A Novel Repressor of the
            <i>ica</i>
            Locus Discovered in Clinically Isolated Super-Biofilm-Elaborating
            <i>Staphylococcus aureus</i>

Yu, Liansheng; Hisatsune, Junzo; Hayashi, Ikue; Tatsukawa, Nobuyuki; Sato’o, Yusuke; Mizumachi, Emiri; Kato, Fuminori; Hirakawa, Hideki; Pier, Gerald B.; Sugai, Motoyuki

doi:10.1128/mbio.02282-16

Cited by 31 publications

(58 citation statements)

References 42 publications

(65 reference statements)

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“…Furthermore, glucose is normally added in the TSB to promote S. aureus biofilm formation in vitro [29]; however, various glucose concentrations have been used in different studies. For example, TSB (which already contains 2.5 g/L glucose and 5.0 g/L NaCl) is often supplemented with additional glucose (0.25%, 0.4%, 0.5%, and 1.0%) [22,30,31,32,33,34]. In addition, brain heart infusion (BHI) (which already contains 2.0 g/L glucose) is also used as a culture media for S. aureus biofilm formation in vitro [35], with additional glucose supplementation (0.5%, 1.0%, and 1.5%) [36,37].…”

Section: Introductionmentioning

confidence: 99%

Biofilm Formation by Staphylococcus aureus Clinical Isolates is Differentially Affected by Glucose and Sodium Chloride Supplemented Culture Media

Lade

Park

Chung

et al. 2019

JCM

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Staphylococcus aureus (S. aureus) causes persistent biofilm-related infections. Biofilm formation by S. aureus is affected by the culture conditions and is associated with certain genotypic characteristics. Here, we show that glucose and sodium chloride (NaCl) supplementation of culture media, a common practice in studies of biofilms in vitro, influences both biofilm formation by 40 S. aureus clinical isolates (methicillin-resistant and methicillin-sensitive S. aureus) and causes variations in biofilm quantification. Methicillin-resistant strains formed more robust biofilms than methicillin-sensitive strains in tryptic soy broth (TSB). However, glucose supplementation in TSB greatly promoted and stabilized biofilm formation of all strains, while additional NaCl was less efficient in this respect and resulted in significant variation in biofilm measurements. In addition, we observed that the ST239-SCCmec (Staphylococcal Cassette Chromosome mec) type III lineage formed strong biofilms in TSB supplemented with glucose and NaCl. Links between biofilm formation and accessory gene regulator (agr) status, as assessed by δ-toxin production, and with mannitol fermentation were not found. Our results show that TSB supplemented with 1.0% glucose supports robust biofilm production and reproducible quantification of S. aureus biofilm formation in vitro, whereas additional NaCl results in major variations in measurements of biofilm formation.

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Section: Introductionmentioning

confidence: 99%

Biofilm Formation by Staphylococcus aureus Clinical Isolates is Differentially Affected by Glucose and Sodium Chloride Supplemented Culture Media

Lade

Park

Chung

et al. 2019

JCM

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“…We chose pKAT as a framework of the shuttle vectors, because we confirmed that this plasmid could be introduced into the S . aureus clinical strain, which did not accept the previous TS-vector [ 33 ]. We also used CRISPR/Cas9 components from pCas9 [ 19 ].…”

Section: Resultsmentioning

confidence: 99%

Tailor-made gene silencing of Staphylococcus aureus clinical isolates by CRISPR interference

et al. 2018

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Preparing the genetically modified organisms have required much time and labor, making it the rate-limiting step but CRISPR/Cas9 technology appearance has changed this difficulty. Although reports on CRISPR/Cas9 technology such as genome editing and CRISPR interference (CRISPRi) in eukaryotes increased, those in prokaryotes especially in Staphylococci were limited. Thus, its potential in the bacteriology remains unexplored. This is attributed to ecological difference between eukaryotes and prokaryotes. Here, we constructed a novel CRISPRi plasmid vector, pBACi for Staphylococcus aureus. The transformation efficiency of S. aureus was ~104 CFU/μg DNA using a vector extracted from dcm negative, which encoded one of DNA modification genes, E. coli. Further, pBACi was introduced into various clinical isolates including that not accepting the conventional temperature-sensitive vector. dcas9 in the vector was expressed throughout the growth phases of S. aureus and this vector decreased various gene mRNA expressions based on the crRNA targeting sequences and altered the knockdown strains’ phenotypes. The targeted genes included various virulence and antibiotic resistant genes. Bioinformatics suggest this vector can be introduced into wide range of low-GC Gram-positive bacteria. Because this new CRISPR/Cas9-based vector can easily prepare knockdown strains, we believe the novel vector will facilitate the characterization of the function of genes from S. aureus and other Gram-positive bacteria.

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“…However, expression of this polymer tends to be suppressed in methicillin-resistant S. aureus strains in favor of fibronectin binding proteins [47]. Staphylococci that predominantly inhabit environments with high shear stress or grow on medical devices produce primarily polysaccharide-dependent biofilms [48, 49], whereas those that interact more directly with host tissues may benefit from protein-based biofilms that allow them to incorporate fibrin or other host proteins into a protective shield [50]. It is tempting to speculate that Yptb strains retain multiple biofilm strategies that provide flexibility according to changing environments, whereas Y. pestis may have jettisoned alternative biofilm strategies to Hms-ECM as it adapted to its restricted lifestyle of flea-rodent transmission.…”

Section: Discussionmentioning

confidence: 99%

A trimeric autotransporter enhances biofilm cohesiveness inYersinia pseudotuberculosisbut not inYersinia pestis

Calder

Christman

Hawkins

et al. 2020

Preprint

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15Cohesion of biofilms made by Yersinia pestis and Yersinia pseudotuberculosis (Yptb) has been 16 attributed solely to an extracellular polysaccharide matrix encoded by the hms genes (Hms-17 ECM). However, mutations in the Yptb BarA/UvrY/CsrB regulatory cascade enhance biofilm 18 stability without dramatically increasing Hms-ECM production. We found that treatment with 19 proteinase K enzyme effectively destabilized Yptb csrB mutant biofilms, suggesting that cell-cell 20 interactions might be mediated by protein adhesins or extracellular matrix proteins. We 21 identified an uncharacterized trimeric autotransporter lipoprotein (YPTB2394), repressed by 22 csrB, which has been referred to as YadE. Biofilms made by a yadE mutant strain were 23 extremely sensitive to mechanical disruption. Overexpression of yadE in wild-type Yptb 24 increased biofilm cohesion, similar to biofilms made by csrB or uvrY mutants. We found that the 25 Rcs signaling cascade, which represses Hms-ECM production, activated expression of yadE. The 26 yadE gene appears to be functional in Yptb but is a pseudogene in modern Y. pestis strains. 27 Expression of functional yadE in Y. pestis KIM6+ altered the production of Hms-ECM and 28 weakened biofilms made by these bacteria. This suggests that although the YadE autotransporter 29 protein increases Yptb biofilm stability, it may be incompatible with Hms-ECM production that 30 is essential for Y. pestis biofilm production in fleas. Inactivation of yadE in Y. pestis may be 31 another instance of selective gene loss in the evolution of flea-borne transmission by this species. 32 33 3 IMPORTANCE 34The evolution of Yersinia pestis from its Y. pseudotuberculosis (Yptb) ancestor involved gene 35 acquisition and gene losses, leading to differences in biofilm production. Characterizing the 36 unique biofilm features of both species may provide better understanding of how each adapts to 37 its specific niches. This study identifies a trimeric autotransporter YadE that promotes biofilm 38 stability of Yptb but which has been inactivated in Y. pestis, likely because it is not compatible 39 with Hms polysaccharide that is crucial for biofilms inside fleas. We also reveal that the Rcs 40 signaling cascade, which represses Hms expression in Y. pestis, activates YadE in Yptb. The 41 ability of Yptb to use polysaccharide or YadE protein for cell-cell adhesion may help it produce 42 biofilms in different environments. 44Environmental persistence, host interaction, and transmission of Yersinia depend on 45 biofilms, which are tightly regulated by both transcriptional and post-transcriptional control 46 mechanisms [1, 2]. Arguably, the best studied Yersinia biofilms are those made by Y. pestis 47 while in the flea digestive tract that block the proventriculus and increase transmission to new 48 hosts during flea feeding. These biofilms require the HmsHFRS proteins to produce and export a 49 polysaccharide extracellular matrix of poly-ß-1,6-N-acetylglucosamine that is crucial in forming 50 and maintaining ba...

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A Novel Repressor of the ica Locus Discovered in Clinically Isolated Super-Biofilm-Elaborating Staphylococcus aureus

Cited by 31 publications

References 42 publications

Biofilm Formation by Staphylococcus aureus Clinical Isolates is Differentially Affected by Glucose and Sodium Chloride Supplemented Culture Media

Biofilm Formation by Staphylococcus aureus Clinical Isolates is Differentially Affected by Glucose and Sodium Chloride Supplemented Culture Media

Tailor-made gene silencing of Staphylococcus aureus clinical isolates by CRISPR interference

A trimeric autotransporter enhances biofilm cohesiveness inYersinia pseudotuberculosisbut not inYersinia pestis

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