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ABSTRACT: To report a clonal outbreak of ST17 vancomycin-resistant Enterococcus faecium (VREfm) carrying Tn1546 (vanA) in a haemo-oncology ward of a tertiary teaching hospital in the south of Spain (January-September 2009).
Twenty-two VREfm strains from 13 patients were characterized by PFGE, multiple-locus variable-number tandem-repeat analysis (MLVA) and multilocus sequence typing (MLST). Genes encoding antibiotic resistance and putative virulence traits and the Tn1546 backbone were investigated by PCR. Plasmid characterization included determination of size (S1-PFGE) and replication modules (PCR, hybridization and sequencing). Patient clinical records were analysed retrospectively.
A single ST17 E. faecium clone (MT-7 MLVA type) carrying esp and hyl plus a 30 kb Inc18-like::Tn1546 (IS1216) plasmid was identified. Ampicillin resistance was linked to PBP5 showing mutations at positions 24, 27, 34, 66, 68, 85, 100, 144, 172, 177, 204, 216, 324, 462, 466', 470, 485, 496, 499, 525, 546, 558, 582, 586, 629, 632, 642 and 667. Other resistance genes identified were erm(B), ant(6')-Ia and aph(3')-IIIa. Fluoroquinolone resistance was attributable to ParC (Arg-61 → Gly and Ser-80 → Arg) and GyrA (Ser-83 → Arg) mutations.
A nosocomial outbreak caused by an ST17 (CC17) E. faecium clone harbouring Esp and Hyl and a 30 kb Inc18-like::Tn1546 plasmid among haemo-oncology patients is reported. The failure of early infection control practices indicates an undetected reservoir and the ability of this strain to persist over long periods. The potential spread of epidemic clones and broad host plasmids carrying vancomycin resistance in Spain is of concern since it might contribute towards a higher rate of VREfm infection.
Journal of Antimicrobial Chemotherapy 01/2012; 67(4):832-6. · 5.07 Impact Factor
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ABSTRACT: The severe morbidity of human brucellosis is one of the main reasons for using molecular typing in the epidemiological surveillance of this worldwide zoonosis. Multiple-locus variable-number repeat analysis (MLVA-16), hypervariable octameric oligonucleotide fingerprinting (HOOF-print), and the differences in the single nucleotide polymorphisms (SNPs) (codons 1249 and 1309) of the DNA-dependent RNA polymerase beta subunit (rpoB) were used to type a human Brucella melitensis population (108 strains) collected from throughout Spain over 13 years. Eighty-six MLVA types (discriminatory index, 0.99) were detected, with a wide-ranging genetic similarity coefficient (37.2 to 93.7%). The population clustered into the following groups: American, with genotypes 47 (1 strain), 48 (13 strains), 53 (12 strains), 55 (2 strains), 80 (1 strain), and a new genotype (2 strains), Western Mediterranean, with genotype 51 (9 strains), and Eastern Mediterranean, with genotypes 42 (60 strains), 43 (4 strains), and 63 (4 strains). Two profession-related and two foodborne acquisitions were confirmed. Distributed throughout Spain, Eastern Mediterranean genotype 42 was the most common (55%). The low MLVA-16 allelic polymorphism (genetic similarity range, 75 to 94%) of the genotype 42 strains suggests that they recently evolved from a common ancestor. rpoB typing grouped the strains as rpoB type 1 (1249-ATG/1309-CTG; 28.7%), rpoB type 2 (1249-ATG/1309-CTA; 62.9%), and rpoB type 3 (1249-ATA/1309-CTG; 8.3%). According to the MLVA-16 results, the population clustered by rpoB type. Given the correlation between B. melitensis MLVA groups and rpoB types (American and rpoB type 1, Eastern Mediterranean and rpoB type 2, and Western Mediterranean and rpoB type 3), the rpoB type could be used as an initial marker for the epidemiological surveillance of brucellosis.
Journal of clinical microbiology 08/2010; 48(8):2734-40. · 4.16 Impact Factor
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ABSTRACT: The aim of this study was to determine, using molecular methods, whether rifampicin and fluoroquinolone resistance was present in a clinical Brucella melitensis population.
Sixty-two B. melitensis strains, isolated from humans-most experiencing their first brucellosis episode-over an 11 year period in Spain, were genotyped by multiple locus variable analysis (MLVA-16) for future studies. In the present work, molecular screening was undertaken to detect the presence of rpoB and gyrA/gyrB/parC/parE mutations (previously described in in vitro Brucella spp. mutants) related to resistance to rifampicin and fluoroquinolones, respectively.
Sixty-two MLVA-16 genotypes were identified among the B. melitensis population, with genetic similarity values ranging from 32% to 94%. rpoB mutations related to rifampicin resistance (positions 154, 526, 536, 539, 541, 574) were not detected. Neither were changes in GyrA described in in vitro mutants (67, 71, 87, 91 and an insertion at 340) detected in these strains. All showed identical GyrA, GyrB, ParC and ParE sequences with respect to B. melitensis 16M, except for one strain (ciprofloxacin and moxifloxacin MICs 0.25-0.50 mg/L) that harboured the Val264Ala replacement outside the GyrA quinolone resistance-determining region (QRDR); no differences were seen, however, in the NorMI/II efflux pump genes.
The absence of rpoB mutations clearly related to rifampicin resistance in clinical B. melitensis strains reinforces the first-choice status of this antibiotic in the treatment of first brucellosis episodes, and demonstrates the usefulness of molecular screening for resistant genotypes. The absence of topoisomerase II-IV mutations, however, cannot rule out fluoroquinolone resistance due to the interplay of different mechanisms.
Journal of Antimicrobial Chemotherapy 10/2009; 65(1):51-3. · 5.07 Impact Factor
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ABSTRACT: Brucella abortus biovar 3 (BAb3) is an uncommon cause of human brucellosis in Spain; where B. melitensis accounted for 97.5% of all cases. ...
Journal of clinical microbiology 07/2009; 47(8):2687-8. · 4.16 Impact Factor
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ABSTRACT: A large clonal outbreak of multidrug-resistant CC17 ST17 Enterococcus faecium containing Tn5382 in a hospital in the north of Spain is described.
We characterized vancomycin-resistant E. faecium isolates from 10 infected and 40 colonized inpatients from a single hospital by PFGE, multiple-locus variable-number tandem-repeat analysis (MLVA) and multilocus sequence typing (MLST). Genes encoding antibiotic resistance (ampicillin, aminoglycosides, macrolides, quinupristin/dalfopristin, quinolones, tetracycline) and putative virulence traits were analysed.
All isolates showed highly similar PFGE profiles and were assigned to the type MT1 by MLVA and to ST17 (CC17) by MLST. The Tn5382 type identified in all isolates was linked to pbp5 and contained a 5 bp deletion and 10 point mutations within the intergenic vanS(B)-vanY(B) region. Other resistance genes identified were erm(B), mef(E), tet(M), ant(6')-Ia, aph(3')-IIIa and aac(6')-Ie-aph(2'')-Ia. All isolates carried the unexpressed tet(M) gene. The high level of ciprofloxacin resistance was attributable to the first described Gly-61 and Ile-80 mutations in ParC and the Tyr-83 or Arg-83 mutations in GyrA. All isolates contained esp. The presence of hyl was variable.
A large clonal outbreak caused by multidrug-resistant CC17 E. faecium containing pbp5-Tn5382 is described. The persistence of this clone, which has been recovered from both hospital and community settings since 2005, and the possibility of transferring this Tn5382 to other epidemic ampicillin-resistant clonal types currently circulating in Spain might contribute to increasing the prevalence of vancomycin-resistant enterococci in our area. This study constitutes the first description of mef(E) in E. faecium.
Journal of Antimicrobial Chemotherapy 12/2008; 63(1):17-20. · 5.07 Impact Factor
