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Background: Environmental and clinical carbapenem-resistant Acinetobacter baumannii (CRAb) isolated in a hospital of the Eastern Region of Saudi Arabia were compared to assess the potential environmental contamination by this pathogen. Methods: Frequent-hand-touch surfaces of intensive care (ICU), medical (MW), and surgical (SW) units were randomly sampled for a month-long period, and the CRAb identified were compared to clinical isolates of the same period by PFGE and blaOXA-51-like gene sequencing. Carbapenemase and ribosomal methylase genes, ISAba1 link to blaOXA51-like or to blaOXA-23, respectively were detected by PCR. Results: CRAb was identified from 35.5% of surfaces. All environmental and clinical isolates were multi- or extremely drug resistant. PFGE of all clinical (n = 21) and selected environmental (n = 30) isolates identified a singleton and four clusters, all of which included both clinical and environmental isolates. In the two largest clusters isolates carried blaOXA-66, ISAba1-linked blaOXA-23, and were from the ICU, MW and the male SW. Isolates of the female SW carried blaOXA-69, ISAba1-linked blaOXA-23 and blaGES-11. A pair of clinical and environmental CRAb from the Male SW harboured blaNDM-1 in addition to ISAba1-linked blaOXA-94. Conclusion: A worrying level of environmental contamination, often by CRAb belonging to international clones, was revealed, highlighting the importance of environmental hygiene. Keywords: Acinetobacter baumannii, Multidrug resistance, Environmental contamination, Molecular typing

Data on the prevalence of MCR-producing

To understand the reasons of successful spread of carbapenem-resistant Klebsiella pneumoniae ST14 (CRKP-ST14) in countries of the Arabian Peninsula, the resistome, capsular locus, carbapenemase carrying plasmid types, and core genome of isolates from the region were compared to global isolates. Thirty-nine CRKP-ST14 strains isolated from 13 hospitals in the United Arab Emirates, Bahrain, and Saudi Arabia were selected for whole genome sequencing on Illumina MiSeq platform based on the variety of carbapenemase genes carried and plasmids bearing these genes. Their resistome, capsular locus, and core genome MLST were compared to 173 CRKP-ST14 genomes available in public databases. The selected 39 CRKP-ST14 produced either NDM-1, OXA-48, OXA-162, OXA-232, KPC-2, or co-produced NDM-1 and an OXA-48-like carbapenemase. cgMLST revealed three clusters: 16 isolates from five UAE cities (C1), 11 isolates from three UAE cities and Bahrain (C2), and 5 isolates from Saudi Arabia (C3), respectively, and seven singletons. Resistance gene profile, carbapenemase genes, and their plasmid types were variable in both C1 and C2 clusters. The majority of CRKP-ST14 had KL2, but members of the C2 cluster and two further singletons possessed KL64 capsular locus. Based on cgMLST comparison of regional and global isolates, CRKP-ST14 with KL64 from four continents formed a distinct cluster, suggesting a recent emergence and spread of this variant. Our findings confirmed clonal transmission coupled with likely horizontal gene transfer in carbapenem-resistant Klebsiella pneumoniae ST14. Dissemination of this genetically flexible, highly resistant clone warrants further monitoring.

Plasmid-borne colistin resistance is considered one of the most complex public health concerns worldwide. Several studies reported the presence of the mcr-1.1 harboring Salmonella from the foodstuffs worldwide; still, there is a knowledge gap about the occurrence of these isolates in the Middle East. In this study, we report an mcr-1.1-mediated colistin resistance in two multidrug-resistant (MDR) S. Minnesota (denoted as Sal_2 and Sal_10), with both being also extended-spectrum β-lactamase (ESBL) producing. These isolates have been recovered from two independent samples out of 315 chilled chicken meat tested from retail supermarkets in the United Arab Emirates (UAE). Based on whole-genome sequencing (WGS) analysis, both isolates belonged to the same Sequence Type (ST) ST548. They shared the same genes encoding resistance to the following antimicrobials: polymyxin (mcr-1.1), phenicol (floR), quinolone (qnrB19), aminoglycoside (aac(6')-Iaa), tetracycline (tet(A)), and sulfonamide (sul2). However, the isolates featured different patterns of β-lactamase resistance genes, which included bla

Purpose Plasmids of the incompatibility group X type 3 (IncX3) were described carrying various carbapenemase genes in carbapenemase-producing Enterobacteriaceae (CPE) worldwide and in the United Arab Emirates (UAE), as well. To understand the driving force behind the emergence of such plasmids in the UAE, the relationship between IncX3 plasmids encountered locally and globally was investigated. Methods CPE strains isolated in the UAE during 2009-2014 were screened by X3 PCR-based replicon typing. The clonal relationship of CPE carrying IncX3 plasmids was determined by multi-locus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). Complete sequence of selected IncX3 plasmids was determined. Phylogenetic relationship between the carbapenemase carrying IncX3 plasmids from the UAE and of those reported worldwide was established by comparing the plasmid backbones. Results 10.2% of the 295 CPE tested were identified to carry IncX3 plasmids: 13 Escherichia coli, 13 Klebsiella pneumoniae, two Enterobacter cloacae, one Citrobacter freundii and one Morganella morganii isolate, respectively. Most of them were non-clonal; with small clusters of triplets and pairs of E. coli and K. pneumoniae, and a cluster of five K. pneumoniae ST11 exhibiting >90% similar PFGE patterns, respectively. The 30 isolates harbored either bla NDM-1, bla NDM-4, bla NDM-5, bla NDM-7, bla OXA-181 or bla KPC-2 carbapenemase genes on IncX3 plasmids. Phylogenetic analysis of the backbone region of IncX3 plasmids carrying various beta-lactamase genes from the UAE (n=23) and that of North-America, Europe, Asia and Australia (n=35) revealed three clusters based on the carbapenemase genes carried: plasmids harboring bla OXA-181 and bla NDM-5 formed two distinct groups, whereas backbones of plasmids with bla NDM-1, bla NDM-4 and bla NDM-7 clustered together. Each cluster contained plasmids of diverse geographical origin. Conclusion The findings suggest that different carbapenemase gene carrying IncX3 plasmids encountered in the UAE do not evolve locally, rather are subtypes of this epidemic plasmid emerging in this country due to international transfer.

Ágnes Sonnevend,1 Nour Yahfoufi,1,2 Akela Ghazawi,1 Wafaa Jamal,3 Vincent Rotimi,3 Tibor Pál1 1Department of Medical Microbiology and Immunology, College of Medicine and Health Sciences, United Arab Emirates University, Al Ain, UAE; 2Department of Cellular and Molecular Medicine, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada; 3Department of Microbiology, Faculty of Medicine, Kuwait University, Kuwait City, Kuwait Abstract: Carbapenem-resistant Enterobacteriaceae encountered in countries of the Arabian Peninsula usually produce OXA-48-like and New Delhi metallo-beta-lactamases (NDM) carbapenemases. However, a temporary increase in VIM-4-producing, clonally unrelated Enterobacteriaceae strains was described earlier in a Kuwaiti hospital. We investigated the genetic support of blaVIM-4 in six Klebsiella pneumoniae strains, one Escherichia coli, and one Enterobacter cloacae strain and compared it to that of VIM-4-producing isolates from other countries of the region. Five K. pneumoniae strains and the E. coli strain from Kuwait carried an ~165kb IncA/C-type plasmid indistinguishable by restriction fragment length polymorphism. The complete sequence of one of them (pKKp4-VIM) was established. pKKp4-VIM exhibited extensive similarities to episomes pKP-Gr642 carrying blaVIM-19 encountered in Greece and to the partially sequenced pCC416 harboring blaVIM-4 detected in Italy. In other countries of the region, the only similar plasmid was the one detected in the isolate from the UAE. In all Kuwaiti strains, irrespective of the species and their VIM plasmids, the blaVIM-4 gene was located within the same integron structure (In416), different from those of other countries of the region. Our data show that the spread of this IncA/C plasmid and particularly that of the In416 integron caused a considerable, albeit temporary, increase in the rate of mostly clonally unrelated VIM-producing Enterobacteriaceae strains of multiple species. Monitoring of such events is of high importance as the interference with the spread of mobile genetic elements may represent a formidable challenge to infection control. Keywords: Enterobacteriaceae, VIM carbapenemase, horizontal gene transfer, multidrug resistance, Middle East

Objectives Searching for the presence of the mcr-1 gene in colistin resistant Enterobacteriaceae in countries of the Arabian Peninsula. Methods Seventy-five independent, colistin resistant Enterobacteriaceae strains isolated from clinical cases in Bahrain, Kuwait, Oman, Saudi Arabia and the United Arab Emirates were tested by PCR for the mcr-1 gene. mcr-1 positive strains were genotyped, and their antibiotic susceptibility was established. The mcr-1 containing plasmids were mobilized into Escherichia coli K-12 and their sequence was determined. Results Four E. coli isolates (two from Bahrain, one from Saudi Arabia and one from the United Arab Emirates) were identified carrying the mcr-1 gene on conjugative plasmids. They belonged to global multidrug resistant E. coli clones, i.e. ST648, ST224, ST68 and ST131, respectively. One strain carried the bla NDM-1 carbapenemase gene. Three strains carried mcr-1 on IncI2 type plasmids, one of them also harboring a bla CTX-M-64 gene. In the fourth strain mcr-1 was located on a 240kb IncHI2 plasmid co-harboring 13 other resistance genes. Conclusions This is the first report on the presence of the plasmid-coded mcr-1 gene in a variety of multi-resistant clinical isolates from the Arabian Peninsula indicating that several commonly used antibiotics can potentially facilitate the spread of mcr-1 carrying strains, or directly, mcr-1 containing plasmids.

Plasmid-encoded colistin resistance is emerging among extraintestinal pathogenic Escherichia coli strains, including those of the epidemic clone sequence type 131 (ST131)-H30. Mcr-1 transfers a phosphoethanolamine to the lipid A portion of lipopolysaccharide (LPS), conferring resistance to polymyxins. We investigated whether this modification changed the activity of the monoclonal antibody ASN-4, specific to the O25b side chain of ST131 LPS. We confirmed that, unlike colistin, ASN-4 retained its bactericidal and endotoxin-neutralizing activities and therefore offers a treatment option against extremely drug-resistant ST131 isolates.

Enterobacteriaceae co-producing NDM- and OXA-48-type carbapenemases were encountered in higher frequency in the United Arab Emirates (UAE) than in the neighboring countries in our earlier study. The aim of this investigation was to characterize the seven double carbapenemase producer Klebsiella pneumoniae found in the region to assess factors contributing to their emergence. Three K. pneumoniae ST14 isolated in the UAE harboring blaNDM-1 on IncHI1b and blaOXA-232 on IncColE plasmids were clonally related. Furthermore, two K. pneumoniae from the UAE, ABC106 and ABC137 belonged to ST307 and ST1318, respectively. ABC106 carried blaNDM-1 on IncHI1b, and blaOXA-162 on IncL/M plasmids, whereas ABC137 possessed blaNDM-1 on IncX3 and blaOXA-48 on IncL/M plasmids. The double carbapenemase-producing K. pneumoniae from Oman (OMABC109) and Saudi Arabia (SA54) belonged to ST11 and ST152, respectively. OMABC109 harbored blaNDM-1 on an IncHI1b plasmid highly similar to the NDM-plasmid of ABC106 and carried a chromosomally coded blaOXA-181 located on Tn2013. SA54 possessed a blaNDM-1 on an IncFIb/FII plasmid and a blaOXA-48 on an IncL/M plasmid. Based on these findings, clonal spread and horizontal transfer of carbapenemase genes located on transposons or self-transmissible plasmids contributed equally to the emergence of double carbapenemase-producing Enterobacteriaceae in the region.

Few studies have addressed the molecular epidemiology of carbapenem-resistant Enterobacteriaceae (CRE) isolates in the Arabian Peninsula, and such investigations have been missing from Dubai, a major economical, tourism and medical centre of the region. The antibiotic susceptibility, the carbapenemase type produced, and the clonality of 89 CRE strains isolated in five major Dubai hospitals in June 2015 to June 2016 were determined. Thirty-three percent of the collection of 70 Klebsiella pneumoniae, 13 Escherichia coli and 6 other Enterobacteriaceae were extremely drug resistant, 27% were resistant to colistin, and 4.5% (4 K. pneumoniae isolates) were resistant to all antibiotics tested. The colistin resistance rate in K. pneumoniae was 31.4%. None of the isolates carried mobile colistin resistance genes. Seventy-seven isolates produced carbapenemase: 53.3% OXA-48-like, 24.7% NDM and 22.1% both OXA-48-like and NDM, respectively. Pulsed-field gel electrophoresis clustered 50% of K. pneumoniae into a 35-membered group, which showed significant association with double carbapenemase production, with extreme drug resistance, and with being isolated from Emirati patients. Members of the cluster belonged to sequence type ST14. The rate of colistin resistance in K. pneumoniae ST14 was 37.1% vs. 27.1% of K. pneumoniae isolates outside of the cluster. Two of the panresistant K. pneumoniae isolates also belonged to ST14, whereas the other two were ST15 and ST231, respectively. In conclusion, beyond the overall high colistin resistance rate in CRE, the emergence of a highly resistant clone of K. pneumoniae ST14 in all Dubai hospitals investigated is a serious problem requiring immediate attention.

To identify plasmid-mediated colistin resistance in clinical Enterobacteriaceae isolates in Oman, where this resistance mechanism has not been encountered yet.Twenty-two colistin-resistant Enterobacteriaceae clinical isolates collected between July 2014 and June 2016 in a tertiary care hospital in Muscat were screened by PCR for the mcr-1 and mcr-2 genes. The strain identified as mcr-1 positive was genotyped and its antibiotic susceptibility was established. The mcr-1 containing plasmid was mobilized into Escherichia coli K-12 and its sequence was determined.A single E. coli isolate (OM97) carrying mcr-1 gene was identified, while no strains carrying the mcr-2 gene was found. E. coli OM97 was isolated in June 2016 from blood culture of a male patient with multiple comorbidities. It belonged to ST10. Beyond colistin, it was resistant to amoxicillin-clavulanic acid, piperacillin-tazobactam, amikacin, ciprofloxacin, tetracycline, and cotrimoxazole. The mcr-1 gene was located on a conjugative IncI2-type plasmid of 63722 bp size, which did not harbor any further resistance genes. The genetic surrounding of the mcr-1 gene lacked the ISApl1 element.Although colistin resistance caused by the mcr-1 gene is not common in our collection of clinical isolates, the occurrence of the plasmid-mediated colistin resistance in an E. coli ST10 strain is of concern as this clonal group was already shown to spread ESBL genes and quinolone resistance worldwide. It is especially worrisome that as the mcr-1 gene occurred in a non-ESBL, carbapenem-susceptible E. coli strain, current susceptibility testing algorithms may not detect its presence.

The emergence of pan-resistant Klebsiella pneumoniae strains is an increasing concern. In the present study, we describe a cluster of 9 pan-resistant K. pneumoniae sequence type 147 (ST147) isolates encountered in 4 patients over nearly 1 year in 3 hospitals of the United Arab Emirates (UAE). The isolates exhibited highly similar genotypes. All produced chromosomally encoded OXA-181, and the majority also produced the NDM-5 carbapenemase. As with the previously described single isolate from the UAE, MS6671, the mgrB was disrupted by a functional, IS Ecp1 -driven bla OXA-181 insertion causing resistance to carbapenems. The mutation was successfully complemented with an intact mgrB gene, indicating that it was responsible for colistin resistance. bla NDM-5 was located within a resistance island of an approximately 100-kb IncFII plasmid carrying ermB , mph (A), bla TEM-1B , rmtB , bla NDM-5 , sul1 , aadA2 , and dfrA12 resistance genes. Sequencing this plasmid (pABC143-NDM) revealed that its backbone was nearly identical to that of plasmid pMS6671E from which several resistance genes, including bla NDM-5 , had been deleted. More extensive similarities of the backbone and the resistance island were found between pABC143C-NDM and the bla NDM-5 -carrying IncFII plasmids of two K. pneumoniae ST147 isolates from South Korea, one of which was colistin resistant, and both also produced OXA-181. Notably, one of these strains was isolated from a patient transferred from the UAE. Our data show that this pan-resistant clone has an alarming capacity to maintain itself over an extended period of time and is even likely to be transmitted internationally.

Aim: The purpose of this study was to characterize the New Delhi metallo-beta lactamase (NDM)-7-producing Enterobacteriaceae isolated in the Arabian Peninsula. Methods: Enterobacteriaceae identified to carry blaNDM-7 in a collection of 157 NDM-producing isolates from Kuwait, Oman, Saudi Arabia, and the United Arab Emirates (UAE) were investigated for their antibiotic and disinfectant susceptibility, and resistance gene content. The virulence profile, phylogenetic and sequence types of the isolates were also determined. The plasmids carrying the blaNDM-7 were transferred, and their complete nucleotide sequence was determined. Results: Four NDM-7-producing Escherichia coli isolated in Kuwait, Oman, and the UAE, respectively, were identified. They were clonally unrelated, carried a few virulence determinants only, and belonged to clonal complexes CC10 and CC23, or ST448. They were all multi-drug resistant but remained susceptible to fosfomycin, tigecycline, and colistin. In all isolates, blaNDM-7 was located...

The purpose of this study was to identify carbapenem-resistant Klebsiella pneumoniae in a tertiary care hospital in Sharjah Emirate, to identify the plasmids carrying the carbapenemase genes and to reveal clonal relationships among the isolates. Two hundred and two clinically relevant isolates collected between September 2011 and October 2012 at Al-Qassimi hospital, Sharjah, were investigated for meropenem resistance. Strains with decreased susceptibility were further tested with the modified Hodge test, by EDTA and phenylboronic acid synergy and by E-test. The genes of New Delhi metallo-β-lactamase (NDM), IMP, VIM, OXA-48, and KPC beta-lactamases were targeted by polymerase chain reaction and the genes were located on plasmids by Southern blotting. Clusters of the isolates were revealed by macrorestriction analysis. Seven percent of the isolates were originally found to be meropenem resistant, one isolate have lost its resistance during storage. All of the 13 resistant isolates were positive for the NDM-1 gene located on plasmids of 125 to170 kb, while three isolates also carried the blaOXA-48-like genes. Clusters having repeatedly been isolated over the study period were identified. Carbapenem-resistant Klebsiella pneumoniae carrying the blaNDM-1 gene is a fast emerging problem, emphasizing the potential role of the Middle East as a secondary reservoir for these organisms.

Among 28 clinically relevant, carbapenem non-susceptible Enterobacteriaceae isolates collected in 2009–2011 in the United Arab Emirates three Klebsiella pneumoniae, two Escherichia coli, one Enterobacter cloacae and one Citrobacter freundi were identified to produce NDM-1 carbapenemase. Unexpectedly, with the exception of a K. pneumoniae strain, sequence type ST11, originally acquired in India and subsequently spread nosocomially in the UAE, the majority of the strains could not be directly linked to foreign travel. All isolates harboured the bla NDM-1 gene on plasmids of IncA/C, IncHI1b and IncX3 types, or were untypable. IncX3 type plasmids with a mass of 50 kb and with the same or highly similar restriction patterns, with regions flanking the bla NDM-1 gene identical to the IncX3 NDM plasmids described from China were present in three different species, Enterobacter cloacae, Escherichia coli and C. freundii. Our findings strongly support the assumptions that, beyond the Indian subcontinent, the Middle East is an important reservoir of NDM-producing organisms. Furthermore, we also provide evidence that IncX3 plasmids, recently implicated in the spread of bla NDM-1 in China, have been widely distributed and are important vehicles of the inter-species spread of the NDM-1 gene.

OBJECTIVES To investigate the mechanism of ceftazidime resistance in two isogenic Acinetobacter baumannii strains from the United Arab Emirates. METHODS Two A. baumannii strains, NM55 and NM128, were isolated 4 months apart from a 6-year-old patient in the United Arab Emirates. Genotypic characterization was performed by PFGE and the MIC of ceftazidime was determined by the agar dilution method. Detection of bla(OXA) and metallo-β-lactamase genes was performed by multiplex PCR. Analysis of bla(PER-7), ISAba1, bla(ADC) and the ISCR1 element was carried out by standard PCR. Plasmid analysis was achieved by Southern blotting. RESULTS Strain NM55 was resistant to ceftazidime, whereas strain NM128 was susceptible. Both isolates carried the bla(OXA-23) and bla(OXA-64) genes and were identical according to their PFGE patterns. ISAba1 was present upstream of the bla(OXA-23) gene, but absent upstream of bla(ADC-26), in both strains. Strain NM55 possessed a bla(PER-7) gene with the presence of gst, a fragment of the abc transporter and a transposase gene downstream of it. The entire structure was part of an ISCR1 element and was located on an ≈ 200 kb plasmid in strain NM55, while the ceftazidime-susceptible NM128 strain carried an ≈ 180 kb plasmid without the bla(PER-7) gene. CONCLUSIONS Ceftazidime resistance was mediated by a PER-7 β-lactamase encoded in an ISCR1 element located on a plasmid. This represents the first detection of a PER-7 β-lactamase encoded by a plasmid in A. baumannii.

The feline immunodeficiency virus (FIV) is a lentivirus that is related to human immunodeficiency virus (HIV), causing a similar pathology in cats. It is a potential small animal model for AIDS and the FIV-based vectors are also being pursued for human gene therapy. Previous studies have mapped the FIV packaging signal (ψ) to two or more discontinuous regions within the 5' 511 nt of the genomic RNA and structural analyses have determined its secondary structure. The 5' and 3' sequences within ψ region interact through extensive long-range interactions (LRIs), including a conserved heptanucleotide interaction between R/U5 and gag. Other secondary structural elements identified include a conserved 150 nt stem-loop (SL2) and a small palindromic stem-loop within gag open reading frame that might act as a viral dimerization initiation site. We have performed extensive mutational analysis of these sequences and structures and ascertained their importance in FIV packaging using a trans-complementation assay. Disrupting the conserved heptanucleotide LRI to prevent base pairing between R/U5 and gag reduced packaging by 2.8-5.5 fold. Restoration of pairing using an alternative, non-wild type (wt) LRI sequence restored RNA packaging and propagation to wt levels, suggesting that it is the structure of the LRI, rather than its sequence, that is important for FIV packaging. Disrupting the palindrome within gag reduced packaging by 1.5-3-fold, but substitution with a different palindromic sequence did not restore packaging completely, suggesting that the sequence of this region as well as its palindromic nature is important. Mutation of individual regions of SL2 did not have a pronounced effect on FIV packaging, suggesting that either it is the structure of SL2 as a whole that is necessary for optimal packaging, or that there is redundancy within this structure. The mutational analysis presented here has further validated the previously predicted RNA secondary structure of FIV ψ.

The feline immunodeficiency virus (FIV) long terminal repeat (LTR), compared with some primate lentiviral LTRs, is quite a strong basal promoter. However, it seems to be highly species-specific in function and generally not very efficient in cells of non-feline origin. This study systematically explored the function of the FIV LTR in simian Cos cells compared with its activity in feline and human cells. Our studies, using biologically relevant two- and three-plasmid trans complementation assays followed by semi-quantitative reverse transcriptase PCR, show that the FIV LTR is functional in Cos cells. The results of the Cos experiment are different from previously reported literature and suggest that the strain specificity of the FIV LTR is an important determinant of whether the LTR will be functional in a particular cell type.

The constitutive transport element (CTE) of Mason-Pfizer monkey virus (MPMV) is a short cis-acting sequence element critical for virus gene expression. Analogous to the Rev/Rev Responsive Element (RRE) of primate lentiviruses, CTE allows the nucleocytoplasmic transport of unspliced viral mRNAs. In fact, CTE can functionally replace Rev/RRE in the genomic context and has been used successfully in the expression of viral and cellular genes from expression vectors as well. However, unlike RRE, CTE accomplishes this by interacting with cellular factors, making CTE function independent of co-expressed trans factors. Thus, CTE has proven to be a valuable tool in the expression of heterologous genes. Our previous studies have shown that close proximity of CTE to the polyadenylation sequences is important for CTE function in the genomic context. However, it is controversial whether CTE needs to be located spatially close to the polyadenylation sequences in the subgenomic context. Since CTE is being frequently used in expression vectors, we investigated the position dependency of CTE in the heterologous, subgenomic background using both genetic and structural analyses. Our results reveal that similar to the genomic situation, close proximity of CTE to the polyadenylation sequences is important for its function in the heterologous subgenomic context.

Screening 155 carbapenem non-susceptible Acinetobacter baumannii strains recovered in Abu Dhabi hospitals identified two metallo-s-lactamase bla NDM gene-carrying isolates. They were isolated 4 months apart from the urine of a cancer patient previously treated in Egypt, Lebanon and in the United Arab Emirates. They were clonally related and carried the bla NDM-2 gene recently identified in A. baumannii in Egypt and Israel. Sequences surrounding the bla NDM-2 gene showed significant similarities with those associated with bla NDM-1 in Enterobacteriaceae and A. baumannii. Repeated isolation of bla NDM-2 -positive A. baumannii in the Middle East raises the possibility of the local emergence and spread of a unique clone.

Sir, Since its first description in 1996, variants of KPC have spread throughout the world mostly in the ST258 clone of Klebsiella pneumoniae within isoforms of the Tn4401 transposon located on various plasmids. While the Middle East is considered as a reservoir of various carbapenemase-producing bacteria, to date the only Enterobacteriaceae carrying the blaKPC gene reported from the Arabian Peninsula has been an Escherichia coli ST131 strain isolated in Kuwait. In a collection of 34 carbapenem-non-susceptible Enterobacteriaceae strains, comprising all clinically relevant, nonrepeat isolates recovered in Rashid Hospital, Dubai, United Arab Emirates (UAE) between March 2012 and February 2014, we identified by PCR: 3 blaNDM and 1 blaOXA-48-like E. coli; and 3 blaNDM, 6 blaOXA-48-like, 17 blaNDM+blaOXA-48-like and 2 blaKPC K. pneumoniae. Since blaKPC-positive K. pneumoniae has not been described from this region to date, we subjected the two blaKPC-positive K. pneumoniae isolates (ABC220 and ABC224) to detailed investigations. K. pneumoniae ABC220 was isolated in October 2012 from a necrotizing wound of a previously healthy, 40-year-old Filipino male, 25 days after he had been admitted for perforated appendix and sepsis. K. pneumoniae ABC224 was recovered in May 2013 from the sputum of an 80-year-old UAE national with multiple chronic illnesses, continuously hospitalized for .1 year in a different ward for left leg amputation and sepsis. Prior to the isolation of these strains, various samples from both patients had been repeatedly negative for carbapenem-resistant Enterobacteriaceae. As detected by Etest (bioMerieux, Marcy-l’Etoile, France) or microdilution, ABC220 and ABC224 exhibited susceptibility to tigecycline (MIC1⁄40.75 mg/L) and amikacin (MIC1⁄412 mg/L) only. The MICs of all antimicrobials tested are listed in Table S1 (available as Supplementary data at JAC Online). The XbaI-digested macrorestriction patterns of strains exhibited .95% similarity. Both ABC220 and ABC224 belonged to ST14 by MLST. Strains were subjected to PCR assays specific for ESBL (blaCTX-M, blaSHV, blaTEM and blaOXA) and AmpC b-lactamase genes, plasmidmediated quinolone and aminoglycoside resistance genes and virulence genes (rmpA, wabG, wcaG, fimH, mrkD, iutA, fyuA, iroN, ireA, kfuBC, traTand clpK). Sequencing of the amplicons revealed that they both possessed blaTEM-1, blaSHV-1, blaCTX-M-15 and aac-6′-1b-cr genes and also virulence-related genes fyuA, kfuBC, fimH, mrkD, traT, clpK, ugeA and wabG. Both strains carried sequences suggestive of a K2 capsule type. Outer membrane protein analysis by SDS–PAGE revealed the lack of a band corresponding to OmpK35 from both isolates as compared with the patterns of control strains CSUB10S (Omp36K) and CSUB10S/pSH16K (OmpK35 and OmpK36). Plasmid transfer and analysis by electrophoresis, Southern blotting, hybridization, replicon typing and RFLP was performed as described previously. Although blaKPC-carrying plasmids from both strains were conjugally transferred into E. coli J53RAZ, they were not stably maintained in this recipient. Therefore, plasmids were transferred into E. coli GM2163 by transformation. Both ABC220 and ABC224 harboured a 45 kb IncX3 plasmid carrying blaKPC as identified by hybridization. The plasmids exhibited the same RFLP profile when digested with EcoRI and HindIII (data not shown). Sequencing of blaKPC and its flanking regions by PCR mapping and direct sequencing identified the blaKPC-2 allele in both strains located within the Tn4401b transposon inserted into the umuD gene of the IncX3 plasmid (Figure 1) (GenBank accession no. KM983022). At the time of submission of our manuscript, no KPC-producing K. pneumoniae had been reported from the Arabian Peninsula; although, not without causing considerable confusion, the terminology ‘KPC’ was used earlier referring to a strain exhibiting putative carbapenemase activity by modified Hodge test, without any analysis of the bla gene. Here, we describe two K. pneumoniae ST14 strains, identical in all features tested, carrying blaKPC-2 on IncX3 plasmids. blaKPC-2-bearing IncX3 plasmids described earlier in ST258 K. pneumoniae in France and Hong Kong originated from Greece and the USA, respectively, i.e. from KPC-endemic regions. Compared with these strains, the genetic context of blaKPC-2 is different in the UAE isolates: they map within the Tn4401b isoform inserted in the opposite direction into the genetic load region of the IncX3 plasmid disrupting the umuD gene (Figure 1). It is noteworthy that neither of our patients had a history of travel or recent hospitalization prior to their current admission, suggesting that both of them acquired the strains while being treated in different wards. Based on the data available, we cannot exclude or confirm the possibility that the clone was concealed in the hospital environment (including personnel) during the considerable time that elapsed between the isolation of the two strains. At present, it is not possible to predict whether these cases indicate the beginning of a local spread of KPC-positive K. pneumoniae or whether their incidence will remain low. To answer these questions, which may have implications in understanding the global

We compared the antibiotic susceptibility, clonal lineages and resistance genes of singleton Acinetobacter baumannii strains to those of isolates representing repeatedly encountered molecular types in five Abu Dhabi hospitals. One hundred and ten clinically relevant, non-repeat strains were typed by bla OXA-51-like allele sequencing and by PFGE, and selected isolates also by MLST. Resistance was assessed by MIC determinations and by disc diffusion. Genotyping was carried out by PCR, targeting 28 genes. The 80 epidemic strains belonged to worldwide lineages 1, 2 and 7, representing 11 pulsotypes and 9 genotypes, while the 30 sporadic isolates exhibited a high level of genetic variability and, with the exception of a small subgroup, were not associated with any recognized epidemic lineages. All epidemic subtypes carried the ISAba1-linked bla OXA-23 gene, and harboured the int, the bla PER and the armA genes significantly more frequently than their sporadic counterparts. They were all multi-drug resistant, including non-susceptibility to carbepenems, and were often extensively drug resistant, a phenomenon rarely seen among sporadic strains. Epidemic strains represented 78.8 % of intensive care unit isolates, causing more respiratory infections, while sporadic strains were more frequently isolated from wound and soft tissue infections. The study showed that among strains collected at the same time and from the same region, the very heterogeneous, sensitive sporadic strains, with the exception of a few non-susceptible singleton isolates, clearly differed from the highly resistant epidemic ones, which belonged to multiple pulsotypes and genotypes clustered into three worldwide clonal lineages carrying bla OXA-64, bla OXA-66 and bla OXA-69, respectively.

Screening 34 carbapenem non-susceptible Enterobacteriaceae recovered in Abu Dhabi hospitals identified an Enterobacter cloacae strain carrying bla(VIM-4) , bla(CMY-4) and bla(CTX-M-15) . It was isolated from the urine of an Egyptian patient repeatedly hospitalized and treated with broad-spectrum antibiotics, including carbapenems, in the United Arab Emirates. The bla(VIM-4) coding class I integron, highly similar to In416, was carried on a 175-kilobase non-conjugative incA/C type plasmid also hybridizing with the bla(CMY-4) probe. This is the first detailed report on the isolation of a Verona integron-encoded metallo-β-lactamase (VIM) -producing enteric bacterium in the Arabian Peninsula with characteristics suggestive of spreading from the Mediterranean region.

Aims Meticillin-resistant Staphylococcus aureus (MRSA) strains isolated in Tawam Hospital, a tertiary care hospital in the United Arab Emirates, were examined in order to understand the reasons for a doubling of its incidence between 2003 and 2008 while maintaining the same infection control measures. Methods All consecutive non-duplicate clinically relevant MRSA isolates recovered between January and December 2003 and between May and October 2008 were studied. The antibiotic susceptibility, pulsed field gel electrophoresis, toxin gene, staphylococcal cassette chromosome mec (SCC mec ), spa , agr and multilocus sequence types of the strains were tested. Results In 2003, typical healthcare-associated (HA-MRSA) genotypes (ST239-MRSA-III, ST22-MRSA-IV and ST5-MRSA-II) represented the majority (61.5%) of the isolates. By 2008 this pattern had changed and clonal types considered as community-associated (CA) MRSA comprised 73.1% of the strains with ST80-MRSA-IV, ST5-MRSA-IV and ST1-MRSA with non-typable SCCmec types being the most frequent. However, further epidemiological investigations showed that only one-third of the CA-MRSA infections were actually acquired in the community, indicating that CA-MRSA clones have entered and spread within the hospital. Conclusions The emergence of CA-MRSA clones with subsequent entry to and spread within the hospital has contributed to the increasing incidence of MRSA observed in Tawam Hospital and probably also in other hospitals in the UAE.

Background The mouse mammary tumor virus (MMTV) is unique from other retroviruses in having multiple viral promoters, which can be regulated by hormones in a tissue specific manner. This unique property has lead to increased interest in studying MMTV replication with the hope of developing MMTV based vectors for human gene therapy. However, it has recently been reported that related as well as unrelated retroviruses can cross-package each other's genome raising safety concerns towards the use of candidate retroviral vectors for human gene therapy. Therefore, using a trans complementation assay, we looked at the ability of MMTV RNA to be cross-packaged and propagated by an unrelated primate Mason-Pfizer monkey virus (MPMV) that has intracellular assembly process similar to that of MMTV. Results Our results revealed that MMTV and MPMV RNAs could be cross-packaged by the heterologous virus particles reciprocally suggesting that pseudotyping between two genetically distinct retroviruses can take place at the RNA level. However, the cross-packaged RNAs could not be propagated further indicating a block at post-packaging events in the retroviral life cycle. To further confirm that the specificity of cross-packaging was conferred by the packaging sequences (ψ), we cloned the packaging sequences of these viruses on expression plasmids that generated non-viral RNAs. Test of these non-viral RNAs confirmed that the reciprocal cross-packaging was primarily due to the recognition of ψ by the heterologous virus proteins. Conclusion The results presented in this study strongly argue that MPMV and MMTV are promiscuous in their ability to cross-package each other's genome suggesting potential RNA-protein interactions among divergent retroviral RNAs proposing that these interactions are more complicated than originally thought. Furthermore, these observations raise the possibility that MMTV and MPMV genomes could also co-package providing substrates for exchanging genetic information.

Feline immunodeficiency virus (FIV) is a lentivirus that infects cats and is related to human immunodeficiency virus (HIV). Although it is a common worldwide infection, and has potential uses as a human gene therapy vector and as a nonprimate model for HIV infection, little detail is known of the viral life cycle. Previous experiments have shown that its packaging signal includes two or more regions within the first 511 nucleotides of the genomic RNA. We have undertaken a secondary structural analysis of this RNA by minimal free-energy structural prediction, biochemical mapping, and phylogenetic analysis, and show that it contains five conserved stem–loops and a conserved long-range interaction between heptanucleotide sequences 5′-CCCUGUC-3′ in R/U5 and 5′-GACAGGG-3′ in gag. This long-range interaction is similar to that seen in primate lentiviruses where it is thought to be functionally important. Along with strains that infect domestic cats, this heptanucleotide interaction can also occur in species-specific FIV strains that infect pumas, lions, and Pallas' cats where the heptanucleotide sequences involved vary. We have analyzed spliced and genomic FIV RNAs and see little structural change or sequence conservation within single-stranded regions of the 5′ UTR that are important for viral packaging, suggesting that FIV may employ a cotranslational packaging mechanism.

To establish the role of local transmission versus possible pathogen import due to previous foreign exposure in infections caused by carbapenem non-susceptible Enterobacteriaceae in the Arabian Peninsula, 200 independent isolates collected in 16 hospitals of Saudi Arabia, Kuwait, Oman and the United Arab Emirates were studied. All strains were multidrug resistant; 42.5% of them also qualified as extremely drug resistant. The frequency of various carbapenemases varied according to the participating countries, but in the collection, as a whole, bla NDM-1 was the most frequently encountered carbapenemase gene (46.5%) followed by bla OXA-48-like gene (32.5%). A comparatively high rate (8.9%) of multi-clonal strains carrying both bla NDM and bla OXA-48-like genes in the United Arab Emirates, representing the most resistant subgroup, was encountered. No KPC-expressing isolates were detected. Three major clones of bla NDM-1 carrying Klebsiella pneumoniae of ST152 (n = 22, Saudi Arabia), ST14 (n = 7, United Arab Emirates) and ST147 types (n = 9, Oman) were identified, the latter two clones carrying similar, but not identical HI1b incompatibility type plasmids of >170kb. While from 78.6% of the cases with documented foreign hospitalization bla NDM positive strains were isolated, these strains formed only 25.6% of all the isolates expressing this enzyme. In fact, 56.8% of the NDM, 75.7% of OXA-48-like and 90.9% of VIM positive strains were recovered from patients without documented foreign exposure, neither in the form of travel or prior hospitalization abroad, suggesting a high rate of autochthonous infections. This, considering the extensive links of these countries to the rest of the world, predicts that trends in the local epidemiology of carbapenem resistant strains may increasingly affect the spread of these pathogens on the global scale. These results call for improved surveillance of carbapenem resistant Enterobacteriaceae in the countries of the Arabian Peninsula.