16S rRNA Methylase Genes Research Articles

Characterization of multidrug-resistant Klebsiella pneumoniae from Australia carrying blaNDM-1

blaNDM genes, encoding metallo-β-lactamases providing resistance to carbapenems, have been reported in many locations since the initial report in 2008, including in several Enterobacteriaceae isolates in Australia/New Zealand. Here, we compare 4 additional carbapenem-resistant Klebsiella pneumoniae carrying blaNDM-1 isolated in Australia. Two are sequence type ST147, previously associated with blaNDM in Australia and elsewhere. They carry blaNDM-1 and different 16S rRNA methylase genes (armA or rmtC) on different conjugative plasmids, in 1 case with an IncFIIY replicon. One isolate belongs to the globally important ST11 but did not transfer a plasmid to Escherichia coli. The fourth isolate belongs to the novel ST1068 and transferred blaNDM-1, armA, and an IncA/C plasmid. Amplification and sequencing of ompK porin genes suggest that, unlike the case for other carbapenemase genes, ompK36 defects may not be required for NDM to cause clinically relevant levels of carbapenem resistance.

P129 Duration of colonisation with extended-spectrum beta-lactamase-producing enterobacteria

The emergence of multidrug-resistant (MDR) Acinetobacter baumannii has become a serious medical problem worldwide. To clarify the genetic and epidemiological properties of MDR A. baumannii strains isolated from a medical setting in Nepal, 246 Acinetobacter spp. isolates obtained from different patients were screened for MDR A. baumannii by antimicrobial disk susceptibility testing. Whole genomes of the MDR A. baumannii isolates were sequenced by MiSeq™ (Illumina), and the complete genome of one isolate (IOMTU433) was sequenced by PacBio RS II. Phylogenetic trees were constructed from single nucleotide polymorphism concatemers. Multilocus sequence types were deduced and drug resistance genes were identified. Of the 246 Acinetobacter spp. isolates, 122 (49.6%) were MDR A. baumannii, with the majority being resistant to aminoglycosides, carbapenems and fluoroquinolones but not to colistin and tigecycline. These isolates harboured the 16S rRNA methylase gene armA as well as blaNDM-1, blaOXA-23 or blaOXA-58. MDR A. baumannii isolates belonging to clonal complex 1 (CC1) and CC2 as well as a novel clonal complex (CC149) have spread throughout a medical setting in Nepal. The MDR isolates harboured genes encoding carbapenemases (OXA and NDM-1) and a 16S rRNA methylase (ArmA).

The First Report of CMY, aac (6′)-Ib and 16S rRNA Methylase Genes Among Pseudomonas aeruginosa Isolates From Iran

Background:: Serious infections by Pseudomonas aeruginosa are commonly treated with the combination of a beta-lactam antibacterial and an aminoglycoside. Therefore, production of a 16S rRNA methylase may result in an extremely important antibacterial resistance profile. Objectives:: The present study was conducted to determine the prevalence of Cephamycinase (CMY), aminoglycoside 6'-N-acetyltransferase (aac(6′)-Ib) and 16S rRNA methylase genes among Pseudomonas aeruginosa isolates from Iran. Patients and Materials:: This descriptive study was performed on hospitalized burnt patients during 2011and 2012. Antibiotics susceptibility tests were performed by disc diffusion and broth microdilution methods. CMY, aac(6′)-Ib, 16S rRNA methylase genes were detected by PCR method. Results:: Seventy-seven (77%) of 100 isolates were resistant to Imipenem and Ceftazidime. aac(6)-Ib, Cephamycinase (CMY), and rRNA methyltransferase (rmtB and rmtD) were detected in 57 (74.02%), 7 (9.09%), 11 (14.28%), and 9 (11.68%) isolates, respectively. PCR results for aminoglycoside resistance methyltransferase (armA) and rRNA methyltransferase (rmtC) were all negative. Aminoglycoside resistance methyltransferase (armA), and rRNA methyltransferase (rmtC) were not detected. Conclusions:: This study detected multiple drug resistance in Pseudomonas aeruginosa including resistance to β-lactams, Aminoglycosides, and Fluoroquinolones. Therefore, identification of drug resistance patterns in P. aeruginosa and detection of pan-resistant producing isolates are of great importance in prevention and control of infections in burn center ward.

Co-existence of bla OXA-23 and armA in multidrug-resistant Acinetobacter baumannii isolated from a hospital in South Korea

The co-existence of carbapenemase, 16S rRNA methylase and mutated quinolone resistance-determining regions (QRDRs) can cause serious difficulty in treating infections with multidrug-resistant Acinetobacter baumannii. In this study, we aimed to determine the mechanisms of imipenem, amikacin and ciprofloxacin resistance in A. baumannii isolates with resistance to these antibiotics. A total of 31 non-duplicate isolates of amikacin- and ciprofloxacin-resistant Acinetobacter isolates were identified from April to August 2010 from a single hospital in South Korea. To assess the clonal relatedness of the 31 Acinetobacter isolates, multilocus sequence typing, network phylogenetic analysis and enterobacterial repetitive intergenic consensus-PCR were utilized. Detection of OXA-type carbapenemase and 16S rRNA methylase was conducted using a multiplex PCR assay. The QRDRs of the gyrA and parC genes were amplified and sequenced. The result showed that 30/31 isolates harboured the blaOXA-23-like carbapenemase, which made them resistant to imipenem (MICs ≥16 µg ml(-1)). Twenty-eight of the 31 isolates were found to possess armA, a 16S rRNA methylase gene, and showed resistance to amikacin, arbekacin, gentamicin and tobramycin (MICs >256 µg ml(-1)). All of the isolates were determined to carry QRDR mutations in both gyrA and parC: a Ser83Leu substitution in gyrA and a Ser80Leu substitution in parC, causing a ciprofloxacin MIC ≥64 µg ml(-1). In conclusion, A. baumannii with co-existence of carbapenemase, 16S rRNA methylase and mutated QRDRs are extremely prevalent in South Korea, which may cause serious problems in the treatment of A. baumannii infections using carbapenem, amikacin and ciprofloxacin.

Detection of plasmid-mediated quinolone resistance determinants and the emergence of multidrug resistance in clinical isolates of Shigella in SiXian area, China

A total of 123 Shigella isolates were collected from SiXian area in Anhui, China. Screening was carried out by polymerase chain reaction (PCR) amplification of plasmid-mediated quinolone resistance (PMQR) determinants. Different β-blactamases genes, plasmid-borne blaAmpC, 16S rRNA methylase genes, integrons, and mutations in quinolone resistance-determining regions were analysed by PCR for the PMQR-positive isolates.

OXA-181-producing Klebsiella pneumoniae establishing in Singapore

BackgroundCarbapenemase producing Enterobacteriaceae are becoming a major public health concern globally, however, relatively little is known about the molecular and clinical epidemiology of these organisms in many parts of the world.MethodsAs part of a laboratory surveillance programme, 96 carbapenem non-susceptible Enterobacteriaceae isolates from clinical samples from patients in seven hospitals were referred for investigation for carbapenemases. Using polymerase chain reaction (PCR) to screen for a collection of genes encoding carbapenemases, 33 of 96 (34.5%) isolates were confirmed as carbapenemase producers. NDM-1 producers were the most prevalent at 64% (21/33) whilst OXA-181 was the second most common carbapenemase constituting 24.5% (8/33) of the carbapenemase producing isolates. Seven of these eight OXA-181 positive isolates underwent further characterisation with screening for other transmissible antimicrobial resistance determinants using PCR. Clonal relatedness was explored using Multilocus sequence typing (MLST) and Pulsed Field Gel Electrophoresis (PFGE). Plasmid characterisation was performed including restriction analysis and transfer by conjugation or transformation.ResultsIn addition to the OXA-181 gene, all contained other transmissible resistance determinants including extended spectrum β-lactamases, oxacillinases or 16S rRNA methylase genes, but none contained metallo-β-lactamases or serine carbapenemases. All isolates had a multidrug resistant phenotype with two isolates being resistant to every antibiotic tested including colistin. Multilocus sequence typing confirmed five isolates belonged to ST17 and two to ST14, with those belonging to the same sequence type having identical PFGE profiles. The OXA-181 gene was typically carried on large plasmids which were mostly non-conjugative.ConclusionsOXA-181 carbapenemase appears to be an important and probably under-recognised cause of carbapenem resistance in Enterobacteriaceae in Singapore. Further coordinated research into clinical and molecular epidemiology of carbapenemases is urgently required in Singapore and throughout Asia.

Detection of Pan‐amino glycoside‐resistant Gram negative Bacteria using a Molecular Method

The 16S rRNA methylases confer and cause pan‐aminoglycoside resistance (PAR). In the present prospective study, aminoglycoside‐resistant isolates were subjected to the disc diffusion and agar dilution methods to determine minimum inhibitory concentrations (MICs) to rule out high‐level aminoglycoside resistance (HLAR). One hundred and two isolates had an MIC ≥ 512 μg/ mL. These 102 isolates were investigated for the presence of 16S rRNA methylase genes including armA, rmtA, rmtB, and rmtC by polymerase chain reaction (PCR). Among isolates belonging to Enterobacteriaceae, armA, rmtB, and rmtC were present in 28, 56, and 4 isolates, respectively. Meanwhile, armA, rmtA, rmtB, and rmtC were present in 6, 1, 4, and 2 non‐fermenting Gram‐negative bacilli (NFGNB) isolates, respectively. All Enterobacteriaceae isolates were extended spectrum beta‐lactamase (ESBL) positive. The present results demonstrate the wide dissemination of PAR among NFGNB compared to Enterobacteriaceae. These results are very similar to those in other countries, in which 16s rRNA methylases are common in Enterobacteriaceae. The rapid spread of multi‐drug‐resistant isolates producing both ESBL and 16s rRNA methylases raises clinical concerns and may become a major therapeutic threat in the future.

Prevalence of β-Lactamase and 16S rRNA Methylase Genes Among ClinicalEscherichia coliIsolates Carrying Plasmid-Mediated Quinolone Resistance Genes from Animals

Plasmid-mediated quinolone-resistance (PMQR) genes were determined by polymerase chain reactions (PCRs) in 250 Escherichia coli isolates from food-producing animals in Guangdong, China, in 2009-2010. Then, the prevalence of plasmid-mediated β-lactamase and 16S rRNA methylase genes was determined by PCRs among the PMQR-positive isolates. One hundred fifty-seven (62.8%) isolates were found to harbor at least one PMQR gene, and qnrS (84) and oqxAB (97) were the most two prevalent PMQR genes. β-lactamase (ESBL and/or AmpC type) genes were detected in 106 of the 157 PMQR-positive strains. The bla(TEM-1) (78) was the most prevalent β-lactamase gene in the 157 PMQR-positive isolates, followed by bla(CMY-2) (28), bla(CTX-M) (25), bla(SHV-1) (3), and bla(DHA-1) (3). Twenty-nine were detected to produce more than one type of β-lactamase. The rmtB was the most prevalent 16S rRNA methylase gene detected (11.5%, 18/157), and armA was detected in only two (1.27%, 2/157) isolates, with one isolate coharboring rmtB and armA. Sixteen isolates were found to coharbor the three types of resistance genes detected in this study. Only 1 transconjugant JGDA2 harboring oqxAB, aac(6')-Ib-cr, bla(DHA-1), and rmtB was obtained from the 16 isolates harboring the three types of resistance genes, by conjugation experiment. The results of Southern blot hybridization revealed that oqxAB, bla(DHA-1), and rmtB were colocated on the same plasmid of ∼54 kb in the JGDA2. To our knowledge, this is the first description of the coexistence of the oqxAB, rmtB, and bla(DHA-1) resistance genes on the same plasmid in one E. coli strain.

Detection of 16S rRNA Methylase Genes in Gram-Negative Bacilli Isolated from Hospitals in Changchun, China

Methylation of 16S rRNA is an important mechanism of aminoglycoside resistance among gram-negative pathogens. In this report, 16S rRNA methylase genes were amplified using PCR among gram-negative bacillus isolates from hospitals in the Changchun area of China and 16S rRNA methylase genotypes (armA, rmtB, rmtA, rmtC, rmtD, and npmA) were identified by direct sequencing. Fifty of the isolates (43.1%) harbored 16S rRNA methylase genes. The common 16S rRNA methylase genes were armA and rmtB (12.1% and 31.0%, respectively), whereas the rmtA, rmtC, rmtD, and npmA genes were absent from the sample. It suggests that the predominant 16S rRNA methylase genes among gramnegative bacilli in the Changchun area are armA and rmtB.

Prevalence and characteristics of rmtB and qepA in Escherichia coli isolated from diseased animals in China

16S rRNA methylase and QepA, a fluoroquinolone efflux pump, are new mechanisms of resistance against aminoglycosides and fluoroquinolone, respectively. One of 16S rRNA methylase genes, rmtB, was found to be associated with qepA, were both located on the same transposable element. In this study, we intended to determine the current prevalence and characteristics of the 16S rRNA methylase genes and qepA, and to study the association between rmtB and qepA. A total of 892 Escherichia coli isolates were collected from various diseased food-producing animals in China from 2004 to 2008 and screened by PCR for 16S rRNA methylase genes and qepA. About 12.6% (112/892) and 0.1% (1/892) of isolates that were highly resistant to amikacin were positive for rmtB and armA, respectively. The remaining five 16S rRNA methlyase genes were not detected. Thirty-six (4.0%) strains carried qepA. About 32.1% of rmtB-positive strains harbored qepA, which was not detected in rmtB-negative strains. Most strains were clonally unrelated, while identical PFGE profiles of rmtB-positive isolates were found in the same farm indicating clonal transmission. Conjugation experiments showed that rmtB was transferred to the recipients, and qepA also cotransferred with rmtB in some cases. The spread of E. coli of food animal origin harboring both rmtB and qepA suggests that surveillance for antimicrobial resistance of animal origin as well as the study of the mechanisms of resistance should be undertaken.

Occurrence and characterization of multidrug-resistant New Delhi metallo-β-lactamase-1-producing bacteria isolated between 2003 and 2010 in Bangladesh

The purpose of this study was to screen for reduced susceptibility against imipenem and the presence of the New Delhi metallo-β-lactamase-1 (NDM-1) gene in a collection of Enterobacteriaceae (Escherichia coli, Shigella spp. and Klebsiella pneumoniae) from different surveillance studies between 2003 and 2010 at the International Centre for Diarrhoeal Disease Research, Bangladesh. None of the E. coli (n = 1789) and Shigella spp. (n = 90) isolated between 2009 and 2010 from stool samples was resistant or had intermediate susceptibility to imipenem. Among 127 extended-spectrum β-lactamase-producing strains isolated during 2003-2009, three Klebsiella pneumoniae isolates (2.4 %) were resistant to imipenem and were positive for bla(NDM-1). All these NDM-1-producing strains were isolated in 2008 and were resistant to all antibiotics tested except for tigecycline and colistin. All three isolates were positive for bla(OXA-1) group, bla(CTX-M-1) group (bla(CTX-M-15)) and bla(SHV) genes, whilst two isolates were positive for 16S rRNA methylase (armA) and qnr (qnrB) genes. One isolate was positive for the bla(CMY) gene and one for the rmtB gene. The bla(NDM-1) gene was located on a conjugative plasmid of ~23-24 MDa. The PFGE patterns of the isolates were different from each other. This study highlights the occurrence of NDM-1-producing organisms in Bangladesh in 2008. The clonal diversity of the isolates and the transferability of bla(NDM-1) plasmids suggest a wider distribution of NDM-1-producing bacteria in Bangladesh.

Diverse carbapenem-resistance mechanisms in 16S rRNA methylase-producing Acinetobacter baumannii

Multidrug-resistant Acinetobacter baumannii has become a global problem. This study characterized amikacin-resistant A. baumannii isolated from eight patients during April 2010-March 2011 in our university hospital and examined the possible mechanisms in three cases in which carbapenem susceptibility changed to carbapenem resistance during treatment of the patients. The armA gene, which is one of the 16S rRNA methylase genes and is associated with high MICs of aminoglycosides, was positive in all isolates. The blaOXA-51 gene and ISAba1 and ISAba1-blaADC were positive in all isolates, but ISAba1-blaOXA-51 was positive in only three isolates. The CarO outer-membrane protein was lost in one isolate. In the first case, both the susceptible and the resistant isolates were positive for ISAba1-blaOXA-51, but the expression of the blaADC gene was increased 3.1-fold in the carbapenem-resistant isolate of the pair. In the second case, the carbapenem-resistant strain became positive for ISAba1-blaOXA-51, resulting in 21.5-fold increased expression of blaOXA-51, compared to the carbapenem-susceptible strain of the pair. In the third case, the carbapenemase genes remained negative despite the carbapenem resistance, but the expression of the adeB gene was increased 4.6-fold after acquisition of carbapenem resistance. Multilocus sequence typing analysis of two isolates showing representative pulsed-field gel electrophoresis patterns demonstrated that both isolates were classified to sequence type 2 (ST2). These results showed that the 16S rRNA methylase-producing A. baumannii, initially susceptible to carbapenem, acquired carbapenem resistance via diverse mechanisms.

Identification of ACT-1 Plasmid-Mediated AmpC β-Lactamase Producing Citrobacter freundii from a Chinese Patient

Identification of ACT-1 Plasmid-Mediated AmpC β-Lactamase Producing Citrobacter freundii from a Chinese Patient

Electron microscopic structures, serum resistance, and plasmid restructuring of New Delhi metallo-β-lactamase-1 (NDM-1)-producing ST42 Klebsiella pneumoniae emerging in Japan

Enterobacteriaceae, carrying the New Delhi metallo-β-lactamase-1 (NDM-1) gene (bla (NDM-1)), have emerged and posed a threat since 2006. In Japan, bla (NDM-1)-carrying Escherichia coli was first described in 2010. In this study, we characterized NDM-1-positive Klebsiella pneumoniae strain 419 in Japan, which was isolated from the urine of a 90-year-old Japanese patient who had never been to the Indian subcontinent. K. pneumoniae 419 belonged to ST42. It possessed a surface capsule (with untypeable capsular PCR types) and was resistant to serum killing. K. pneumoniae 419 cells were occasionally flagellated or piliated and autoaggregated. K. pneumoniae 419 was resistant to β-lactams (including carbapenems), aminoglycosides, and fluoroquinolones, and was susceptible to imipenem (or biapenem), aztreonam, polymixin B, and colistin. It possessed at least eight plasmids; of those, a 74-kb plasmid (pKPJ1) of the replicon FIIA carried bla (NDM-1) and was conjugally transferred to E. coli strains, with a 71-kb transferable azithromycin-resistant (mphA (+)) plasmid of the replicon F (pKPJ2), as a large (145-kb) plasmid (pKPJF100) through a transposition event. In addition to bla (NDM-1), pKPJ1 carried arr-2, pKPJ2 carried mphA, and pKPJF100 carried both. They were negative for the 16S rRNA methylase gene, e.g., which is frequently associated with bla (NDM-1). The data demonstrate that K. pneumoniae 419 possessed virulence- and fitness-associated surface structures, was resistant to serum killing, and possessed a unique (or rare) genetic background in terms of ST type and bla (NDM-1)-carrying plasmid.

Prevalence of plasmid-mediated quinolone resistance determinants in association with β-lactamases, 16S rRNA methylase genes and integrons amongst clinical isolates of Shigella flexneri

Prevalence of plasmid-mediated quinolone resistance determinants in association with β-lactamases, 16S rRNA methylase genes and integrons amongst clinical isolates of Shigella flexneri

Characterization of Clinical Multidrug-ResistantEscherichia coliandKlebsiella pneumoniaeIsolates, 2007–2009, China

Various resistance mechanisms facilitate the emergence and spread of multidrug-resistance (MDR) phenotypes of Escherichia coli and Klebsiella pneumoniae. To elucidate the MDR mechanisms of E. coli and K. pneumoniae in China, we analyzed the antimicrobial susceptibilities of strains isolated from clinical samples in a large tertiary care hospital in Beijing, China, during 2007-2009 and characterized the isolates with a cefotaxime-ciprofloxacin-amikacin (CTX-CIP-AK) resistance pattern. In total, 98 and 52 clinical isolates of E. coli and K. pneumoniae, respectively, with a CTX-CIP-AK resistance pattern were subjected to antimicrobial susceptibility testing and screening of common β-lactamase genes, plasmid-mediated quinolone resistance (PMQR) genes, quinolone resistance-determining region (QRDR) substitutions, and 16S rRNA methylase genes by polymerase chain reaction amplification and DNA sequencing. Pulsed-field gel electrophoresis (PFGE) was used to determine the genetic relatedness of the isolates. Approximately 6.86% and 8.05% of the clinical E. coli and K. pneumoniae isolates, respectively, exhibited MDR phenotypes. The MDR K. pneumoniae isolates exhibited significantly higher ceftazidime resistance than the MDR E. coli isolates (90.4% vs. 76.5%, p=0.0339); a similar result was noted for piperacillin-tazobactam resistance (28.8% vs. 2%, p=0.0001). The common resistance determinants among the MDR E. coli and K. pneumoniae isolates were as follows: CTX-M (88.8% vs. 82.7%), PMQR genes (70.4% vs. 90.4%), gyrA mutations (100% vs. 90.4%), and 16S rRNA methylase genes (93.9% vs. 94.2%). Half (50%) of the MDR E. coli isolates belonged to phylogenetic group D, followed by group A (39.8%). For the E. coli isolates, 94 PFGE patterns and 23 clusters were identified, whereas 51 PFGE patterns and 11 clusters were identified for the K. pneumoniae isolates. Clinical E. coli and K. pneumoniae isolates seem to have a low prevalence of MDR phenotypes in China. The great genetic variation indicates a considerable transmission of common resistance determinants, including a high prevalence of QRDR substitutions in E. coli and K. pneumoniae.

Identification and molecular characterisation of New Delhi metallo-β-lactamase-1 (NDM-1)- and NDM-6-producing Enterobacteriaceae from New Zealand hospitals

The global spread of New Delhi metallo-β-lactamase (NDM) is of significant public health concern. This study sought to determine whether blaNDM was present in Enterobacteriaceae isolates displaying resistance to carbapenems that were submitted to the National Antibiotic Reference Laboratory, Institute of Environmental Science and Research (Porirua, New Zealand) during 2009 and 2010. Isolates were tested for the presence of β-lactamase genes and 16S rRNA methylase genes by polymerase chain reaction (PCR) and sequencing. Plasmid transfer studies were undertaken on isolates found to be harbouring blaNDM. Molecular typing was performed by multilocus sequence typing (MLST). The blaNDM-1 gene was identified in four Enterobacteriaceae isolates (two Escherichia coli, one Klebsiella pneumoniae and one Proteus mirabilis) from four patients in New Zealand hospitals in 2009 and 2010. In addition, the blaNDM-6 gene, which differed from blaNDM-1 by a point mutation at position 698 (C→T), was also identified in an E. coli isolate from the same patient who harboured the blaNDM-1-positive P. mirabilis. All four patients had recently been hospitalised or received health care in India. Four of the isolates also produced a CTX-M-15 extended-spectrum β-lactamase and/or plasmid-mediated AmpC β-lactamase, and all five isolates harboured the plasmid-mediated 16S rRNA methylase rmtC gene. The E. coli types were diverse by MLST, and the K. pneumoniae isolate belonged to the internationally disseminated sequence type 11 (ST11) clone. These findings further illustrate the diversity of phenotypic and genotypic features found in association with blaNDM, in addition to documenting the international spread of this resistance mechanism, notably into a country with historically low rates of antimicrobial resistance.

Complete sequencing of an IncH plasmid carrying the blaNDM-1, blaCTX-M-15 and qnrB1 genes

To characterize plasmid pNDM-MAR recovered from a Klebsiella pneumoniae isolate of sequence type (ST) 15 from Morocco, carrying the genes encoding the metallo-β-lactamase NDM-1, the extended-spectrum β-lactamase (ESBL) CTX-M-15 and the qnrB1 quinolone resistance determinant. The plasmid DNA sequence was obtained by using the 454-Genome Sequencer FLX procedure on a library constructed from total plasmid DNA obtained from an Escherichia coli J53 transconjugant. Contig assembly and predicted gaps were confirmed and filled by PCR-based gap closure. Plasmid pNDM-MAR was 267 242 bp long and encoded 177 predicted proteins. It harboured novel replicons and transfer loci, defining a novel plasmid type within the IncH plasmid family. The bla(NDM-1) gene was flanked by genetic elements that are distinct from those observed in other bla(NDM-1)-positive plasmids, including the groES and groEL chaperonin genes. This plasmid harboured the ESBL gene bla(CTX-M-15) together with the quinolone resistance gene qnrB1. In addition, it harboured genes encoding resistance to tellurite, mercury, chloramphenicol and aminoglycosides. Interestingly, pNDM-MAR did not carry any 16S rRNA methylase gene, in contrast to other bla(NDM-1)-positive plasmids. This study underlines the diversity of genetic vehicles involved in the spread of the bla(NDM-1) gene. Plasmid pNDM-MAR differed significantly from all known bla(NDM-1)-bearing plasmids. Comparative analysis of the pNDM-MAR sequence identified a novel type of IncH plasmid.

Emergence of multidrug-resistant NDM-1-producing Gram-negative bacteria in Bangladesh

The main objective of this study was to investigate the prevalence of bla (NDM-1) in Gram-negative bacteria in Bangladesh. In October 2010 at the International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR,B) laboratories, 1,816 consecutive clinical samples were tested for imipenem-resistant Gram-negative organisms. Imipenem-resistant isolates were tested for the bla (NDM-1) gene. Among 403 isolates, 14 (3.5 %) were positive for bla (NDM-1), and the predominant species were Klebsiella pneumoniae, Acinetobacter baumannii, and Escherichia coli. All bla (NDM-1)-positive isolates were resistant to multiple antibiotics. Among β-lactamase genes, bla (CTX-M-1-group) was detected in ten isolates (eight bla (CTX-M-15)), bla (OXA-1-group) in six, bla (TEM) in nine, bla (SHV) in seven, and bla (VIM) and bla (CMY) in two isolates each. The 16S rRNA methylase gene, armA, was detected in five K. pneumoniae isolates and in one E. coli isolate. rmtB and rmtC were detected in a Citrobacter freundii and two K. pneumoniae isolates, respectively. qnr genes were detected in two K. pneumoniae isolates (one qnrB and one qnrS) and in an E. coli isolate (qnrA). Transferable plasmids (60-100 MDa) carrying bla (NDM-1) were detected in 7 of the 11 plasmid-containing isolates. Pulsed-field gel electrophoresis (PFGE) analysis grouped K. pneumoniae isolates into three clusters, while E. coli isolates differed significantly from each other. This study reports that approximately 3.5 % of Gram-negative clinical isolates in Bangladesh are NDM-1-producing.

Prevalence of 16S rRNA methylase genes in Enterobacteriaceae isolates from a Greek University Hospital

16S ribosomal RNA methylase-mediated high-level resistance to 4-,6-aminoglycosides has been reported in clinical isolates of gram-negative bacilli from several countries. Three of 1534 (0.2%) isolates of Klebsiella pneumoniae and three of 734 (0.4%) Proteus mirabilis isolates from a university hospital in Athens, Greece, were positive for rmtB and highly resistant to all aminoglycosides tested (MICs ≥256 mg/L). Two of the K. pneumoniae rmtB-bearing isolates, were KPC-2 and OXA-10 producers and the third was a DHA-1 producer. One of the P. mirabilis isolates was a VIM-1 and OXA-10 producer and one was an OXA-10 producer. All rmtB-harbouring isolates were clonally unrelated. None of the E. coli (n = 1398) and Enterobacter spp. (n = 414) isolates were positive for armA, rmtA, rmtB, rmtC or rmtD.