped to 22 clonal complexes. Comparison of these current data with previous work suggest that the widespread use of staphylococcal decolonization regimens over the past decade or more has not had an adverse impact on resistance rates, PVL carriage or the prevalence of specific S. aureus lineages.

scholarly journals Synergistic Effects of the Lactobacillus acidophilus Surface Layer and Nisin on Bacterial Growth

2009 ◽  
Vol 76 (3) ◽  
pp. 974-977 ◽  
Author(s):  
Mariano Prado-Acosta ◽  
Sandra M. Ruzal ◽  
Mariana C. Allievi ◽  
María Mercedes Palomino ◽  
Carmen Sanchez Rivas
Keyword(s):  
Staphylococcus Aureus ◽  
Surface Layer ◽  
Lactobacillus Acidophilus ◽  
Synergistic Effects ◽  
Proton Motive Force ◽  
Hydrolase Activity ◽  
Gram Positive ◽  
Gram Negative ◽  
Gram Positive Bacteria ◽  
Murein Hydrolase

ABSTRACT We have previously described a murein hydrolase activity for the surface layer (S-layer) of Lactobacillus acidophilus ATCC 4356. Here we show that, in combination with nisin, this S-layer acts synergistically to inhibit the growth of pathogenic Gram-negative Salmonella enterica and potential pathogenic Gram-positive bacteria, Staphylococcus aureus and Bacillus cereus. In addition, bacteriolytic effects were observed for the Gram-positive species tested. We postulate that the S-layer enhances the access of nisin into the cell membrane by enabling it to cross the cell wall, while nisin provides the sudden ion-nonspecific dissipation of the proton motive force required to enhance the S-layer murein hydrolase activity.

scholarly journals Human- and Plant-Pathogenic Pseudomonas Species Produce Bacteriocins Exhibiting Colicin M-Like Hydrolase Activity towards Peptidoglycan Precursors

2009 ◽  
Vol 191 (11) ◽  
pp. 3657-3664 ◽  
Author(s):  
Hélène Barreteau ◽  
Ahmed Bouhss ◽  
Martine Fourgeaud ◽  
Jean-Luc Mainardi ◽  
Thierry Touzé ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Hydrolase Activity ◽  
New Class ◽  
Low Concentrations ◽  
Pseudomonas Species ◽  
Genes Encoding ◽  
Colicin M ◽  
Undecaprenyl Phosphate

ABSTRACT Genes encoding proteins that exhibit similarity to the C-terminal domain of Escherichia coli colicin M were identified in the genomes of some Pseudomonas species, namely, P. aeruginosa, P. syringae, and P. fluorescens. These genes were detected only in a restricted number of strains. In P. aeruginosa, for instance, the colicin M homologue gene was located within the ExoU-containing genomic island A, a large horizontally acquired genetic element and virulence determinant. Here we report the cloning of these genes from the three Pseudomonas species and the purification and biochemical characterization of the different colicin M homologues. All of them were shown to exhibit Mg2+-dependent diphosphoric diester hydrolase activity toward the two undecaprenyl phosphate-linked peptidoglycan precursors (lipids I and II) in vitro. In all cases, the site of cleavage was localized between the undecaprenyl and pyrophospho-MurNAc moieties of these precursors. These enzymes were not active on the cytoplasmic precursor UDP-MurNAc-pentapeptide or (or only very poorly) on undecaprenyl pyrophosphate. These colicin M homologues have a narrow range of antibacterial activity. The P. aeruginosa protein at low concentrations was shown to inhibit growth of sensitive P. aeruginosa strains. These proteins thus represent a new class of bacteriocins (pyocins), the first ones reported thus far in the genus Pseudomonas that target peptidoglycan metabolism.

scholarly journals Characterization of Phage Obtained from Methicillin -Resistant Staphylococcus aureus (MRSA)

2021 ◽  
Author(s):  
Özlem Ulusan Bağcı ◽  
Fikret Şahin ◽  
Mehmet Kıyan
Keyword(s):  
Staphylococcus Aureus ◽  
Phage Therapy ◽  
Relevant Literature ◽  
Open Reading Frames ◽  
Recombinant Phage ◽  
Protein Map ◽  
Genes Encoding ◽  
New Antibiotics ◽  
Lysogenic Phage

Objective: The emergence of Staphylococcus aureus strains resistant to all antimicrobials and failure to discover new antibiotics have led researchers to phage therapy, which lost popularity after the discovery of antibiotics. The development of recombinant technology introduced the idea of creating lysogenic recombinant phages that provide controlled bacterial death and this required small- sized phages that were easy to manipulate. Our aim is to identify small-sized lysogenic bacteriophages that can be used safely in therapy. Method: The gene and protein map of the phage was created by analysis of sequencing after extracting a phage from the MRSA strain that is known to contain a small phage. Results: The phage was classified in Caudovirales spp. as it contains genes encoding tail proteins, and in Podoviridae spp. due to its genomic size and arrangement. Conclusion: To date, there are only sixteen phages from Podoviridae family uploaded on NCBI, and the phage described in this study is the seventeenth one. Only 41.4% of the ORFs (Open Reading Frames) in the genome could be matched with proteins using the NCBI BLAST. Recent studies suggest that 50-75% of bacteriophage ORFs do not correspond to any organism in GenBank. For better understanding of bacteriophages and their utilization in phage therapy, it is essential to sequence greater number of phages, and to discover their genomes and corresponding proteins. Since the genes and proteins of a lysogenic phage that can be used safely in recombinant phage therapies have been identified in our study, it will contribute to the relevant literature.

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