Impact of agr dysfunction on virulence profiles and infections associated with a novel methicillin-resistant Staphylococcus aureus (MRSA) variant of the lineage ST1-SCCmec IV

BMC Microbiology

Agnes Marie Sá Figueiredo²  José Nelson dos Santos Silva Couceiro²  Sérgio Eduardo Longo Fracalanzza²  Marco Antônio Américo²  Ana Maria de Amorim Ferreira¹  Bruno de Sousa Moraes²  Raquel Rodrigues Souza²  Fabienne Antunes Ferreira² 

[1]Departmento de Virologia, Universidade Federal do Rio de Janeiro, Instituto de Microbiologia Paulo de Góes Rio de Janeiro, Rio de Janeiro, RJ, Brazil

[2]Departamento de Microbiologia Médica, Instituto de Microbiologia Paulo de Góes, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brazil

关键词: Virulence factors;    Biofilm;    agr;    USA400;    ST1-SCCmecIV;    MRSA;   

Others  :  1143767
DOI  :  10.1186/1471-2180-13-93

 received in 2012-11-05, accepted in 2013-04-22,  发布年份 2013

【 摘 要 】

Background

A novel variant of the ST1-SCCmecIV methicillin-resistant Staphylococcus aureus (MRSA) lineage, mostly associated with nosocomial bloodstream infections (BSI), has emerged in Rio de Janeiro. Bacterial biofilm has been considered a major virulence factor in central venous catheter-associated BSI. The mechanisms involved in biofilm formation/accumulation are multifactorial and complex. Studies have suggested that biofilm production was affected in vitro and vivo for agr-null mutants of S. aureus.

Results

The impact of naturally occurring inhibition of agr signaling on virulence profiles and infections associated with the ST1 variant was investigated. agr dysfunction was detected in a significant percentage (13%) of the isolates with concomitant increase in biofilm accumulation in vitro and in vivo, and enhanced ability to adhere to and invade airway cells. The biofilm formed by these ST1 isolates was ica-independent and proteinaceous in nature. In fact, the improved colonization properties were paralleled by an increased expression of the biofilm-associated genes fnbA, spa and sasG. The transcription of sarA, a positive regulator of agr, was two-times reduced for the agr-dysfunctional MRSA. Remarkably, the agr inhibition was genetically stable. Indeed, agr-dysfunctional isolates succeed to colonize and cause both acute and chronic infections in hospitalized patients, and also to effectively accumulate biofilm in a mouse subcutaneous catheter implant model.

Conclusion

The ability of agr-dysfunctional isolates to cause infections in humans and to form biofilm in the animal model suggests that therapeutic approaches based on agr-inactivation strategies are unlikely to be effective in controlling human-device infections caused by ST1 isolates. The increased biofilm accumulation associated with the acquisition of multiple antimicrobial resistant traits might have influenced (at least in part) the expansion of this USA400 related clone in our hospitals.

【 授权许可】

   
2013 Ferreira et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Centers for Disease Control and Prevention: Community-acquired methicillin–resistant staphylococcus aureus infections-Michigan. MMWR Morb Mortal Wkly Rep 1981, 30:185-187.
• [2]Centers for Disease Control and Prevention: Four pediatric deaths from community-acquired methicillin-resistant staphylococcus aureus: Minnesota and north Dakota, 1997–1999. MMWR Morb Mortal Wkly Rep 1999, 48:707-710.
• [3]Herold BC, Immergluck LC, Maranan MC, Lauderdale DS, Gaskin RE, Boyle-Vavra S, Leitch CD, Daum RS: Community-acquired methicillin-resistant staphylococcus aureus in children with no identified predisposing risk. JAMA 1998, 279:593-598.
• [4]David MZ, Daum RS: Community-associated methicillin-resistant staphylococcus aureus: epidemiology and clinical consequences of an emerging epidemic. Clin Microbiol Rev 2010, 23:616-687.
• [5]Skov R, Christiansen K, Dancer SJ, Daum RS, Dryden M, Huang YC, Lowy FD: Update on the prevention and control of community-acquired methicillin-resistant staphylococcus aureus (CA-MRSA). Int J Antimicrob Agents 2012, 39:193-200.
• [6]Diep BA, Chan L, Tattevin P, Kajikawa O, Martin TR, Basuino L, Mai TT, Marbach H, Braughton KR, Whitney AR, Gardner DJ, Fan X, Tseng CW, Liu GY, Badiou C, Etienne J, Lina G, Matthay MA, DeLeo FR, Chambers HF: Polymorphonuclear leukocytes mediate staphylococcus aureus Panton-valentine leukocidin-induced lung inflammation and injury. Proc Natl Acad Sci USA 2010, 107:5587-5592.
• [7]Löffler B, Hussain M, Grundmeier M, Brück M, Holzinger D, Varga G, Roth J, Kahl BC, Proctor RA, Peters G: Staphylococcus aureus Panton-valentine leukocidin is a very potent cytotoxic factor for human neutrophils. PLoS Pathog 2010, 6:e1000715.
• [8]Otto M: A MRSA-terious enemy among us: end of the PVL controversy? Nat Med 2011, 17:169-170.
• [9]Hudson LO, Murphy CR, Spratt BG, Enright MC, Terpstra L, Gombosev A, Hannah P, Mikhail L, Alexander R, Moore DF, Huang SS: Differences in methicillin-resistant staphylococcus aureus (MRSA) strains isolated from pediatric and adult from hospitals in a large California county. J Clin Microbiol 2011, 50:573-579.
• [10]David MZ, Rudolph KM, Hennessy TW, Boyle-Vavra S, Daum RS: Molecular epidemiology of methicillin-resistant staphylococcus aureus rural southwestern Alaska. Emerg Infect Dis 2008, 14:1693-1699.
• [11]Golding GR, Levett PN, McDonald RR, Irvine J, Quinn B, Nsungu M, Woods S, Khan M, Ofner-Agostini M, Mulvey MR, Northern Antibiotic Resistance Partnership: High rates of staphylococcus aureus USA400 infection, northern Canada. Emerg Infect Dis 2011, 17:722-725.
• [12]Silva-Carvalho MC, Bonelli RR, Souza RR, Moreira S, dos Santos LC, de Souza Conceição M, de Mello Junior SJ, Carballido JM, Vieira VV, Teixeira LA, Sá Figueiredo AM: Emergence of multiresistant variants of the community-acquired methicillin-resistant staphylococcus aureus lineage ST1-SCCmecIV in 2 hospitals in Rio de Janeiro, brazil. Diagn Microbiol Infect Dis 2009, 65:300-305.
• [13]Shukla SK, Karow ME, Brady JM, Stemper ME, Kislow J, Moore N, Wroblewski K, Chyou PH, Warshauer DM, Reed KD, Lynfield R, Schwan WR: Virulence genes and genotypic associations in nasal carriage, community-associated methicillin-susceptible and methicillin-resistant USA400 staphylococcus aureus isolates. J Clin Microbiol 2010, 48:3582-3592.
• [14]Amaral MM, Coelho LR, Flores RP, Souza RR, Silva-Carvalho MC, Teixeira LA, Ferreira-Carvalho BT, Figueiredo AM: The predominant variant of the Brazilian epidemic clonal complex of methicillin-resistant staphylococcus aureus has an enhanced ability to produce biofilm and to adhere to and invade airway epithelial cells. J Infect Dis 2005, 192:801-810.
• [15]Datta R, Huang SS: Risk of infection and death due to methicillin-resistant staphylococcus aureus in long-term carriers. Clin Infec Dis 2008, 47:176-181.
• [16]Sinha B, Herrmann M: Mechanism and consequences of invasion of endothelial cells by staphylococcus aureus. Thromb Haemost 2005, 94:266-277.
• [17]Merino N, Toledo-Arana A, Vergara-Irigaray M, Valle J, Solano C, Calvo E, Lopez JA, Foster TJ, Penadés JR, Lasa I: Protein a-mediated multicellular behavior in staphylococcus aureus. J Bacteriol 2009, 191:832-843.
• [18]Geoghegan JA, Corrigan RM, Gruszka DT, Speziale P, O´Gara JP, Potts JR, Foster TJ: Role of surface protein SasG in biofilm formation in stapylococcus aureus. J Bacteriol 2010, 192:5663-5673.
• [19]Houston P, Rowe SE, Pozzi C, Waters EM, O´Gara JP: Essential role for the major autolysin in the fibronectin-binding protein-mediated staphylococcus aureus biofilm phenotype. Infect Immun 2011, 79:1153-1165.
• [20]Kiedrowski MR, Kavanaugh JS, Malone CL, Mootz JM, Voyich JM, Smeltzer MS, Bayles KW, Horswill AR: Nuclease modulates biofilm formation in community-associated methicillin-resistant staphylococcus aureus. PLoS One 2011, 6:e26714.
• [21]Bronner S, Monteil H, Prévost G: Regulation of virulence determinants in staphylococcus aureus: complexity and applications. FEMS Microbiol Rev 2004, 28:183-200.
• [22]Novick RP, Ross HF, Figueiredo AMS, Abramochkin G, Muir T: Activation and inhibition of staphylococcal Agr system. Science 2000, 287:391a.
• [23]Mayville P, Ji G, Beavis R, Yang H, Goger M, Novick RP, Muir TW: Structure-activity analysis of synthetic autoinducing thiolactone peptides from staphylococcus aureus responsible for virulence. Proc Natl Acad Sci USA 1999, 96:1218-1223.
• [24]Balaban N, Cirioni O, Giacometti A, Ghiselli R, Braunstein JB, Silvestri C, Mocchegiani F, Saba V, Scalise G: Treatment of staphylococcus aureus biofilm infection by the quorum-sensing inhibitor RIP. Antimicrob Agents Chemother 2007, 51:2226-2229.
• [25]Lopez-Leban F, Kiran MD, Wolcott R, Balaban N: Molecular mechanisms of RIP, an effective inhibitor of chronic infections. Int J Artif Organs 2010, 33:582-589.
• [26]Shenkman B, Varon D, Tamarin I, Dardik R, Peisachov M, Savion N, Rubinstein R: Role of agr (RNAIII) in staphylococcus aureus adherence to fibrinogen, fibronectin, platelets and endothelial cells under static and flow conditions. J Med Microbiol 2002, 51:747-754.
• [27]Coelho LR, Souza RR, Ferreira FA, Guimarães MA, Ferreira-Carvalho BT, Figueiredo AMS: agr RNAIII divergently regulates glucose-induced biofilm formation in clinical isolates of staphylococcus aureus. Microbiology 2008, 154:3480-3490.
• [28]Ferreira FA, Souza RR, Bonelli RR, Américo MA, Fracalanzza SEL, Figueiredo AMS: Comparison of in vitro and in vivo systems to study ica-independent staphylococcus aureus biofilm. J Microbiol Methods 2012, 88:393-398.
• [29]Zautner AE, Krause M, Stropahl G, Holtfreter S, Frickmann H, Maletzki C, Kreikemeyer B, Pau HW, Podbielski A: Intracellular persisting staphylococcus aureus is the major pathogen in recurrent tonsillitis. PLoS One 2010, 5:e9452.
• [30]Trotonda MP, Tamber S, Memmi G, Cheung AL: MgrA represses biofilm formation in staphylococcus aureus. Infect Immun 2008, 76:5645-5654.
• [31]Kaito C, Saito Y, Nagano G, Ikuo M, Omae Y, Hanada Y, Han X, Kuwahara-Arai K, Hishinuma T, Baba T, Ito T, Hiramatsu K, Sekimizu K: Transcription and translation products of the cytolysin gene psm-mec on the mobile genetic element SCCmec regulate staphylococcus aureus regulation. PLoS Pathog 2011, 7:e1001267.
• [32]Joshi SG, Paff M, Friedman G, Fridman G, Fridman A, Brooks AD: Control of methicillin-resistant staphylococcus aureus planktonic form and biofilms: a biocidal efficacy study of nonthermal dielectric-barrier discharge plasma. Am J Infect Control 2010, 38:293-301.
• [33]O´Neill E, Pozzi C, Houston P, Humphreys H, Robinson DA, Loughman A, Foster TJ, O'Gara JP: A novel staphylococcus aureus biofilm phenotype mediated by the fibronectin-binding proteins, FnBPA and FnBPB. J Bacteriol 2008, 190:3835-3850.
• [34]Vergara-Irigaray M, Valle J, Merino N, Latasa C, García B, Ruiz de Los Mozos I, Solano C, Toledo-Arana A, Penadés JR, Lasa I: Relevant role of fibronectin-binding proteins in staphylococcus aureus biofilm-associated foreign-body infections. Infect Immun 2009, 77:3978-3991.
• [35]Lauderdale KJ, Boles BR, Cheung AL, Horswill AR: Interconnections between sigma B, agr, and proteolytic activity in staphylococcus aureus biofilm maturation. Infect Immun 2009, 7:1623-1635.
• [36]Wolz C, Pöhlmann-Dietze P, Steinhuber A, Chien YT, Manna A, van Wamel W, Cheung A: Agr-independent regulation of fibronectin-binding protein(s) by the regulatory locus sar in staphylococcus aureus. Mol Microbiol 2000, 36:230-243.
• [37]Bartlett AH, Foster TJ, Hayashida A, Park PW: Alpha-toxin facilitates the generation of CXC chemokine gradients and stimulates neutrophil homing in Staphylococcus aureus pneumonia. J Infect Dis 2008, 198:1529-1535.
• [38]Bubeck Wardenburg J, Bae T, Otto M, DeLeo FR, Schneewind O: Poring over pores: alpha-hemolysin and Panton-valentine leukocidin in staphylococcus aureus pneumonia. Nat Med 2007, 13:1405-1406.
• [39]Wang R, Braughton KR, Kretschmer D, Bach TH, Queck SY, Li M, Kennedy AD, Dorward DW, Klebanoff SJ, Peschel A, DeLeo FR, Otto M: Identification of novel cytolytic peptides as key virulence determinants for community-associated MRSA. Nat Med 2007, 13:1510-1514.
• [40]Wright JS 3rd, Jin R, Novick RP: Transient interference with staphylococcal quorum sensing blocks abscess formation. Proc Natl Acad Sci USA 2005, 102:1691-1696.
• [41]Traber KE, Lee E, Benson S, Corrigan R, Cantera M, Shopsin B, Novick RP: agr function in clinical staphylococcus aureus isolates. Microbiology 2008, 154:2265-2274.
• [42]Park SY, Chong YP, Park HJ, Park KH, Moon SM, Jeong JY, Kim MN, Kim SH, Lee SO, Choi SH, Woo JH, Kim YS: agr dysfunction and persistent methicillin-resistant staphylococcus aureus bacteremia in patients with removed eradicable foci. Infection 2013, 41:111-119.
• [43]Falkow S: Bacterial entry into eukaryotic cells. Cell 1991, 65:1099-1102.
• [44]Garzoni C, François P, Huyghe A, Couzinet S, Tapparel C, Charbonnier Y, Renzoni A, Lucchini S, Lew DP, Vaudaux P, Kelley WL, Schrenzel J: A global view of staphylococcus aureus whole genome expression upon internalization in human epithelial cells. BMC Genomics 2007, 8:171. BioMed Central Full Text
• [45]Wesson CA, Liou LE, Todd KM, Bohach GA, Trumble WR, Bayles KW: Staphylococcus aureus Agr and Sar global regulators influence internalization and induction of apoptosis. Infect Immun 1998, 66:5238-5243.
• [46]Pozzi C, Waters EM, Rudkin JK, Schaeffer CR, Lohan AJ, Tong P, Loftus BJ, Pier GB, Fey PD, Massey RC, O'Gara JP: Methicillin resistance alters the biofilm phenotype and attenuates virulence in staphylococcus aureus device-associated infections. PLoS Pathog 2012, 8:e1002626.
• [47]Rudkin JK, Edwards AM, Bowden MG, Brown EL, Pozzi C, Waters EM, Chan WC, Williams P, O'Gara JP, Massey RC: Methicillin resistance reduces the virulence of healthcare-associated methicillin-resistant staphylococcus aureus by interfering with the agr quorum sensing system. J Infect Dis 2012, 205:798-806.
• [48]Morfeldt E, Tegmark K, Arvidson S: Transcriptional control of the agr-dependent virulence gene regulator, RNAIII, in staphylococcus aureus. Mol Microbiol 1996, 21:1227-1237.
• [49]Beenken KE, Mrak LN, Griffin LM, Zielinska AK, Shaw LN, Rice KC, Horswill AR, Bayles KW, Smeltzer MS: Epistatic relationships between sarA and agr in staphylococcus aureus biofilm formation. PLoS One 2010, 5:e10790.
• [50]CLSI: Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; approved standard. 9th edition. Wayne, PA: CLSI document M07-A9. Clinical and Laboratory Standards Institute; 2012.
• [51]De Jonge BLM, De Lencastre H, Tomasz A: Supression of autolysis and cell wall turnover in heterogeneous Tn551 mutants of a methicillin-resistant staphylococcus aureus strain. J Bacteriology 1991, 173:1105-1110.
• [52]Adhikari RP, Arvidson S, Novick RP: A nonsense mutation in agrA accounts for the defect in agr expression and the avirulence of staphylococcus 8325–4 traP::kan. Infect Immun 2007, 75:4534-4540.
• [53]Li M, Cheung GYC, Hu J, Wang D, Joo H, De Leo FR, Otto M: Comparative analysis of virulence and toxin expression of global community-associated MRSA strains. J Infect Dis 2010, 202:1866-1876.
• [54]Oliveira DC, de Lencastre H: Multiplex PCR strategy for rapid identification of structural types and variants of the mec element in methicillin-resistant staphylococcus aureus. Antimicrob Agents Chemother 2002, 46:2155-2161.
• [55]Dunn OJ: Basic statistics: a primer for the biomedical sciences. New York: John Wiley & Sons; 1964.