First Genome-Based Characterisation and Staphylococcal Enterotoxin Production Ability of Methicillin-Susceptible and Methicillin-Resistant Staphylococcus aureus Strains Isolated from Ready-to-Eat Foods in Algiers (Algeria)

Abstract

Staphylococcus aureus is a pathogenic microorganism of humans and animals, able to cause foodborne intoxication due to the production of staphylococcal enterotoxins (SEs) and to resist antibiotic treatment as in the case of methicillin-resistant S. aureus (MRSA). In this study, we performed a genomic characterisation of 12 genetically diverse S. aureus strains isolated from ready-to-eat foods in Algiers (Algeria). Moreover, their ability to produce some classical and new staphylococcal enterotoxins (SEs) was investigated. The 12 S. aureus strains resulted to belong to nine known sequence types (STs) and to the novel ST7199 and ST7200. Furthermore, S. aureus SA46 was assigned to the European clone MRSA-ST80-SCCmec-IV. The 12 strains showed a wide endowment of se and sel (staphylococcal enterotoxin-like toxin) genes (sea, seb, sed, seg, seh, sei, selj, sek, sem, sen, seo, seq, ser, selu2, selw, selx, sey, sel30; ψent1-ψent2), including variants and pseudogenes, and harboured the enterotoxin gene cluster (egc) types 1 and 5. Additionally, they produced various amounts of SEA (64.54-345.02 ng/mL), SEB (2871.28-14739.17 ng/mL), SED (322.70-398.94 ng/mL), SEH (not detectable-239.48 ng/mL), and SER (36,720.10-63,176.06 ng/mL) depending on their genotypes. The genetic determinants related to their phenotypic resistance to β-lactams (blaZ, mecA), ofloxacin (gyrA-S84L), erythromycin (ermB), lincomycin (lmrS), kanamycin (aph(3')-III, ant(6)-I), and tetracyclin (tet(L), tet(38)) were also detected. A plethora of virulence-related genes, including major virulence genes such as the tst gene, determinant for the toxic shock syndrome toxin-1, and the lukF-PV and lukS-PV genes, encoding the panton-valentine leukocidin (PVL), were present in the S. aureus strains, highlighting their pathogenic potential. Furthermore, a phylogenomic reconstruction including worldwide foodborne S. aureus showed a clear clustering based on ST and geographical origin rather than the source of isolation.

Keywords: Staphylococcus aureus; antibiotic resistance; food safety; genomics; lukF-PV and lukS-PV genes; methicillin-resistant Staphylococcus aureus (MRSA); panton-valentine leukocidin (PVL); ready-to-eat food; staphylococcal enterotoxins; virulence.

Figure 1

Phylogeny of S. aureus strains. Genome-based phylogenetic tree inferred by using maximum likelihood method RAxML with progressive refinement. S. argenteus MSHR1132 was used as outgroup. Support values are represented by scaled circles at each node. Tree is annotated with sequence type (ST), source of isolation (S), and geographical origin (O). See Supplementary Table S4 for detailed information.

Figure 2

sea and seh prophagic regions in S. aureus SA04 and SA51 genomes. (A) ΦMu3A region comprising sea gene in SA51 genome and φSA3 region comprising sea, sek, and seq genes in SA04 genome. se genes are shown in red; sak: staphylokinase; scn: complement inhibitor SCIN-A; spH: sphingomyelin phosphodiesterase; int: integrase; pAI: pathogenicity island family protein; ter: terminase; lukGH: bi-component leukocidin LukGH; groEL: chaperonin GroEL; * indicates pseudogenes. (B) φSa3ms region comprising seh gene in SA04 genome. act: acetyltransferase; safE: SafE family protein; cstR: persulfide-sensing transcriptional repressor CstR; cstA: persulfide response sulfurtransferase CstA; cstB: persulfide dioxygenase-sulfurtransferase CstB; * indicates pseudogenes.

Figure 3

pIB485-like plasmid in S. aureus SA20 genome. Enterotoxin genes ser and selj are represented in red. rep: replication protein; cadD: cadmium resistance transporter CadD; RNApol: RNA-directed DNA polymerase; rec: recombinase; IS6: IS6 family transposase; blaZ: beta-lactamase; blaR1: regulatory sensor/signal transducer BlaR1; blaI: beta-lactamase repressor BlaI; marR: MarR family transcriptional regulator; ox: oxidoreductase; * indicates pseudogenes.

Figure 4

Virulence determinants in S. aureus genomes. Heatmap of virulence determinants in S. aureus genomes. Presence and numbers of genes are indicated by green colour intensity; absence of gene was indicated by a white square; pseudogenes are indicated as red-striped squares. Heatmap was visualised by using heatmapper web server [43] with average linkage as clustering method and Euclidean distance measurement method.

Figure 5

seh relative gene expression. seh relative gene expression in S. aureus SA04 and SA46 at 5 h and 24 h of incubation in BHI + YE broth.