Neil Woodford

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.

Rapid evolution and spread of carbapenemases among Enterobacteriaceae in Europe

Plasmid-acquired carbapenemases in Enterobacteriaceae, which were first discovered in Europe in the 1990s, are now increasingly being identified at an alarming rate. Although their hydrolysis spectrum may vary, they hydrolyse most β-lactams, including carbapenems. They are mostly of the KPC, VIM, NDM and OXA-48 types. Their prevalence in Europe as reported in 2011 varies significantly from high (Greece and Italy) to low (Nordic countries). The types of carbapenemase vary among countries, partially depending on the cultural/population exchange relationship between the European countries and the possible reservoirs of each carbapenemase. Carbapenemase producers are mainly identified among Klebsiella pneumoniae and Escherichia coli, and still mostly in hospital settings and rarely in the community. Although important nosocomial outbreaks with carbapenemase-producing Enterobacteriaceae have been extensively reported, many new cases are still related to importation from a foreign country. Rapid identification of colonized or infected patients and screening of carriers is possible, and will probably be effective for prevention of a scenario of endemicity, as now reported for extended-spectrum β-lactamase (mainly CTX-M) producers in all European countries.

Identification of Acinetobacter baumannii by Detection of the blaOXA-51-like Carbapenemase Gene Intrinsic to This Species

ABSTRACT bla OXA-51-like was sought in clinical isolates of Acinetobacter species in a multiplex PCR, which also detects blaOXA-23-like and class 1 integrase genes. All isolates that gave a band for blaOXA-51-like identified as A. baumannii. This gene was detected in each of 141 isolates of A. baumannii but not in those of 22 other Acinetobacter species.

Outbreak of Klebsiella pneumoniae Producing a New Carbapenem-Hydrolyzing Class A β-Lactamase, KPC-3, in a New York Medical Center

ABSTRACT From April 2000 to April 2001, 24 patients in intensive care units at Tisch Hospital, New York, N.Y., were infected or colonized by carbapenem-resistant Klebsiella pneumoniae. Pulsed-field gel electrophoresis identified a predominant outbreak strain, but other resistant strains were also recovered. Three representatives of the outbreak strain from separate patients were studied in detail. All were resistant or had reduced susceptibility to imipenem, meropenem, ceftazidime, piperacillin-tazobactam, and gentamicin but remained fully susceptible to tetracycline. PCR amplified a blaKPC allele encoding a novel variant, KPC-3, with a His(272)→Tyr substitution not found in KPC-2; other carbapenemase genes were absent. In the outbreak strain, KPC-3 was encoded by a 75-kb plasmid, which was transferred in vitro by electroporation and conjugation. The isolates lacked the OmpK35 porin but expressed OmpK36, implying reduced permeability as a cofactor in resistance. This is the third KPC carbapenem-hydrolyzing β-lactamase variant to have been reported in members of the Enterobacteriaceae, with others reported from the East Coast of the United States. Although producers of these enzymes remain rare, the progress of this enzyme group merits monitoring.

Rapid detection of the O25b-ST131 clone of Escherichia coli encompassing the CTX-M-15-producing strains

OBJECTIVES Recently, a CTX-M-15 extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli O25b-ST131 clone, belonging to the B2 phylogenetic group and with a high virulence potential, has been reported all over the world, representing a major public health problem. The present study was carried out to develop a rapid and simple detection assay that identifies members of this clone. METHODS A total of 627 E. coli isolates of which 373 produced an ESBL, collected across four continents, were screened using a O25b-ST131 clone allele-specific PCR for the pabB gene. RESULTS One hundred and forty-three ESBL isolates were found positive with the assay. These isolates were all of O25b type and, when studied by multilocus sequence typing (25 cases), were all of ST131. The O25b-ST131 clone was found to produce ESBLs other than CTX-M-15, specifically CTX-M-2, -3, -14, -27, -32 and -61 as well as TEM-24. This clone represents 3% of non-ESBL B2 isolates originating from urinary tract infections in Paris. CONCLUSIONS We have developed a PCR-based assay that easily identifies a clone with high likelihood of producing ESBLs, including CTX-M-15.

Molecular mechanisms disrupting porin expression in ertapenem-resistant Klebsiella and Enterobacter spp. clinical isolates from the UK

OBJECTIVES The aim of this study was to investigate relatedness and molecular mechanism(s) of ertapenem resistance in clinical isolates of Klebsiella spp. (n = 28) and Enterobacter spp. (n = 27) referred from multiple hospitals to the UK national reference laboratory. METHODS Investigations included genotyping by PFGE, resistance gene analysis by PCR and antimicrobial susceptibility testing with and without inhibitors of efflux and beta-lactamase activity. Outer membrane proteins (OMPs) were profiled by SDS-PAGE; porin genes were sequenced and their expression was examined by RT-PCR. The contribution of porin deficiency to resistance was investigated by restoring functional porin genes on plasmids. RESULTS PFGE showed significant clonal diversity among ertapenem-resistant isolates, with only small clusters identified. SHV- and CTX-M-type extended-spectrum beta-lactamases were identified in the Klebsiella spp. isolates, whereas AmpC overexpression or KPC carbapenemase was detected in the Enterobacter cloacae isolates. SDS-PAGE showed that Klebsiella pneumoniae and Enterobacter aerogenes with high-level ertapenem resistance (MICs > or = 16 mg/L) consistently lacked both of the two major non-specific porins, whereas variable patterns of OmpC and OmpF were seen in E. cloacae with lower-level ertapenem resistance. Various point mutations or insertion sequences were identified as disrupting the porin-coding sequences, as well as mutations in the promoter region. Functional restoration of OmpK35 or OmpK36 in Klebsiella and OmpC or OmpF in Enterobacter spp. isolates significantly decreased the MICs of all carbapenems, but particularly of ertapenem. We found no evidence of efflux contributing to resistance. CONCLUSIONS Ertapenem resistance was exclusively due to combinations of beta-lactamases with impermeability caused by loss of OMPs. Efflux was not implicated and there was no national spread of resistant clones.

Identification and screening of carbapenemase-producing Enterobacteriaceae

Carbapenem-hydrolysing β-lactamases are the most powerful β-lactamases, being able to hydrolyse almost all β-lactams. They are mostly of the KPC, VIM, IMP, NDM and OXA-48 types. Their current extensive spread worldwide in Enterobacteriaceae is an important source of concern, as these carbapenemase producers are multidrug-resistant. Detection of infected patients and of carriers are the two main approaches for prevention of their spread. Phenotypic and molecular-based techniques are able to identify these carbapenemase producers, although with variable efficiencies. The detection of carriers still relies mostly on the use of screening culture media.

Complete Nucleotide Sequences of Plasmids pEK204, pEK499, and pEK516, Encoding CTX-M Enzymes in Three Major Escherichia coli Lineages from the United Kingdom, All Belonging to the International O25:H4-ST131 Clone

ABSTRACT We determined the complete nucleotide sequences of three plasmids that encode CTX-M extended-spectrum β-lactamases (ESBLs) in pulsed-field gel electrophoresis-defined United Kingdom variants (strains A, C, and D) of the internationally prevalent Escherichia coli O25:H4-ST131 clone. Plasmid pEK499 (strain A; 117,536 bp) was a fusion of type FII and FIA replicons and harbored the following 10 antibiotic resistance genes conferring resistance to eight antibiotic classes: blaCTX-M-15, blaOXA-1, blaTEM-1,aac6′-Ib-cr, mph(A), catB4, tet(A), and the integron-borne dfrA7, aadA5, and sulI genes. pEK516 (strain D; 64,471 bp) belonged to incompatibility group IncFII and carried seven antibiotic resistance genes: blaCTX-M-15, blaOXA-1, blaTEM-1, aac6′-Ib-cr, catB4, and tet(A), all as in pEK499. It also carried aac3-IIa, conferring gentamicin resistance, and was highly related to pC15-1a, a plasmid encoding the CTX-M-15 enzyme in Canada. By contrast, pEK204 (strain C; 93,732 bp) belonged to incompatibility group IncI1 and carried only two resistance genes, blaCTX-M-3 and blaTEM-1. It probably arose by the transposition of Tn3 and ISEcp1-blaCTX-M-3 elements into a pCOLIb-P9-like plasmid. We conclude that (i) United Kingdom variants of the successful E. coli ST131 clone have acquired different plasmids encoding CTX-M ESBLs on separate occasions, (ii) the blaCTX-M-3 and blaCTX-M-15 genes on pEK204 and pEK499/pEK516 represent separate escape events, and (iii) IncFII plasmids harboring blaCTX-M-15 have played a crucial role in the global spread of CTX-M-15 ESBLs in E. coli.

Editorial: Assessing the antimicrobial susceptibility of bacteria obtained from animals

The accurate performance of antimicrobial susceptibility testing of bacteria from animal sources and the correct presentation of the results is a complex matter. A review of the published literature revealed a number of recurring errors with regard to methodology, quality control, appropriate interpretive criteria, and calculation of MIC(50) and MIC(90) values. Although more subjective, there is also no consensus regarding the definition of multiresistance. This Editorial is intended to provide guidance to authors on how to avoid these frequently detected shortcomings.

Global spread of antibiotic resistance: the example of New Delhi metallo-β-lactamase (NDM)- mediated carbapenem resistance

The rapidity with which new types of antibiotic resistance can disseminate globally following their initial emergence or recognition is exemplified by the novel carbapenemase New Delhi metallo-β-lactamase (NDM). The first documented case of infection caused by bacteria producing NDM occurred in 2008, although retrospective analyses of stored cultures have identified the gene encoding this enzyme (blaNDM) in Enterobacteriaceae isolated in 2006. Since its first description, NDM carbapenemase has been reported from 40 countries worldwide, encompassing all continents except South America and Antarctica. The spread of NDM has a complex epidemiology involving the spread of a variety of species of NDM-positive bacteria and the inter-strain, inter-species and inter-genus transmission of diverse plasmids containing blaNDM, with the latter mechanism having played a more prominent role to date. The spread of NDM illustrates that antibiotic resistance is a public health problem that transcends national borders and will require international cooperation between health authorities if it is to be controlled.