Expression of superantigens and the agr system in Staphylococcus epidermidis

Highlights

• The clinical coagulase-negative staphylococci strains are reservoirs of enterotoxin genes.

• Coagulase-negative staphylococci confirm the toxin-producing ability.

• The agr group II may be associated with enterotoxin C production by S. epidermidis.

Abstract

Infections with coagulase-negative staphylococci (CoNS) can involve the production of toxins such as superantigens, which contribute to tissue degradation and inflammatory immune responses. The accessory gene regulator (agr) quorum sensing system coordinates the expression of most S. aureus virulence factors. Therefore, the aim of this study was to investigate the expression of these superantigens and the presence of the agr locus in CoNS strains isolated from blood cultures. PCR was used to detect enterotoxin and agr genes and expression was analyzed by RT-PCR. Expression of the sea gene was observed in one S. epidermidis isolate andof sec-1 in two, seg and sei were expressed concomitantly in one isolate, and sei was expressed in another isolate. The agr group I was detected in S. epidermidis expressing the sea, seg and sei genes, whereas agr group II was detected in isolates expressing thesec-1 gene. The agr groups were only expressed in strains expressing thesec-1 gene. The results show that enterotoxin genes are highly frequent in CoNS isolated from clinical specimens and confirm the toxin-producing ability of these strains. The agr group II may be associated with enterotoxin C production by S. epidermidis, increasing the virulence of strains isolated from blood cultures and consequently the severity of sepsis caused by these organisms.

Introduction

Most species belonging to the genus Staphylococcus are coagulase-negative staphylococci (CoNS), i.e., they cannot produce the coagulase enzyme. Although CoNS are part of the human microbiota, they are considered opportunistic microorganisms because they take advantage of situations such as post-trauma tissue damage and the presence of foreign bodies to proliferate and spread to other tissues, developing a pathogenic behavior [1]. CoNS are the main cause of bacteremia in hospitals and their pathogenesis is complex, involving the production of a variety of virulence factors such as toxins [2], [3].

Staphylococcal toxins can contribute to tissue degradation and elicit anti-inflammatory immune responses [4]. Staphylococcal enterotoxins and toxic shock syndrome toxin 1 (TSST-1) are superantigens whose toxic effects can trigger the nonspecific proliferation of T cells through direct binding to major histocompatibility complex class II molecules and to the Vβ region of the T cell antigen receptor. Unlike normal antigen processing, they stimulate many T cells that overproduce cytokines such as interleukin 1 (IL-1), IL-2, interferon gamma (IFN-γ), and tumor necrosis factor alpha (TNF-α) [5].

During infection, toxin production by staphylococci is a multi-stage process that is coordinated by a complex system of communication between bacterial cells. This mechanism, called quorum sensing, allows bacteria to share information about cell density and to establish a phenotypic reaction according to the growth stage of the culture [6], [7]. Quorum sensing via the accessory gene regulator (agr) system is one of the main systems that coordinate staphylococcal virulence factors [8]. This system comprises promoters P2 and P3, which operate in opposite directions and produce transcripts RNAII and RNAIII, respectively. RNAIII is responsible for the gene transcription of a number of virulence factors, including extracellular toxins and enzymes and surface proteins [9]. The agr system is composed of four genes (agrA, agrB, agrC, and agrD) arranged in an operon. The products of these genes are proteins AgrA, AgrB, AgrC and AgrD, which are necessary for the function of the system. Proteins AgrB and AgrD combine to produce the autoinducing polypeptide (AIP), which is released into the extracellular medium. When the transmembrane protein AgrC detects AIP in the external environment, it phosphorylates AgrA that induces expression from P2 and P3. The final product of the agr locus is RNAIII, an mRNA that induces or inhibits toxin genes [7].

Three polymorphisms of the agr locus (agr group I, group II and group III) have been described in Staphylococcus epidermidis [10]. These groups show variations in the agrB, agrC and agrD genes and thus in AIP and AgrC, with AIP binding to a specific receptor for each allelic group. When AIP of one allele group binds to the AgrC receptor of another group, it does not produce intrinsic factors and thereby behaves as an antagonist. AIP is the only agonist to its own allele group and bacteria of one agr group can therefore affect the regulation of accessory proteins of bacteria from another agr group [7].

In view of the increasing incidence of CoNS infection, studies on associated virulence factors are important to better understand the toxigenic potential of these microorganisms. Therefore, the present study evaluated the occurrence and expression of superantigens and the role of the agr system in the production of virulence factors by CoNS species isolated from blood cultures of patients hospitalized in the University Hospital of the Botucatu Medical School.