Polymyxin resistance in Enterobacterales: overview and epidemiology in the Americas.

The worldwide spread of carbapenem- and polymyxin-resistant Enterobacterales represents an urgent public-health threat. However, for most countries in the Americas, the available data are limited, although Latin America has been suggested as a silent spreading reservoir for isolates carrying plasmid-mediated polymyxin resistance mechanisms. This work provides an overall update on polymyxin and polymyxin resistance and focuses on uses, availability and susceptibility testing. Moreover, a comprehensive review of the current polymyxin resistance epidemiology in the Americas is provided. We found that reports in the English and Spanish literature show widespread carbapenemase-producing and colistin-resistant Klebsiella pneumoniae in the Americas determined by the clonal expansion of the pandemic clone ST258 and mgrB-mediated colistin resistance. In addition, widespread IncI2 and IncX4 plasmids carrying mcr-1 in Escherichia coli come mainly from human sources; however, plasmid-mediated colistin resistance in the Americas is underreported in the veterinary sector. These findings demonstrate the urgent need for the implementation of polymyxin resistance surveillance in Enterobacterales as well as appropriate regulatory measures for antimicrobial use in veterinary medicine.

Phenotypic and molecular characterization of Klebsiella spp. isolates causing community-acquired infections.

Klebsiella spp. isolates from community-acquired infections were characterized. A total of 39 Klebsiella spp. isolates were obtained from outpatients at four rural hospitals in Mexico (2013-2014). The biochemical tests identified all as being K. pneumoniae. The molecular multiplex-PCR test identified 36 (92.4%) K. pneumoniae isolates and one (2.5%) K. variicola isolate, and phylogenetic analysis of the rpoB gene identified two isolates (5.1%) belonging to K. quasipneumoniae subsp. quasipneumoniae and K. quasivariicola. The last one was confirmed by phylogenetic analysis of six-loci concatenated genes. Mostly the isolates were multidrug resistant; however, a minority were extended-spectrum ß-lactamase producing (10.2%). The extended-spectrum ß-lactamase CTX-M-15 gene was identified in these isolates. Analysis of biofilm production and the hypermucoviscosity phenotype showed a total of 35 (92.3%) and seven (17.9%) of the isolates were positive for these phenotypes respectively. The K2 (4/39, 10.2%), K5 (2/39, 5.1%) and K54 (1/39, 2.5%) serotypes were identified in seven (17.9%) of the isolates, and only 28.5% (2/7) hypermucoviscous isolates were positive for the K2 and K5 serotypes. In general, the sequence type (ST) analysis and phylogenetic analysis of seven multilocus sequence typing loci were heterogeneous; however, ST29 was the most prevalent ST in the analysed isolates, accounting for 19% (4/21) of the total isolates. Two of the four ST29 isolates had the hypermucoviscosity phenotype. The virulence factors for fimbriae were the most prevalent, followed by siderophores. Community-acquired infections are caused by various species from Klebsiella genus, with different profiles of antibiotic resistance and heterogeneous virulence factors.

Draft Genome Sequence of a Hypermucoviscous Extended-Spectrum-ß-Lactamase-Producing Klebsiella quasipneumoniae subsp. similipneumoniae Clinical Isolate.

A clinical isolate of extended-spectrum-ß-lactamase-producing Klebsiella quasipneumoniae subsp. similipneumoniae 06-219 with hypermucoviscosity phenotypes obtained from a urine culture of an adult patient was used for whole-genome sequencing. Here, we report the draft genome sequences of this strain, consisting of 53 contigs with an ~5.6-Mb genome size and an average G+C content of 57.36%. The annotation revealed 6,622 coding DNA sequences and 77 tRNA genes.

Identification of OXA-23 carbapenemases: novel variant OXA-239 in Acinetobacter baumannii ST758 clinical isolates in Mexico.

A collection of 15 carbapenem-resistance Acinetobacter baumannii clinical isolates was analysed on two tertiary hospitals in Mexico. The OXA-51 was identified in all isolates, followed by OXA-239 and OXA-58; OXA-239 is described as a new OXA-23-like allele. These carbapenemases were identified on four clonal groups, distributed between two neighbouring hospitals. Acinetobacter baumannii is poorly studied in Mexico; this situation urges the implementation of strategies to prevent its dissemination.

Identification of SHV-type and CTX-M-12 extended-spectrum beta-lactamases (ESBLs) in multiresistant Enterobacteriaceae from Colombian Caribbean hospitals.

The production of extended-spectrum beta-lactamases (ESBLs) in Enterobacteriaceae is the most prevalent resistance mechanism to third-generation cephalosporins. The aim of this study was to identify the ESBLs produced in Escherichia coli and Klebsiella pneumoniae clinical isolates from two hospitals of the Colombian Caribbean Region. A total of 30 clinical isolates of K. pneumoniae (21) and E. coli (9) ESBL-producers were collected in two hospitals from January, 2001 to June, 2003. Isoelectric point values were indicative of SHV-, and CTX-M-type beta-lactamases. PCR amplification and sequencing of SHV genes revealed that SHV-12 was the most prevalent ESBL followed by SHV-5, SHV-2a, the novel SHV-86 and CTX-M-12. There was a geographic distribution of two particular PFGE subtypes in these two distant hospitals. Clonal and horizontal dissemination of resistance was observed.

The blaSHV-5 gene is encoded in a compound transposon duplicated in tandem in Enterobacter cloacae.

The presence of bla(SHV-5) is described in a compound transposon, duplicated in tandem and flanked by IS26 copies on a 70-kb conjugative plasmid (pHNM1), in an Enterobacter cloacae strain associated with a nosocomial outbreak that occurred in Mexico.

Outbreak of infection by extended-spectrum beta-lactamase SHV-5-producing Serratia marcescens in a Mexican hospital.

Previous outbreaks caused by Serratia marcescens have been associated with contaminated medical equipment, intravenous fluids and inadequate hygiene. We carried out the molecular characterization of an outbreak produced by a cephalosporin-resistant S. marscescens that occurred in a Mexican hospital in August 1999. The lethality of this outbreak was 26%. Positive isolates were collected from 20 patients, one medical staff and three chlorhexidine disinfectant solutions. Results of PFGE, beta-lactamase patterns, sequencing of PCR amplifications, plasmid profiles, and mating experiments showed that the outbreak occurred by the dissemination of a S. marcescens SHV-5 producing strain. The adequate enforcement of procedures under the supervision of an infection control resulted in the abrupt end of the outbreak.

Metallo-beta-lactamase gene bla(IMP-15) in a class 1 integron, In95, from Pseudomonas aeruginosa clinical isolates from a hospital in Mexico.

During 2003, 40 carbapenem-resistant Pseudomonas aeruginosa clinical isolates collected in a Mexican tertiary-care hospital were screened for metallo-beta-lactamase production. Thirteen isolates produced IMP-15, and 12 had a single pulsed-field gel electrophoresis pattern. The bla(IMP-15) gene cassette was inserted in a plasmid-borne integron with a unique array of gene cassettes and was named In95.

Outbreak of infection with extended-spectrum beta-lactamase-producing Klebsiella pneumoniae in a Mexican hospital.

Thirty-one strains of Klebsiella pneumoniae (including 10 duplicates) from 21 septicemic pediatric patients (age, <2 months) were studied during a 4-month period (June to October 1996) in which the fatality rate was 62% (13 of 21). These isolates identified by the API 20E system yielded the same biotype. Pulsed-field gel electrophoresis experiments revealed the same clone in 31 strains. The isolates were multidrug-resistant but were still susceptible to ciprofloxacin, imipenem, and cefoxitin. A 135-kb plasmid was harbored in all of the isolates. No transconjugants were obtained that were resistant to ampicillin, cefotaxime, tetracycline, or gentamicin. Isoelectric focusing for beta-lactamases was performed on all strains, and three bands with pIs of 5.4, 7.6, and 8.2 were obtained. Of these, the pI 8.2 beta-lactamase had an extended-spectrum beta-lactamase phenotype. PCR amplification of both TEM- and SHV-type genes was obtained. The sequence analysis of the SHV PCR product indicated a mutation corresponding to the SHV-5 beta-lactamase.

TLA-1: a new plasmid-mediated extended-spectrum beta-lactamase from Escherichia coli.

Escherichia coli R170, isolated from the urine of an infected patient, was resistant to expanded-spectrum cephalosporins, aztreonam, ciprofloxacin, and ofloxacin but was susceptible to amikacin, cefotetan, and imipenem. This particular strain contained three different plasmids that encoded two beta-lactamases with pIs of 7.0 and 9.0. Resistance to cefotaxime, ceftazidime, aztreonam, trimethoprim, and sulfamethoxazole was transferred by conjugation from E. coli R170 to E. coli J53-2. The transferred plasmid, RZA92, which encoded a single beta-lactamase, was 150 kb in length. The cefotaxime resistance gene that encodes the TLA-1 beta-lactamase (pI 9.0) was cloned from the transconjugant by transformation to E. coli DH5alpha. Sequencing of the bla(TLA-1) gene revealed an open reading frame of 906 bp, which corresponded to 301 amino acid residues, including motifs common to class A beta-lactamases: (70)SXXK, (130)SDN, and (234)KTG. The amino acid sequence of TLA-1 shared 50% identity with the CME-1 chromosomal class A beta-lactamase from Chryseobacterium (Flavobacterium) meningosepticum; 48.8% identity with the VEB-1 class A beta-lactamase from E. coli; 40 to 42% identity with CblA of Bacteroides uniformis, PER-1 of Pseudomonas aeruginosa, and PER-2 of Salmonella typhimurium; and 39% identity with CepA of Bacteroides fragilis. The partially purified TLA-1 beta-lactamase had a molecular mass of 31.4 kDa and a pI of 9.0 and preferentially hydrolyzed cephaloridine, cefotaxime, cephalothin, benzylpenicillin, and ceftazidime. The enzyme was markedly inhibited by sulbactam, tazobactam, and clavulanic acid. TLA-1 is a new extended-spectrum beta-lactamase of Ambler class A.

RegulonDB (version 2.0): a database on transcriptional regulation in Escherichia coli.

RegulonDB version 2.0, a database on transcriptional regulation and operon organization in Escherichia coli, is now available on the web at the following URL: http://www.cifn.unam. mx/Computational_Biology/regulondb/. In this paper we describe the main computational changes to the database, which include migrating the database to Sybase, providing graphical descriptions of the internal organization of operons and regulons, and direct links to MEDLINE references. The web interface offers searching either by mechanisms of regulation or by operon organization. The results of a search (operon organization, or site collection) are displayed as hypertext, and can also be displayed graphically. In terms of its contents, RegulonDB contains a large number of operons, as well as the absolute position in the completed genome sequence of sites, promoters, and individual genes of E.coli.