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*
Centre Nationale des Toxémies à Staphylococques, Faculté de Médecine Laennec, Lyon, France;
Institute National de la Santé et de la Recherche Médicale Unité 463, Institut de Biologie, Nantes, France;
Institute National de la Santé et de la Recherche Médicale Unité 277, Institut Pasteur, Paris, France; and
Unité Mixte de Recherche Centre National de la Recherche Scientifique 5557, Laboratoire d’Ecologie Microbienne, Universite Claude Bernard, Villeurbanne, France
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResults and DiscussionReferences |
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ent1 and ent2.
RT-PCR analysis showed that all these genes, including
seg and sei, belong to an operon,
designated the enterotoxin gene cluster (egc).
Recombinant SEG, SEI, SEK, SEL, and SEM showed superantigen activity,
each with a specific V
pattern. Distribution studies of genes
encoding superantigens in clinical S. aureus isolates
showed that most strains harbored such genes and in particular the
enterotoxin gene cluster, whatever the disease they caused.
Phylogenetic analysis of enterotoxin genes indicated that they all
potentially derived from this cluster, identifying egc
as a putative nursery of enterotoxin genes. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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SEs and TSST-1 share common structural and biological properties,
suggesting that they derived from a common ancestor (3).
They display significant homology in their primary sequence and
secondary and tertiary structures (3). Based on amino acid
sequence comparisons, SEs have been divided into several groups; one
includes SEA, SEE, SEJ, SED, and SEH, and another SEB and SEC, whereas
SEG and SEI could not be clearly attributed to a specific group
(4, 5). Biologically, SEs and TSST-1 exhibit
superantigen activity, stimulating polyclonal T cell proliferation
through coligation between MHC class II molecules on APCs and the
variable portion of the T cell Ag receptor -chain (TCR V)
(3). The pattern of V activation is specific for each
of these superantigens (3). T cell/APC activation by these
toxins leads to the release of various cytokines/lymphokines and IFN,
enhances endotoxic shock, and causes T and B cell immunosuppression,
all of which may hinder the immune response against bacterial infection
(5, 6, 7, 8).
SEG and SEI are recently described SEs (4). We have
previously reported the involvement of these toxins in TSS and SSF
(9). The SEG and SEI genes (seg and
sei) were originally identified in two separate strains
(4), but we have shown that, when present, seg
and sei coexist in all clinical isolates of S.
aureus examined to date (9). Moreover, we found that
the two genes were in tandem orientation on the same 3.2-kb DNA
fragment. As Munson et al. (4) have found that the
seg transcript is unusually large (
6.7 kb), we postulated
that the seg transcript might encode additional genes,
including sei. The aim of this study was to identify and
characterize the genes that are cotranscribed with seg.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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S. aureus A900322, isolated from a patient with TSS, was shown to have the genotype sea-, seb-, sec-, sed-, see-, seg+, seh-, sei+ (9), and was used as a seg and sei reference strain. S. aureus RN450 (sea-, seb-, sec-, sed-, see-, seg-, seh-, sei-) was used as a negative control for SE genes. S. aureus MJB1316 (a gift from Sibyl Munson, University of Wisconsin, Madison, WI), an RN450 derivative that contains the cloned seg gene on the staphylococcal expression vector pRN5548 (4), was used as seg positive control. The following S. aureus strains were used to check the specificity of PCR amplification: FDA-S6 (ATCC 13566 (sea+ seb+)), FRI-137 (ATCC 19095 (sec+ seg+ seh+ sei+)), FRI-1151 m (sed+), FRI-326 (ATCC 27664 (see+)), FRI-569 (ATCC 51811 (seh+)), FRI-1169 (tst+), TC-7 (eta+ seg+ sei+), and TC-146 (etb+ seg+ sei+) (9). Two hundred thirty S. aureus clinical isolates were collected by the Center National de Référence des Toxémies à Staphylocoques (Lyon, France) between January 1998 and December 1999. They were isolated from 58 patients with S. aureus infection (arthritis, skin infection, pneumonia, or infective endocarditis), 102 patients with acute toxemia (TSS, SSF, or SSSS), and 70 asymptomatic nasal carriers. All strains were collected from hospitals located throughout France and were identified as S. aureus by their ability to coagulate citrated rabbit plasma (bioMérieux, Marcy-l’Etoile, France) and to produce a clumping factor (Staphyslide Test; bioMérieux). Escherichia coli TG1 was used for plasmid amplification and genetic manipulations.
DNA amplification and sequencing
DNA was extracted from A900322 cultures and used as a template
for amplification with primers sei-1 and seg-2 (Table I
) in conditions described in detail
elsewhere (9). Primers wsei and wseg (Table I) were
designed following identification of suitable hybridization sites in
the sei and seg genes and were compatible with
the Clontech Genome Walker kit (Ozyme; Montigny-Le Bretonneux, France),
which is suitable for cloning unknown DNA sequences adjacent to a known
sequence. This kit was used, according to the supplier’s instructions,
to identify sei and seg flanking regions using
primers hindIII and wsei (Table I) on a HindIII chromosomal
digest for the amplification of the sei-upstream region; and
primers hpaI and wseg (Table I) on an HpaI chromosomal
digest for the amplification of the seg-downstream region.
PCR products were analyzed by electrophoresis through 0.8% agarose
gels (Sigma, St. Louis, MO), purified using the High Pure PCR Product
Purification kit (Boehringer Mannheim, Meylan, France), and sequenced
(Genome Express, Grenoble, France). Sequences were compiled, analyzed,
and compared using Blast (http://www.ncbi. nlm.nih.gov/BLAST),
GeneJokey, and ClustalX software (European Bioinformatics Institute,
Cambridge, U.K., http://www.ebi.ac.uk) (10).
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The sequences of SE-related genes were obtained from GenBank: sea accession number M18970; seb accession number M11118; sec1 accession number X05815; sed accession number M28521; see accession number M21319; seh accession number U11702; and ent accession number U93688. Nucleotide sequences of these genes and open reading frames (ORFs) encoded by egc were aligned using the multiple alignment ClustalX software (10). The evolutionary distances were determined by the method of Kimura, and these values were used to construct a dendrogram by means of the neighbor-joining method using the Phylip package (European Bioinformatics Institute). At least 1000 bootstrap trees were generated for each data set to investigate the stability of phylogenic relationships using the Seqboot module of Phylip package. Similar phylogenic analysis was conducted using the corresponding amino acids sequences.
Toxin-gene detection
Sequences specific for sea-e, seg-i, tst, eta, and
etb, encoding SEA-E, SEG-I, TSST-1, ETA, and ETB,
respectively, were detected by PCR, as previously described
(9). DNA from clinical isolates was extracted from
cultures and used as a template for amplification with the primers
described in Table I
(Eurogentec, Seraing, Belgium). Amplification of
gyrA was used as a control to confirm the quality of each
DNA extract and the absence of PCR inhibitors (9). All PCR
products were analyzed by electrophoresis through 1% agarose gels
(Sigma).
Detection of bacterial RNA by RT-PCR
Total RNA was extracted from staphylococcal cultures by using
RNeasy spin columns (Qiagen, Courtaboeuf, France). cDNA was synthesized
using Ready-To-Go RT-PCR beads (Pharmacia Biotech, Orsay, France) by
incubating 0.1 µg of total RNA with the following pairs of primers
(primer 5', sel3), (sel-4, sel-5), (sel1, sel2), (invsel2, invsem1),
(sem1, invsei1), (sei1, sei2), (invsei2, ent2), (ent1, invsek1),
(sek1, sek2), (invsek2, invseg1), (seg1, seg2), (invseg2, primer
3') (Table I). The reaction mixtures were incubated with each primer
pair described above, at 42°C for 30 min for reverse transcription,
followed by 30 cycles of amplification (1-min denaturation at 94°C,
1-min annealing at 55°C, and 1-min extension at 72°C). The RT-PCR
products were then analyzed by electrophoresis through 1% agarose gel.
RNA extracts were tested for DNA contamination by preincubating the
reaction mixtures at 95°C for 10 min to inactivate reverse
transcriptase before the RT-PCR.
Production and purification of recombinant enterotoxins
Primers were designed following identification of suitable
hybridization sites in sel, sem, sei, sek, and
seg (Table I). The 5' primers were chosen within the coding
sequence of each gene, omitting the region predicted to encode the
signal peptide, as determined by hydrophobicity analysis according to
Kyte and Doolitttle (11) with GeneJockey software and
SignalP V1.1 World Wide Web Prediction Server
(http://www.cbs.dtu.dk/services/SignalP/) (12); the 3'
primers were chosen to overlap the stop codon of each gene. A
restriction site was included in each primer (Table I). DNA was
extracted from A900322 or MJB1316 and used as a template for PCR
amplification. PCR products and plasmid DNA were prepared using the
Qiagen plasmid kit. PCR fragments were digested with EcoRI
and PstI (Boehringer Mannheim) and ligated (T4 DNA ligase;
Boehringer Mannheim) with the pMAL-c2 expression vector from New
England Biolabs (Ozyme) digested with the same restriction enzymes. The
resulting plasmids were transformed into E. coli TG1. The
integrity of the ORF of each construct was verified by DNA sequencing
of the junction between pMAL-c2 and the different inserts. The fusion
proteins were purified from cell lysates of transfected E.
coli by affinity chromatography on an amylose column according to
the supplier’s instructions (New England Biolabs).
T cell proliferation assays
PBL from healthy donors were cultured in 24-well plates (106 cells/well) in RPMI 1640 medium supplemented with 8% pooled human serum and 10 µg/ml recombinant staphylococcal toxin. rIL-2 (50 IU/ml) was added on day 5. When necessary, T cell cultures were diluted in IL-2-supplemented medium until TCR analysis. We used as controls T cells from the same donors that were stimulated with 0.5 µg/ml Phaseolus vulgaris leucoagglutinin (PHAL) (Sigma).
Flow cytometry
The following mAb (mAb; specificity indicated in brackets) were
used for flow cytometry: E2.2E7.2 (V2), LE89 (V3), IMMU157
(V5.1), 3D11 (V5.3), CRI304.3 (V6.2), 3G5D15 (V7), 56C5.2
(V8.1/8.2), FIN9 (V9), C21 (V11), S511 (V12), IMMU1222
(V13.1), JU74 (V13.6), CAS1.1.13 (V14), Tamaya1.2 (V16),
E17.5F3 (V17), BA62.6 (V18), ELL1.4 (V20), IG125 (V21.3),
IMMU546 (V22), and HUT78.1 (V23). These mAb, and CD4- and
CD8-specific mAb, were purchased from Beckman/Coulter/Immunotech
(Marseille, France). Cells were phenotyped by indirect
immunofluorescence, as described previously (13). Briefly,
cells were incubated with unconjugated mAb for 30 min at room
temperature, then washed and incubated with FITC-conjugated rabbit
anti-mouse Ig antiserum (BioAtlantic, Nantes, France) for 30 min on
ice. After washing, cells were analyzed by flow cytometry on a FACScan
apparatus (Becton Dickinson, Mountain View, CA) using the LYSYS II
software package on a FACstation.
Immunoscope analysis
Total RNA was extracted using the Trizol reagent (Life
Technologies, Gaithersburg, MD). TCR -chain-specific primers were as
described previously (14), and reverse transcription, PCR
amplification, and run-off steps were performed as reported previously
(15). Fluorescent DNA products were loaded on a sequencing
gel and analyzed with the Immunoscope software (16).
Statistical analysis
2 test was used to determine whether
the distribution of egc, sea, seb,
sec, sed, see, seh,
tst, eta, and etb was significantly
different in isolates from asymptomatic nasal carriers and patients
with S. aureus infection or acute toxemia; p
< 0.05 was considered statistically significant.
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Results and Discussion |
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TopAbstractIntroductionMaterials and MethodsResults and DiscussionReferences |
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When this work was initiated, the coding regions of only
seg and sei were available, and the two genes
were known to be in tandem orientation, separated by a 1.9-kb DNA
fragment in S. aureus strain A900322 (4, 9). A
3.2-kb fragment was thus amplified by PCR with primers sei1 and seg2
and was then sequenced. The intergenic 1.9-kb DNA sequence contained
three open reading frames (ORF1, 2, and 3) of 399, 327, and 777 bp,
respectively. Comparison of the deduced amino acid sequences of these
ORFs with translated sequences from GenBank showed that the putative
proteins corresponding to these ORFs had substantial sequence
similarities to known SEs: ORF1 exhibited homology to the N-terminal
region of SEB; ORF2 to the C-terminal region of SEC; and ORF3 to SEA
(Table II). The PCR "walking"
strategy was chosen to identify the seg and sei
flanking regions. The use of primers wsei and hindIII on
HindIII digests allowed us to amplify and sequence the 3.2
kb of DNA upstream of sei. Analysis of this sequence showed
two significant ORFs (ORF4 and ORF5) of 783 and 720 bp, respectively.
ORF4 exhibited homology with SEJ, and ORF5 with SEI (Table II). The use
of primers wseg and hpaI on HpaI digests amplified a 0.8-kb
fragment downstream of seg. Sequence analysis of this
fragment revealed no other significant ORFs. The concatenated sequence
of seg-sei-intergenic, -upstream and -downstream regions was
validated by sequencing a 6.189-kb PCR fragment encompassing the whole
region (Fig. 1). Although sei
in strain A900322 was 100% homologous with the sequence deposited in
GenBank (accession number AF064774), seg in strain A900322
showed one mutation, corresponding to a Leu
Pro substitution at
position 29. This new variant was designated
SEGL29P.
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ent1 and 2,
respectively, meaning they represent pseudogenes with no likely
biological function. In contrast, ORFs 3, 4, and 5 had sizes consistent
with active enterotoxin-like molecules. ORF5 possesses a satisfactory
SD sequence and translation start site, whereas ORF3 and ORF4 have an
adequate SD sequence in front of a noncanonical, although suitable
(18), translation start site (ATT) coding thethionine
(Fig. 1). Thus, ORF3, ORF4, and ORF5 were designated sek,
sel, and sem, respectively. Thus, the 6301-bp DNA
region identified in this study contains seg and
sei plus three potential enterotoxin genes (sek,
sel, and sem) and two pseudogenes (
ent1,
ent2), all in the same orientation (Fig. 1). We
designated this region egc for "enterotoxin gene
cluster." With the exception of plasmid pIB485, which contains
sed and sej in opposite orientations separated by
895 nucleotides, and the staphylococcal pathogenicity island, which
contains tst and ent separated by 10.234 kb
(19), no such gene cluster organization has ever been
described for enterotoxin genes. It is likely that this organization
was generated through gene duplication and variation from an ancestral
gene. The proposed molecular mechanism involved in egc
formation is unequal crossing-over, which can be generated when
recombination occurs between nonallelic regions by misalignment
(20). This hypothesis is supported by the presence of
homologous genes and pseudogenes in the same DNA region. Transcriptional analysis
As mentioned above, Munson et al. (4) reported an
unusually large (6.7-kb) seg transcript. To investigate
whether this transcript was polycistronic, i.e., encoded one or more of
the ORFs identified in egc, c-DNA was generated from strain
A900322 total RNA by reverse transcription and amplified by PCR using
primer pairs located within each gene and bracketing adjacent genes.
Abundant RT-PCR products (B to K) of the expected size were obtained
using the corresponding primer pairs (Fig. 2). In contrast, no RT-PCR product A
(primer 5', sel3) nor L (primer invseg2 and primer 3') was obtained
(Fig. 2). These results suggest that the seven genes and pseudogenes
composing egc are cotranscribed, and that the 5' and 3' ends
of the transcript must be close to the beginning of sel and
to the end of seg, respectively. Sequence analysis revealed
putative -10 and -35 promotor sequences (TTGTCT-N15-TAATTT-N134-ATT)
upstream of the sel start codon. The 3' end may lie at an
inverted repeat at position 6018–6067, which is a potential
transcription termination signal, 5830 nucleotides downstream of the
putative transcription start site. These results suggest that
egc is an operon. However, we could not rule out the
coexistence of alternative transcription start sites and/or termination
sites resulting in partial egc transcription. The size of
the egc transcript was slightly shorter than that previously
estimated by means of Northern blot analysis (6.7 kb) by Munson et al.
This discrepancy is most probably due to technical reasons, as Northern
blot analysis permits only a rough estimate of RNA size.
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The association of related genes that are cotranscribed
suggested that the resulting peptides might have complementary effects
on the host’s immune response. One hypothesis was that gene
recombination had created new variants of toxins differing by their
superantigen profiles. Purified recombinant SEL, SEM, SEI, SEK, and
SEGL29P expressed in E. coli were
studied for their ability to induce selective expansion of T cells
bearing particular TCR V regions in short-term PBL culture. As shown
in Tables III and IV, recombinant SEL SEM, SEI, and SEK
consistently induced selective expansion of distinct sets of V
subpopulations. By contrast, SEGL29P failed to
trigger expansion of any of the 23 V subsets. The sum of results
obtained with each of these recombinant toxins globally corresponded to
the selective expansion of V subpopulations induced by crude
supernatant of staphylococcal culture of strains that harbored
egc (data not shown). This suggested that the
maltose-binding protein portion of the fusion toxins did not
significantly influence the V specificity of these superantigens. To
investigate whether the L29P mutation could explain the lack of
superantigen activity, a rSEG with an L29 codon was constructed from
S. aureus strain MJB1316 (which contains the cloned
seg on a plasmid) and then expressed in E. coli,
and the superantigen activity of this toxin was tested.
SEGL29.induced selective expansion of V14 and,
to a lesser extent, V13.6,0T cells (Table III). The L29P mutation
thus accounts for the complete loss of superantigen activity.
Computer modeling of the two-dimensional structure (21) of
the wild-type and mutated proteins revealed no major conformational
differences between the two proteins (not shown). It is likely that L29
is located at a position crucial for proper superantigen/MHC II
interaction. In addition to the selective expansion of TCR V subsets
observed with the different toxins, flow cytometry revealed
preferential expansion of CD4 T cells in SEI and SEM cultures (Table III). By contrast, the CD4/CD8 ratios in SEK-, SEL-, and SEG-stimulated
T cell lines were close to those in fresh PBL. This phenomenon, which
was observed with cells from several donors, may reflect a variable
contribution of the CD4 coreceptor to the T cell activation process,
depending on the affinity of the TCR for the superantigen/MHC complex
(22, 23).
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composition of superantigen-stimulated T cell
lines and the clonal diversity of the expanded TCR V subsets, the
size distribution of PCR-amplified TCR -chain junctional products
was studied using the Immunoscope technique (14, 15, 16).
Results of this molecular analysis were in good overall agreement with
those obtained by flow cytometry, as similar dominant TCR V subsets
were identified with the two approaches (Fig. 3, Tables III and IV, and data not shown
for rSEK). Slight discrepancies observed in some instances may have
been due to selective expansion of particular members of a given V
subfamily not recognized by available mAb (e.g., V6 or V13), and
to the fact that TCR V frequencies were estimated with a
semiquantitative PCR technique, which might have led to slight over- or
underestimation of particular V subsets. Additionally, Immunoscope
analysis showed that the complementarity-determining region 3 size
distribution of TCR -chain junctional transcripts within expanded
V subsets was pseudogaussian in all superantigen-stimulated
cultures, reflecting a high level of polyclonality (data not shown).
This was further confirmed by sequence analysis of TCR junctional
transcripts derived from some expanded TCR V subsets (e.g.,
V5+ cells in SEL and SEI cultures) (not
shown). Taken together, these TCR repertoire studies confirmed the
superantigenic nature of the new toxins identified in this study.
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We then analyzed the distribution of egc (by
PCR amplification encompassing sei to seg) and
that of all known enterotoxins (by selective PCR (9)) in
230 S. aureus strains isolated in various clinical settings
(nasal carriage, suppurative infection, and toxemia). As shown in Table V, the majority of the isolates harbored
gene(s) encoding superantigenic toxins, whatever the clinical setting.
seg-sei (and thus egc) were present in most
toxemia strains (59% in TSS, 48% in SSF, and 92% in SSSS), and also
in most strains associated with suppurative infections (67%) and nasal
carriage (57%). Moreover, egc appeared to be the most
frequent superantigens in S. aureus, whatever the clinical
setting. The prevalence of egc in strains associated with
SSSS, a disease caused by ETs, was significantly higher than that in
nasal carriage strains (2 test,
p = 0.03) (Table V). This could reflect the clonal
origin of the strains associated with SSSS, as previously suggested by
phage typing, pulsed-field gel electrophoresis, and amplified fragment
length polymorphism (24 , G. Lina, manuscript in
preparation). The strains associated with TSS were significantly more
frequently TSST-1 producers than were nasal carriage strains
(2 test, p = 0.04) (Table V), whereas no
significant difference was observed between the two groups of strains
regarding the presence of egc. Thus, the superantigens
produced by egc must have a role other than the induction of
toxemia. As each toxin encoded by egc was associated with a
complementary pattern of V subset usage, a bacterium that produces
such a panel of superantigens theoretically has a marked capacity for
stimulating polyclonal T cell proliferation and thus for inducing
several deleterious effects, including immune anergy by T cell
suppressor activity, B cell depletion, and inhibition of Ab responses
(6, 7, 8). We speculate that the apparent redundancy of these
superantigens confers a selective advantage toward colonization and/or
invasion of human and not only for toxemia.
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The high prevalence of egc among staphylococcal
isolates raises the possibility that this locus acts as a reservoir of
enterotoxin genes. Phylogenic analysis was conducted, including all
known enterotoxins and enterotoxin-like toxins in S. aureus.
Phylogenetic trees, constructed from the nucleic acid sequences of
these genes and from the amino acid sequences of the corresponding
toxins by using the neighbor-joining method, were superimposable. The
position of ent1 and its products was unstable, as
reflected by the low bootstrap value at the node from which they
branched (52.5% and 62%, respectively). As the Phylip package was not
able to confidently branch ent2, this gene is not
presented in the tree. All other nodes were well supported (>70%
bootstrap values) (Fig. 4). We identified
three monophyletic groups within the tree: one composed of
sea, see, sej, sed, sek,
sel, and seh; another including seb,
sec, ent2, seg, and probably
ent1; and a third including sei, sem,
and ent (a putative enterotoxin identified in the
staphylococcal pathogenicity island (19)). Each of these
clusters contained one or more genes encoded in egc.
Remarkably, each of the predicted egc products showed the
strongest homology with one of the known enterotoxins encoded outside
egc on monocistronic loci. This phylogenic organization
could be interpreted as suggesting that gene ancestors of enterotoxin
genes encoded outside egc derive from egc. Thus,
egc would appear to be an enterotoxin gene nursery. The
mechanism by which gene diversity has been generated in egc
and then exported on the mode of a single gene to other regions of the
chromosome remains to be elucidated.
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subsets. The wide distribution of
egc in clinical isolates suggests that it is beneficial for
S. aureus. Finally, phylogenetic analysis of all known
enterotoxins indicates that they all potentially derived from this
cluster, inferring that egc is in an enterotoxin gene
nursery. Note added in proof. While the present article was under review, Williams et al. 25 reported the discovery of a novel genetic locus within S. aureus that encodes a cluster of at least five exotoxin-like proteins designated the staphylococcal exotoxin-like genes 1 to 5 (set1 to set5). Comparison of the nucleotide sequences of set1-5 with that of egc revealed that the two clusters are distinct.
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Acknowledgments |
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Footnotes |
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2 Abbreviations used in this paper: SE, staphylococcal enterotoxin; ET, exfoliative toxin; ORF, open reading frame; SD, Shine-Dalgarno; SSF, staphylococcal scarlet fever; SSSS, scalded skin syndrome; TSS, toxic shock syndrome; TSST, TSS toxin.
Received for publication July 26, 2000. Accepted for publication October 4, 2000.
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D. S. Blanc, C. Petignat, A. Wenger, G. Kuhn, Y. Vallet, D. Fracheboud, S. Trachsel, M. Reymond, N. Troillet, H. H. Siegrist, et al. Changing Molecular Epidemiology of Methicillin-Resistant Staphylococcus aureus in a Small Geographic Area over an Eight-Year Period J. Clin. Microbiol., November 1, 2007; 45(11): 3729 - 3736. [Abstract] [Full Text] [PDF]
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P. M. Schlievert, L. C. Case, K. L. Strandberg, T. J. Tripp, Y.-C. Lin, and M. L. Peterson Vaginal Staphylococcus aureus Superantigen Profile Shift from 1980 and 1981 to 2003, 2004, and 2005 J. Clin. Microbiol., August 1, 2007; 45(8): 2704 - 2707. [Abstract] [Full Text] [PDF]
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S. Holtfreter, D. Grumann, M. Schmudde, H. T. T. Nguyen, P. Eichler, B. Strommenger, K. Kopron, J. Kolata, S. Giedrys-Kalemba, I. Steinmetz, et al. Clonal Distribution of Superantigen Genes in Clinical Staphylococcus aureus Isolates J. Clin. Microbiol., August 1, 2007; 45(8): 2669 - 2680. [Abstract] [Full Text] [PDF]
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D. S. Smyth, W. J. Meaney, P. J. Hartigan, and C. J. Smyth Occurrence of ssl genes in isolates of Staphylococcus aureus from animal infection J. Med. Microbiol., March 1, 2007; 56(3): 418 - 425. [Abstract] [Full Text] [PDF]
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W. R. Morgan, M. D. Caldwell, J. M. Brady, M. E. Stemper, K. D. Reed, and S. K. Shukla Necrotizing Fasciitis Due to a Methicillin-Sensitive Staphylococcus aureus Isolate Harboring an Enterotoxin Gene Cluster J. Clin. Microbiol., February 1, 2007; 45(2): 668 - 671. [Abstract] [Full Text] [PDF]
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M. M. Collery and C. J. Smyth Rapid differentiation of Staphylococcus aureus isolates harbouring egc loci with pseudogenes {psi}ent1 and {psi}ent2 and the selu or seluv gene using PCR-RFLP J. Med. Microbiol., February 1, 2007; 56(2): 208 - 216. [Abstract] [Full Text] [PDF]
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D. Y. Thomas, S. Jarraud, B. Lemercier, G. Cozon, K. Echasserieau, J. Etienne, M.-L. Gougeon, G. Lina, and F. Vandenesch Staphylococcal Enterotoxin-Like Toxins U2 and V, Two New Staphylococcal Superantigens Arising from Recombination within the Enterotoxin Gene Cluster. Infect. Immun., August 1, 2006; 74(8): 4724 - 4734. [Abstract] [Full Text] [PDF]
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T. Ferry, M. Bes, O. Dauwalder, H. Meugnier, G. Lina, F. Forey, F. Vandenesch, and J. Etienne Toxin Gene Content of the Lyon Methicillin-Resistant Staphylococcus aureus Clone Compared with That of Other Pandemic Clones. J. Clin. Microbiol., July 1, 2006; 44(7): 2642 - 2644. [Abstract] [Full Text] [PDF]
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F. Layer, B. Ghebremedhin, W. Konig, and B. Konig Heterogeneity of Methicillin-Susceptible Staphylococcus aureus Strains at a German University Hospital Implicates the Circulating-Strai |
