Research paperMolecular analysis of accessory gene regulator functionality and virulence genes in Staphylococcus aureus derived from pediatric wound infections
Introduction
Staphylococcus aureus is an ever-present opportunistic pathogen that can cause a variety of diseases. The severity of S. aureus-associated infections ranges from benign localized skin abscesses to life-threatening diseases, such as arthritis, osteomyelitis, and endocarditis (Francois et al., 2006; von Eiff et al., 2004). In recent decades, methicillin-resistant S. aureus (MRSA) strains have emerged as a predominant cause of invasive diseases, namely skin and soft tissues, as well as musculoskeletal infections in children (Kaushik and Kest, 2018). This bacterium is one of the most dominant commensals on human skin and nasal mucosa and can express a multitude of virulence factors, such as surface adhesins, enterotoxins and hemolysins which are central in the development of disease. (Kassam et al., 2017; Stevens et al., 2017). The synchronized expression of these virulence determinants is tightly controlled by the cumulative action of several regulatory elements, such as the accessory gene regulator (agr), staphylococcal accessory regulator A (sarA), and the alternative sigma factor B (σB) (Manna and Cheung, 2001).
The Agr system plays a central role in the growth-phase dependent modulation of virulence gene expression (Bronner et al., 2004; Sakoulas et al., 2003a). The agr operon is an autocatalytic system controlled in a cell density-dependent fashion through the production and sensing of auto-inducing peptides (AIP). At high cell density, the Agr system increases the production of many secreted virulence factors, including Toxic shock syndrome toxin −1 (TSST-1), delta-hemolysin and exfoliative toxins A and B (ETA and ETB). In contrast, Agr decreases the expression of several colonization factors such as fibronectin binding proteins, important in adhesion and biofilm formation (Li et al., 2018). The agr locus consists of two distinct transcripts, RNAII and RNAIII, which are transcribed by two promoters, P2 and P3 respectively. The activation of P2 induces the expression of the components involved in cell-to-cell quorum-sensing communication (AgrBDCA) (Bibalan et al., 2014). Both AgrB and AgrD function to process and secrete the auto-inducing peptide (AIP), which acts as the chemical messenger critical for Agr activity (Wang et al., 2014). Upon reaching a critical density, AIPs interact with the sensor kinase, AgrC which promotes phosphorylation of the DNA binding response regulator AgrA. Phosphorylated AgrA undergoes a conformational change permitting interaction and binding to the intergenic region between P2 and P3 facilitating their expression. P3 activation leads to the expression of RNAIII, the effector of target gene regulation (Novick and Geisinger, 2008).
Several studies have demonstrated a correlation between agr types and particular diseases. For example phylogenetic group AF1 (agr group IV) strains are closely related to generalized exfoliative syndromes and bullous impetigo whereas endocarditis is mainly caused by phylogenetic group AF2 (agr groups II and I) strains (Jarraud et al., 2002). In addition, it has been suggested that agr group III and IV strains are associated with toxic shock syndrome (Gomes et al., 2005). To the best of our knowledge, there is no published study evaluating Agr functionality among Iranian S. aureus isolates. The present study was conducted to determine dominant Agr types, Agr activity and presence of specific virulence genes in S. aureus isolates derived from pediatric wound infections.
Section snippets
Bacterial isolation and identification
In the present study, 48 S. aureus isolates were collected from skin and soft tissue infections (SSTIs) of pediatric patients referred to the Children's Medical Center Hospital Tehran, Iran over one year from April 2017–2018. The School of Medicine, Shahid Beheshti University of Medical Sciences ethics committee approved this study (IR.SBMU.MSP.REC.1395.369). The isolates were identified as S. aureus according to phenotypic (colonial morphology and Gram-stain), biochemical (catalase,
Bacterial strains and antimicrobial resistance profiles
In this study, 48 S. aureus clinical isolates were collected from pediatric SSTIs from children aged between 1 day and 14 years. 14 (29.2%) samples were collected from patients admitted to the in infectious disease ward, 9 (18.8%) from post-surgery ward, 15 (31.2%) from infants, 6 (12.5%) from OPD, 3 (6.3%) from in-patient ward, 2 (4.2%) from emergency cases, and 3 (6.3%) from gastrointestinal, 2 (4.2%) from neurosurgery, 2 (4.2%) from intensive care unit, 1 (2.1%) from coronary intensive care
Discussion
The present study was conducted to evaluate the activity of the Agr system among clinical isolates of S. aureus derived from pediatric SSTIs. In addition, the association between Agr activity and the presence of several virulence determinants and antibiotic susceptibility was examined. Several techniques can be used to determine Agr function including the CAMP synergistic haemolysis assay, the Vesicle Lysis Test (VLT) and qRT-PCR detection of RNAIII.
Agr activity is traditionally evaluated using
Conclusion
In the present study, the data revealed that there was no significant correlation between Agr activity and the ability to cause wound infections by S. aureus strains.
Acknowledgments
The present article is financially supported by ‘Research Department of the School of Medicine Shahid Beheshti University of Medical Science’ (Grant No 12200). Special thanks to Dr. Mohammad Emaneini and Dr. Ruth Massey for all cooperation and guidance.
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