Genotypic and Phenotypic Assessment of Hyaluronidase among Type Strains of a Select Group of Staphylococcal Species

Hyaluronidases degrade hyaluronic acid, a major polysaccharide of the extracellular matrix of tissues, and are considered important for virulence in a number of Gram-positive and -negative bacteria. The purpose of the present study was to determine the prevalence of hyaluronidase among clinical strains ofStaphylococcus aureusand among otherStaphylococcusspecies. Spent media and chromosomal DNA were assessed for hyaluronidase activity and the absence or presence of a hyaluronidase gene (hysA) by Southern analysis, respectively. AllS. aureusstrains examined exhibited at least one hybridizing band (half of the strains exhibited two or more hybridizing bands) when probed forhysAand all but three of these strains produced hyaluronidase. In contrast, none of the type strains of 19 other species exhibited either hyaluronidase activity or hybridizing bands when probed forhysA. These data support the hypothesis that among members of theStaphylococcusgenus only strains ofS. aureuspossess the enzyme hyaluronidase. This would suggest that hyaluronidase represents yet another potential virulence factor employed byS. aureusto cause disease and may represent a diagnostically important characteristic for distinguishingS. aureusfrom other members of this genus.

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sae is essential for expression of the staphylococcal adhesins Eap and Emp

Microbiology ◽

10.1099/mic.0.27902-0 ◽

2005 ◽

Vol 151 (6) ◽

pp. 1789-1800 ◽

Cited By ~ 56

Author(s):

Niamh Harraghy ◽

Jan Kormanec ◽

Christiane Wolz ◽

Dagmar Homerova ◽

Christiane Goerke ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Extracellular Matrix ◽

Virulence Factor ◽

Eukaryotic Cells ◽

Transcription Start Point ◽

Chronic Infections ◽

Transcription Start ◽

Regulatory Mutants ◽

Global Regulators ◽

Glucose Analysis

Eap and Emp are two Staphylococcus aureus adhesins initially described as extracellular matrix binding proteins. Eap has since emerged as being important in adherence to and invasion of eukaryotic cells, as well as being described as an immunomodulator and virulence factor in chronic infections. This paper describes the mapping of the transcription start point of the eap and emp promoters. Moreover, using reporter-gene assays and real-time PCR in defined regulatory mutants, environmental conditions and global regulators affecting expression of eap and emp were investigated. Marked differences were found in expression of eap and emp between strain Newman and the 8325 derivatives SH1000 and 8325-4. Moreover, both genes were repressed in the presence of glucose. Analysis of expression of both genes in various regulatory mutants revealed that sarA and agr were involved in their regulation, but the data suggested that there were additional regulators of both genes. In a sae mutant, expression of both genes was severely repressed. sae expression was also reduced in the presence of glucose, suggesting that repression of eap and emp in glucose-containing medium may, in part, be a consequence of a decrease in expression of sae.

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Investigation of the antibiotic resistance and biofilm formation of Staphylococcus aureus strains isolated from gangrenous mastitis of ewes

Acta Veterinaria Hungarica ◽

10.1556/avet.2012.016 ◽

2012 ◽

Vol 60 (2) ◽

pp. 189-197 ◽

Cited By ~ 7

Author(s):

Osman Tel ◽

Özkan Aslantaş ◽

Oktay Keskin ◽

Ebru Yilmaz ◽

Cemil Demir

Keyword(s):

Staphylococcus Aureus ◽

Antibiotic Resistance ◽

Virulence Factor ◽

Resistance Genes ◽

Biofilm Formation ◽

Antibiotic Resistance Genes ◽

Penicillin G ◽

Beta Lactamase ◽

Potential Virulence Factor ◽

Amoxicillin Clavulanic Acid

In this study,Staphylococcus aureusstrains (n = 110) isolated from seven ewe flocks in Sanliurfa, Turkey were screened for antibiotic resistance and biofilmforming ability as well as for genes associated with antibiotic resistance and biofilm-forming ability. All isolates were found to be susceptible to oxacillin, gentamicin, clindamycin, cefoxitin, tetracycline, vancomycin, amoxicillin-clavulanic acid, ciprofloxacin and sulphamethoxazole-trimethoprim. The percent proportions of strains resistant to penicillin G, ampicillin and erythromycin were 27.2% (n = 30), 25.4% (n = 28) and 6.3% (n = 7), respectively. Regarding the antibiotic resistance genes, 32 (29%) isolates carried theblaZ and 8 (7.2%) theermC gene. Other resistance genes were not detected in the isolates. All isolates showed biofilm-forming ability on Congo red agar (CRA), while 108 (98.18%) and 101 (91.81%) of them were identified as biofilm producers by the use of standard tube (ST) and microplate (MP) methods, respectively. All isolates carried theicaA andicaD genes but none of them harboured thebapgene. The results demonstrated thatS. aureusisolates from gangrenous mastitis were mainly resistant to penicillins (which are susceptible to the staphylococcal beta-lactamase enzyme), and less frequently to erythromycin. Furthermore, all of theS. aureusisolates produced biofilm which was considered a potential virulence factor in the pathogenesis of staphylococcal mastitis.

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Extra-Chromosomal DNA Sequencing Reveals Episomal Prophages Capable of Impacting Virulence Factor Expression in Staphylococcus aureus

Frontiers in Microbiology ◽

10.3389/fmicb.2018.01406 ◽

2018 ◽

Vol 9 ◽

Cited By ~ 7

Author(s):

Douglas R. Deutsch ◽

Bryan Utter ◽

Kathleen J. Verratti ◽

Heike Sichtig ◽

Luke J. Tallon ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Dna Sequencing ◽

Virulence Factor ◽

Chromosomal Dna ◽

Factor Expression

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Varied Contribution of Phospholipid Shedding From Membrane to Daptomycin Tolerance in Staphylococcus aureus

Frontiers in Molecular Biosciences ◽

10.3389/fmolb.2021.679949 ◽

2021 ◽

Vol 8 ◽

Author(s):

Tianwei Shen ◽

Kelly M. Hines ◽

Nathaniel K. Ashford ◽

Brian J. Werth ◽

Libin Xu

Keyword(s):

Staphylococcus Aureus ◽

Wild Type ◽

Bacterial Membranes ◽

Strain Pair ◽

Time Kill ◽

The Impact ◽

Growth Controls ◽

Type Strains ◽

Spent Media

It has been suggested that daptomycin can be inactivated by lipids released by Staphylococcus aureus and that this effect is antagonized by phenol soluble modulins (PSMs), which bind to the shed lipids. PSM production is regulated by the Agr system, and others have shown that loss of the Agr function enhances S. aureus survival in the presence of daptomycin. Here we assessed the impact of Agr function on daptomycin activity and lipid metabolism under various conditions. Daptomycin activity was evaluated against three sets of isogenic strain series with wild-type or dysfunctional Agr using static daptomycin time-kills over 24 h and against one strain pair using in vitro pharmacokinetic/pharmacodynamic (PK/PD) models simulating clinical daptomycin exposure for 48 h. We performed comprehensive lipidomics on bacterial membranes and the spent media to correlate lipid shedding with survival. In static time-kill experiments, two agr-deficient strains (SH1000- and USA300 LAC ΔagrA) showed improved survival for 8 h compared with their corresponding wild-type strains as seen in previous studies, but this difference did not persist for 24 h. However, four other agr-deficient strains (SH1001 and JE2 agr KOs) did not demonstrate improved survival compared to isogenic wild-type strains at any time in the time-kills. Lipidomics analysis of SH1000, SH1001, and SH1000- strains showed daptomycin exposure increased lipid shedding compared to growth controls in all strains with phosphatidylglycerols (PGs), lysylPGs and cardiolipins predominating. In the cell pellets, PGs and lysylPGs decreased but cardiolipins were unchanged with daptomycin exposure. The shed lipid profiles in SH1001 and SH1000- were similar, suggesting that the inability to resist daptomycin by SH1001 was not because of differences in lipid shedding. In the PK/PD model, the agr mutant SH1000- strain did not show improved survival relative to SH1000 either. In conclusion, inactivation of daptomycin by shed lipids may be dependent on genetic background, the specific agr mutations, or the techniques used to generate these KOs rather than the overall function of the Agr system, and its contribution to daptomycin tolerance seems to be varied, transient, and growth-condition dependent.

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Identification, Cloning, and Initial Characterization of rot, a Locus Encoding a Regulator of Virulence Factor Expression in Staphylococcus aureus

Journal of Bacteriology ◽

10.1128/jb.182.11.3197-3203.2000 ◽

2000 ◽

Vol 182 (11) ◽

pp. 3197-3203 ◽

Cited By ~ 126

Author(s):

Peter J. McNamara ◽

Kathy C. Milligan-Monroe ◽

Shirin Khalili ◽

Richard A. Proctor

Keyword(s):

Staphylococcus Aureus ◽

Virulence Factor ◽

Insertion Site ◽

Mutant Gene ◽

Northern Analysis ◽

Alpha Toxin ◽

Wild Type ◽

Chromosomal Dna ◽

Erythromycin Resistance ◽

Mutant Strains

ABSTRACT A chromosomal insertion of transposon Tn917 partially restores the expression of protease and alpha-toxin activities to PM466, a genetically defined agr-null derivative of the wild-type Staphylococcus aureus strain RN6390. In co-transduction experiments, transposon-encoded erythromycin resistance and a protease- and alpha-toxin-positive phenotype are transferred at high frequency from mutant strains to agr-null strains ofS. aureus. Southern analysis of chromosomal DNA and sequence analysis of DNA flanking the Tn917 insertion site in mutant strains revealed that the transposon interrupted a 498-bp open reading frame (ORF). Similarity searches using a conceptual translation of the ORF identified a region of homology to the known staphylococcal global regulators AgrA and SarA. To verify that the mutant allele conferred the observed phenotype, a wild-type allele of the mutant gene was introduced into the genome of a mutant strain by homologous recombination. The resulting isolates had a restoredagr-null phenotype. Virulence factor gene expression in mutant, restored mutant, and wild-type strains was quantified by measuring alpha-toxin activity in culture supernatant fluids and by Northern analysis of the alpha-toxin transcript. We named this ORFrot (for repressor of toxins) (GenBank accession no.AF189239 ) because of the activity associated withrot::Tn917 mutant strains.

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Staphylococcus aureus Hyaluronidase Is a CodY-Regulated Virulence Factor

Infection and Immunity ◽

10.1128/iai.01710-14 ◽

2014 ◽

Vol 82 (10) ◽

pp. 4253-4264 ◽

Cited By ~ 38

Author(s):

Carolyn B. Ibberson ◽

Crystal L. Jones ◽

Shweta Singh ◽

Matthew C. Wise ◽

Mark E. Hart ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Hyaluronic Acid ◽

Virulence Factor ◽

Bacterial Infections ◽

Lung Pathology ◽

Diverse Range ◽

Content Type ◽

Global Regulators ◽

Protein Levels ◽

Unit Reduction

ABSTRACTStaphylococcus aureusis a Gram-positive pathogen that causes a diverse range of bacterial infections. InvasiveS. aureusstrains secrete an extensive arsenal of hemolysins, immunomodulators, and exoenzymes to cause disease. Our studies have focused on the secreted enzyme hyaluronidase (HysA), which cleaves the hyaluronic acid polymer at the β-1,4 glycosidic bond. In the study described in this report, we have investigated the regulation and contribution of this enzyme toS. aureuspathogenesis. Using the Nebraska Transposon Mutant Library (NTML), we identified eight insertions that modulate extracellular levels of HysA activity. Insertions in thesigBoperon, as well as in genes encoding the global regulators SarA and CodY, significantly increased HysA protein levels and activity. By altering the availability of branched-chain amino acids, we further demonstrated CodY-dependent repression of HysA activity. Additionally, through mutation of the CodY binding box upstream ofhysA, the repression of HysA production was lost, suggesting that CodY is a direct repressor ofhysAexpression. To determine whether HysA is a virulence factor, a ΔhysAmutant of a community-associated methicillin-resistantS. aureus(CA-MRSA) USA300 strain was constructed and found to be attenuated in a neutropenic, murine model of pulmonary infection. Mice infected with this mutant strain exhibited a 4-log-unit reduction in bacterial burden in their lungs, as well as reduced lung pathology and increased levels of pulmonary hyaluronic acid, compared to mice infected with the wild-type, parent strain. Taken together, these results indicate thatS. aureushyaluronidase is a CodY-regulated virulence factor.

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