New Delhi metallo-ß-lactamase in Jamaica.

INTRODUCTION: The global dissemination of the New Delhi metallo-beta-lactamase (NDM) gene among certain strains of bacteria has serious implications since the infections caused by such organisms pose a therapeutic challenge. Although the NDM gene has been detected in various parts of the world, this is the first report of its detection in the English-speaking Caribbean. The NDM producing Klebsiella pneumoniae was isolated from an Indian patient who had recently relocated to Jamaica. METHODOLOGY: Identification and susceptibility testing of the K. pneumoniae isolate was performed using the Vitek 2 automated system) in keeping with Clinical and Laboratory Standards Institute (CLSI) standards. It was identified as a metallobetalactamase producer using the Rosco KPC+MBL kit. Genotypic screening for common betalactamase (including carbapenemase) genes, was carried out  using two multiplex PCRs: one for SHV-, TEM-, CTX-M-, OXA-1-, and CMY-2-types, and one for VIM-, KPC-, IMP-, OXA-48, GES-, and NDM-types. Strain typing was conducted by pulsed-field gel electrophoresis (PFGE) using XbaI and multi-locus sequencing (MLS). Plasmid isolation and analysis was also performed. RESULTS: K. pneumoniae (N11-02395), not previously associated with the dissemination of the NDM in India, Sweden or the UK, was found to harbor the NDM-1 gene on plasmid pNDM112395. CONCLUSION: The identification of the NDM-1 gene underscores the need for effective surveillance and infection control measures to identify and prevent spread of multidrug resistant Gram negative bacilli. Strict infection control measures implemented for this patient helped to prevent the spread of this organism to other patients.

Emergence of an outbreak-associated Clostridium difficile variant with increased virulence.

The prevalence of Clostridium difficile infections has increased due to the emergence of epidemic variants from diverse genetic lineages. Here we describe the emergence of a novel variant during an outbreak in a Costa Rican hospital that was associated with severe clinical presentations. This C. difficile variant elicited higher white blood cell counts and caused disease in younger patients than did other strains isolated during the outbreak. Furthermore, it had a recurrence rate, a 30-day attributable disease rate, and disease severity as great as those of the epidemic strain NAP1. Pulsed-field gel electrophoresis genotyping indicated that the outbreak strains belong to a previously undescribed variant, designated NAPCR1. Whole-genome sequencing and ribotyping indicated that the NAPCR1 variant belongs to C. difficile ribotype 012 and sequence type 54, as does the reference strain 630. NAPCR1 strains are resistant to fluoroquinolones due to a mutation in gyrA, and they possess an 18-bp deletion in tcdC that is characteristic of the epidemic, evolutionarily distinct, C. difficile NAP1 variant. NAPCR1 genomes contain 10% more predicted genes than strain 630, most of which are of hypothetical function and are present on phages and other mobile genetic elements. The increased virulence of NAPCR1 was confirmed by mortality rates in the hamster model and strong inflammatory responses induced by bacteria-free supernatants in the murine ligated loop model. However, NAPCR1 strains do not synthesize toxin A and toxin B at levels comparable to those in NAP1 strains. Our results suggest that the pathogenic potential of this emerging C. difficile variant is due to the acquisition of hypothetical functions associated with laterally acquired DNA.

Isolation of a toxigenic and clinical genotype of clostridium difficile in retail meats in Costa Rica.

We isolated a regional toxigenic genotype of Clostridium difficile, previously found in human infection in 4 of 200 (2%) samples of retail meats for human consumption: 1 of 67 samples of beef, 2 of 66 of pork, and 1 of 67 of poultry meat. These four isolates were positive for the tcdA and tcdB genes but negative for deletion of the tcdC and cdtB genes. All strains induced cytopathic effects in HeLa cells. However, they were susceptible to some antibiotics to which clinical isolates are often resistant. All strains were susceptible to vancomycin, metronidazole, moxifloxacin, and rifampicin but resistant to clindamycin and ciprofloxacin. This first report of isolation of C. difficile in foodstuff from Latin America lends support to the notion that animal products serve as a reservoir for clinical strains of this pathogen in the community.

Identification of a progenitor of the CTX-M-9 group of extended-spectrum beta-lactamases from Kluyvera georgiana isolated in Guyana.

Chromosomal beta-lactamase genes (bla(KLUY)) from six Kluyvera georgiana strains isolated in Guyana were cloned and expressed in Escherichia coli. KLUY-1 exhibited 100% amino acid identity with the extended-spectrum beta-lactamase CTX-M-14. We also show that a 2.7-kb Kluyvera chromosomal region exhibits 99% nucleotide identity to a portion of In60 that includes bla(CTX-M-9).

Characterization of a Salmonella enterica serovar Agona strain harbouring a class 1 integron containing novel OXA-type beta-lactamase (blaOXA-53) and 6'-N-aminoglycoside acetyltransferase genes [aac(6')-I30].

OBJECTIVE: To characterize by molecular methods a multidrug-resistant Salmonella enterica serovar Agona (S. enterica Agona) isolated from a hospitalized patient in Rio de Janeiro, Brazil. METHODS: The S. enterica Agona strain was screened by PCR and DNA sequencing for TEM, SHV and CTX-M-type beta-lactamase genes, tet(A), (B), (C) and (D) tetracycline resistance genes, chloramphenicol resistance genes and class 1 integrons. Plasmid characterization was carried out by PCR and Southern hybridization analysis. PCR and PFGE were used to characterize nine other S. enterica Agona strains collected from hospitals in Rio de Janeiro. RESULTS: The study strain was found to harbour a 105 kb plasmid, which contained catA1, bla(TEM-1), a class 1 integron with two novel genes labelled bla(OXA-53) and aac(6')-I30, respectively, and an additional unidentified aminoglycoside resistance gene. A second 53 kb plasmid from the same strain contained tet(D) and bla(SHV-5). OXA-53 was shown to provide reduced susceptibility to ceftazidime, and its activity was inhibited in the presence of clavulanic acid. PFGE analysis of the nine other S. enterica Agona strains revealed two clusters of related strains (78% similarity), and PCR analysis showed that all strains contained the novel integron. CONCLUSION: An S. enterica Agona strain was found to harbour three plasmid-encoded beta-lactamases, one (OXA-53) on a novel class 1 integron that also contains a new aminoglycoside resistance gene, aac(6')-I30. The multidrug resistance plasmids appear to have disseminated to other city hospitals via other S. enterica Agona strains.