Rachel Pike

Emergence of a new antibiotic resistance mechanism in India, Pakistan, and the UK: a molecular, biological, and epidemiological study.

Summary Background Gram-negative Enterobacteriaceae with resistance to carbapenem conferred by New Delhi metallo-β-lactamase 1 (NDM-1) are potentially a major global health problem. We investigated the prevalence of NDM-1, in multidrug-resistant Enterobacteriaceae in India, Pakistan, and the UK. Methods Enterobacteriaceae isolates were studied from two major centres in India—Chennai (south India), Haryana (north India)—and those referred to the UKs national reference laboratory. Antibiotic susceptibilities were assessed, and the presence of the carbapenem resistance gene blaNDM-1 was established by PCR. Isolates were typed by pulsed-field gel electrophoresis of XbaI-restricted genomic DNA. Plasmids were analysed by S1 nuclease digestion and PCR typing. Case data for UK patients were reviewed for evidence of travel and recent admission to hospitals in India or Pakistan. Findings We identified 44 isolates with NDM-1 in Chennai, 26 in Haryana, 37 in the UK, and 73 in other sites in India and Pakistan. NDM-1 was mostly found among Escherichia coli (36) and Klebsiella pneumoniae (111), which were highly resistant to all antibiotics except to tigecycline and colistin. K pneumoniae isolates from Haryana were clonal but NDM-1 producers from the UK and Chennai were clonally diverse. Most isolates carried the NDM-1 gene on plasmids: those from UK and Chennai were readily transferable whereas those from Haryana were not conjugative. Many of the UK NDM-1 positive patients had travelled to India or Pakistan within the past year, or had links with these countries. Interpretation The potential of NDM-1 to be a worldwide public health problem is great, and co-ordinated international surveillance is needed. Funding European Union, Wellcome Trust, and Wyeth.

Occurrence of Carbapenem-Resistant Acinetobacter baumannii Clones at Multiple Hospitals in London and Southeast England

ABSTRACT From late 2003 to the end of 2005, the Health Protection Agencys national reference laboratories received approximately 1,600 referrals of Acinetobacter spp., including 419 and 58 examples, respectively, of two carbapenem-resistant Acinetobacter baumannii lineages, designated OXA-23 clones 1 and 2. Representatives of these clones were obtained from 40 and 8 hospitals, respectively, in London or elsewhere in Southeast England. Both clones had blaOXA-23-like genes, as well as the intrinsic (but downregulated) blaOXA-51-like carbapenemase genes typical of A. baumannii. Both were highly multiresistant: only colistin and tigecycline remained active versus OXA-23 clone 1 isolates; OXA-23 clone 2 isolates were also susceptible to amikacin and minocycline. These lineages increase the burden created by the southeast (SE) clone, a previously reported A. baumannii lineage with variable carbapenem resistance contingent on upregulation of the blaOXA-51-like gene. Known since 2000, the SE clone had been referred from over 40 hospitals by the end of 2005, with 627 representatives received by the reference laboratories. The OXA-23 clone 2 is now in decline, but OXA-23 clone 1 continues to be referred from new sites, as does the SE clone. Their spread is forcing the use of unorthodox therapies, principally colistin and tigecycline, although the optimal regimens remain uncertain.

Detection and Typing of Integrons in Epidemic Strains of Acinetobacter baumannii Found in the United Kingdom

ABSTRACT Integrons were sought in Acinetobacter isolates from hospitals in the United Kingdom by integrase gene PCR. Isolates were compared by pulsed-field gel electrophoresis, and most belonged to a small number of outbreak strains or clones of A. baumannii, which are highly successful in the United Kingdom. Class 1 integrons were found in all of the outbreak isolates but in none of the sporadic isolates. No class 2 integrons were found. Three integrons were identified among the main outbreak strains and clones. While a particular integron was usually associated with a strain or clone, some members carried a different integron. Some integrons were associated with more than one strain. The cassette arrays of two of the integrons were very similar, both containing gene aacC1, which confers resistance to gentamicin, two open reading frames coding for unknown products (orfX, orfX′), and gene aadA1a, which confers resistance to spectinomycin and streptomycin. The larger of these integrons had two copies of the first (orfX) of the gene cassettes coding for unknown products. The third integron, with a cassette array containing gene aacA4, which codes for amikacin, netilmicin, and tobramycin resistance; a chloramphenicol acetyltransferase, catB8; and gene aadA1, conferring resistance to spectinomycin and streptomycin, was associated with an OXA-23 carbapenemase-producing clone, which has spread rapidly in hospitals in the United Kingdom during 2003 and 2004. These integron cassette arrays have been found in other outbreak strains of A. baumannii from other countries. We conclude that integrons are useful markers for epidemic strains of A. baumannii and that integron typing provides valuable information for epidemiological studies.

Phylogenetic diversity of Escherichia coli strains producing NDM-type carbapenemases

BACKGROUND The global accumulation of Escherichia coli with CTX-M extended-spectrum β-lactamases partly reflects the dissemination of clonal lineages, notably ST131 and ST405. More recently, E. coli have emerged that produce NDM carbapenemase. We sought to determine the clonal diversity of E. coli with this enzyme from English hospitals, and to compare them with isolates from Pakistan and India. METHODS The 18 NDM-positive E. coli were from hospitals in England (n = 10), Pakistan (n = 7) and India (n = 1). Isolates were compared by phylogenetic grouping, multilocus sequence typing and PFGE of XbaI-digested DNA. Isolates were screened by PCR for acquired AmpC genes, bla(CTX-M), and the 16S rRNA methylase genes armA and rmtC. RESULTS Most of the isolates belonged to phylogenetic groups B1 (n = 9) or D (n = 7); two were group A and none was group B2. Nine isolates from England and Pakistan belonged to the B1 lineage ST101, with seven of these clustering at >77% similarity by PFGE. Other lineages included ST405 (n = 3, group D), ST648 (n = 3, group D), the ST23 complex (one each of ST90 and ST410, both group A) and ST156 (n = 1, group D). Sixteen of 18 isolates had a group 1 CTX-M gene, 13 had a CIT-type acquired AmpC, and 16 had either or both of armA and rmtC. CONCLUSIONS The E. coli isolates producing NDM-1 carbapenemase belonged to six sequence types and included diverse clonal lineages. Nevertheless, isolates of B1-ST101 accounted for half the collection, and included isolates from both England and Pakistan. None of the isolates belonged to ST131 or to phylogroup B2.

Incidence of Acinetobacter Species Other than A. baumannii among Clinical Isolates of Acinetobacter: Evidence for Emerging Species

ABSTRACT Six hundred ninety nonduplicate isolates of Acinetobacter species were identified using a combination of detection of bla OXA-51-like and rpoB sequence cluster analysis. Although most isolates were identified as A. baumannii (78%), significant numbers of other species, particularly A. lwoffii/genomic species 9 (8.8%), A. ursingii (4%), genomic species 3 (1.7%), and A. johnsonii (1.7%), were received, often associated with bacteremias.

Comparison of Acinetobacter baumannii Isolates from the United Kingdom and the United States That Were Associated with Repatriated Casualties of the Iraq Conflict

ABSTRACT Acinetobacter isolates associated with casualties from the Iraq conflict from the United States were compared with those from the United Kingdom by pulsed-field gel electrophoresis and integron analysis. Representatives of the main outbreak strain associated with casualties from both countries were indistinguishable in DNA profile. Two further outbreak strains were common to both sets of isolates.

Characterization of Enterobacteriaceae producing OXA-48-like carbapenemases in the UK

OBJECTIVES To characterize UK clinical isolates of Enterobacteriaceae producing OXA-48-like carbapenemases and to compare their resistance plasmids. METHODS Twenty-six enterobacteria producing OXA-48-like enzymes were studied. These were from 22 diverse hospitals in the UK. Isolates of Escherichia coli and Klebsiella pneumoniae were assigned to clonal lineages by multilocus sequence typing. Carbapenemase genes and their genetic environments were characterized by PCR and sequencing. Resistance plasmids were transferred by transformation or conjugation and compared by restriction analysis and PCR for genes encoding critical plasmid functions. RESULTS Thirteen isolates of K. pneumoniae, 10 E. coli and 2 Enterobacter cloacae harboured a classical bla(OXA-48) gene; the K. pneumoniae isolates belonged to 11 sequence types (STs) and the E. coli to 7 STs, including ST131 and ST38. The bla(OXA-48) genes were located within either Tn1999 or Tn1999.2 transposons on related ≈ 50 kb or ≈ 62 kb plasmids, which lacked other resistance genes or, in one isolate, on an ≈ 140 kb plasmid that also encoded OXA-9 and CTX-M group-9 β-lactamases. One India-linked K. pneumoniae isolate had a bla(OXA-181) gene in association with an ISEcp1 insertion sequence on a 7 kb plasmid. CONCLUSIONS Horizontal transfer of related plasmids has facilitated the spread of OXA-48 carbapenemase into multiple strains of several Enterobacteriaceae species. The clonal diversity of the producers suggests repeated introduction into the UK. Low carbapenem MICs for some producers complicates detection and creates a risk for unrecognized spread.

Antimicrobial treatment and clinical outcome for infections with carbapenem- and multiply-resistant Acinetobacter baumannii around London

Carbapenem- and multiply-resistant Acinetobacter baumannii (C-MRAB) are challenging pathogens, often susceptible only to polymyxins and tigecycline. We reviewed clinical outcomes in relation to antibiotic treatment for 166 consecutive patients infected or colonised with these organisms at 18 hospitals around London, UK. Clinical data were obtained along with the isolates, which were typed by pulsed-field gel electrophoresis (PFGE). Outcomes were compared for colonised and infected patients and in relation to treatment, with associations examined by logistic regression. Most subjects (103/166; 62%) were in Intensive Care Units (ICUs) or high dependency units; 84 (50.6%) were judged to be infected and 73 (44.0%) were colonised, with 9 indeterminate. Among the 166 C-MRAB isolates, 141 belonged to OXA-23 clone 1, a European clone II lineage. Survival rates among infected and colonised patients were 68% and 67%, respectively (P > 0.05), indicating little attributable mortality. Univariate and multivariate analyses indicated poorer outcomes among ICU-infected patients and those with pulmonary infection or bacteraemia, whereas trauma patients had significantly better outcomes than the generality. Outcomes varied with hospital, even in multivariate analysis, reflecting either differences in management or case mix. There was little association between outcome and therapy with colistin and/or tigecycline except that, among patients with respiratory infection, 12/15 treated with intravenous colistin alone had poor outcome compared with 1/8 whose therapy include nebulised colistin. This difference was significant (P=0.003), although the patients receiving nebulised drug were mostly younger, included trauma cases and were at a hospital with good outcomes.

Linezolid-resistant ST36 methicillin-resistant Staphylococcus aureus associated with prolonged linezolid treatment in two paediatric cystic fibrosis patients

OBJECTIVES To describe the emergence of linezolid-resistant methicillin-resistant Staphylococcus aureus (MRSA) of sequence type (ST)36 lineage in two paediatric patients with cystic fibrosis, after long-term low-dose linezolid treatment. METHODS Two paediatric males with cystic fibrosis had sputum samples quantitatively cultured during hospitalization. After the isolation of MRSA from both patients, oral treatment with 300 mg linezolid twice daily was initiated for periods of 1-2 months separated by up to 6 months. Isolates cultured 9 months after the start of treatment were tested for resistance to linezolid by agar dilution (BSAC). Resistant isolates were examined for 23S rDNA mutations, and typed by phage and macrorestriction with SmaI. Isolates from follow-up sputum samples were obtained until 44-51 months after treatment with linezolid. RESULTS Colonization with MRSA was at a density of approximately 10(6) cfu/mL sputum for both subjects. Initial isolates were susceptible to linezolid, but, 9 months later, isolates from both patients were resistant (MICs > 16 mg/L). Both isolates were epidemic MRSA-16 variant A1 (ST36-MRSA-II), which is widespread in UK hospitals. Both isolates were heterozygous for a G2576T mutation in their 23S rDNA genes, but one was resistant to fusidic acid and tetracycline. In follow-up sampling, the younger patient yielded linezolid-resistant EMRSA-16 for a further 42 months, whilst the other lost the linezolid-resistant MRSA and had alternately Pseudomonas aeruginosa or linezolid-susceptible EMRSA-16 variant A1 isolated over 35 further months. CONCLUSIONS Linezolid resistance emerged in two isolates of ST36 MRSA colonizing the lungs of two paediatric cystic fibrosis patients. Subtherapeutic levels of linezolid may have facilitated the selection of resistance.

NDM carbapenemases in the United Kingdom: an analysis of the first 250 cases

OBJECTIVES Gram-negative bacteria with diverse carbapenemases, including New Delhi metallo-β-lactamase (NDM) enzymes, have been increasingly recorded in the UK since 2007. We analysed patient data for NDM-positive isolates confirmed by the national reference laboratory from UK laboratories from February 2008 to July 2013. METHODS Isolates resistant to carbapenems and with imipenem MICs reduced ≥8-fold by EDTA were tested by PCR for genes encoding acquired class B carbapenemases. MICs were determined by BSAC agar dilution methodology. When requested by the sender, or when they were members of apparent clusters, NDM-positive isolates were typed by variable number tandem repeat (VNTR) analysis or PFGE. Data provided by the sending laboratories were collated and reviewed. RESULTS From February 2008 to July 2013 the reference laboratory confirmed 326 NDM-positive isolates from 250 patients, submitted by 83 laboratories. Most (85%, 213/250) patients were already hospitalized when the NDM-positive bacteria were detected, were male (61%, 152/250) and were aged >60 years (58%, 145/250). Travel history was available for only 40% of patients, but 52% (53/101) of these had documented healthcare contact within or travel to the Indian subcontinent. Most NDM-positive isolates (94%, 306/326) were Enterobacteriaceae with just 6% (20/326) non-fermenters; the predominant hosts were Klebsiella spp. (55%, 180/326) and Escherichia coli (25%, 80/326). Almost all NDM-positive isolates were resistant to multiple antibiotic classes, but 90% remained susceptible to colistin. CONCLUSIONS Gram-negative bacteria with NDM carbapenemases are a growing challenge, especially for elderly hospitalized patients, including those with healthcare contact in the Indian subcontinent, and leave few therapeutic options. UK outbreaks remain rare and contained.