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Characterization of enterotoxin A-producing Staphylococcus aureus.

ABSTRACT: Purpose:This study aims to characterize the wild-type staphylococcal enterotoxin A (SEA)-producing Staphylococcus aureus. Materials and methods:We identified 29 wild-type sea-positive S. aureus isolates from dairy and meat samples, as well as from patients, measured the amount of SEA produced under favorable cultivation conditions using enzyme-linked immunosorbent assay and sea mRNA transcriptional level and investigated the phage type as well as genetic diversity by means of pulsed-field gel electrophoresis and multilocus sequence typing. Results:Among 29 sea-positive isolates, 22 were from food sources (including one outbreak case) and seven from clinical patients. Five enterotoxin gene profiles, namely, sea (14 isolates), sea+sec (9 isolates), sea+seb (4 isolates), sea+seb+sec (1 isolate) and sea+seb+sed (1 isolate), were identified. Multilocus sequence typing generated sequence type (ST)1 (13 isolates), ST6 (5 isolates), ST59 (3 isolates), ST239 (3 isolates), ST5 (2 isolates), ST188 (2 isolate) and ST15 (1 isolate). The amount of SEA per 108 colony-forming unit (CFU) after 24 h of incubation was 1.1-33.5 (mean, 8.74; SD, 7.7) ng/108 CFU. The amount of SEA per hour incubation in the log growth phase was 0.1-12.0 (mean, 2.37; SD, 3.06) ng/108 CFU. Overall, 54.2% of SEA was produced in the log growth phase. Both the transcriptional level of sea mRNA and the amount of SEA in the log growth phase correlated well with the amount of SEA after 24 h of cultivation. Four isolates, namely, SA-212, SA-217, SA-340 and SA-341, were categorized to be of high SEA production (877-1,109 ng/mL, 24 h). The total amount of SEA was mainly based on the amount of SEA in 108 CFU, not the relatively fixed bacterial cell counts (21.1-43×108 CFU/mL). Seven isolates from patients all carried the ?Mu3A phage, whereas 21 of the 22 isolates from the environmental sources all carried the ?Sa3ms phage. Conclusion:The present study exhibits varied SEA production capacity of the wild sea-positive S. aureus strains. An apparent boundary in phage types between strains from the clinical samples and strains from the environment was also identified.

SUBMITTER: Chen D

PROVIDER: S-EPMC5905522 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Characterization of enterotoxin A-producing <i>Staphylococcus aureus</i>.

Chen Dong D Song Qifa Q Xu Zhaojun Z Zhang Dandan D

Infection and drug resistance 20180413

<h4>Purpose</h4>This study aims to characterize the wild-type staphylococcal enterotoxin A (SEA)-producing <i>Staphylococcus aureus</i>.<h4>Materials and methods</h4>We identified 29 wild-type <i>sea</i>-positive <i>S. aureus</i> isolates from dairy and meat samples, as well as from patients, measured the amount of SEA produced under favorable cultivation conditions using enzyme-linked immunosorbent assay and <i>sea</i> mRNA transcriptional level and investigated the phage type as well as geneti ...[more]

PMID: 29695922

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Project description:Staphylococcal enterotoxins are exotoxins produced by Staphylococcus aureus that possess emetic and superantigenic properties. Prior to this research there were six characterized enterotoxins, staphylococcal enterotoxin types A to E and H (referred to as SEA to SEE and SEH). Two new staphylococcal enterotoxin genes have been identified and designated seg and sei (staphylococcal enterotoxin types G and I, respectively). seg and sei consist of 777 and 729 nucleotides, respectively, encoding precursor proteins of 258 (SEG) and 242 (SEI) deduced amino acids. SEG and SEI have typical bacterial signal sequences that are cleaved to form toxins with 233 (SEG) and 218 (SEI, predicted) amino acids, corresponding to mature proteins of 27,043 Da (SEG) and 24,928 Da (SEI). Biological activities for SEG and SEI were determined with recombinant S. aureus strains. SEG and SEI elicited emetic responses in rhesus monkeys upon nasogastric administration and stimulated murine T-cell proliferation with the concomitant production of interleukin 2 (IL-2) and gamma interferon (IFN-gamma), as measured by cytokine enzyme-linked immunoassays. SEG and SEI are related to other enterotoxins of S. aureus and to streptococcal pyrogenic exotoxin A (SpeA) and streptococcal superantigen (SSA) of Streptococcus pyogenes. Phylogenetic analysis and comparisons of amino acid and nucleotide sequence identities were performed on related staphylococcal and streptococcal protein toxins to group SEG and SEI among the characterized toxins. SEG is most similar to SpeA, SEB, SEC, and SSA (38 to 42% amino acid identity), while SEI is most similar to SEA, SEE, and SED (26 to 28% amino acid identity). Polyclonal antiserum was generated against purified histidine-tagged SEG and SEI (HisSEG and HisSEI). Immunoblot analysis of the enterotoxins, toxic-shock syndrome toxin 1, and SpeA with antiserum prepared against HisSEG and HisSEI revealed that SEG shares some epitopes with SEC1 while SEI does not.

Project description:Enterotoxin-producing Escherichia coli (ETEC) are one of the main causative agents of diarrhea in children especially in developing countries and travel diarrhoea in adults. Pathogenic properties of ETEC associated with their ability to produce a heat-stable (ST) and/or heat-labile (LT) enterotoxins, as well as adhesins providing bacterial adhesion to intestinal epithelial cells. This study presents the molecular characterization of the ETEC isolates collected from the Central and Far-Eastern regions of Russia in 2011-2012. It was shown that all ETEC under study (n=18) had the heat-labile enterotoxin-coding operon elt, and had no the genes of the heat-stable enterotoxin operon est. DNA sequencing revealed two types of nucleotide exchanges in the eltB gene coding subunit B of LT in isolates collected from Cherepovets city (Central region, Russia) and Vladivostok city (Far-East region, Russia). Only one ETEC strain carried genes cfaA, cfaB, cfaC and cfaD coding adhesion factor CFA/I. Expression of LT in four ETEC isolates in the agglutination reaction was detected using a latex test-system. The isolates were assigned to serogroups O142 (n = 6), ?6 (n = 4), ?25 (n = 5), ?26 (n = 2), and O115 (n = 1). Genotyping showed that they belonged to an earlier described sequence-type ST4 (n = 3) as well as to 11 novel sequence-types ST1043, ST1312, ST3697, ST3707, ST3708, ST3709, ST3710, ST3755, ST3756, ST3757 and ST4509. The ETEC isolates displayed different levels of antimicrobial resistance. Eight isolates were resistant to only one drug, three isolates-to two drugs, one isolate-to three drugs, two isolates-to four antibacterials, and only one isolate to each of the five, six and ten antibacterials simultaneously. Genetic determinants of the resistance to beta-lactams and other classes of antibacterials on the ETEC genomes were identified. There are blaTEM (n = 10), blaCTX-M-15 (n = 1), class 1 integron (n = 3) carrying resistance cassettes to aminoglycosides and sulphonamides dfrA17-aadA5 and dfrA12-orfF-aadA2. One isolate ETEC_Ef-6 was found to be a multidrug-resistant (MDR) pathogen that carried both the beta-lactamase gene and class 1 integron. These data suggest the circulation of ETEC in Russia. Further investigations are necessary to study the spread of the revealed ETEC sequence types (STs) and serotypes. Their role in the etiology of diarrhea should be also estimated.

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Characterization of enterotoxin A-producing Staphylococcus aureus.

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Characterization of enterotoxin A-producing <i>Staphylococcus aureus</i>.

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