Akela Ghazawi

Emergence and spread of NDM-1 producer Enterobacteriaceae with contribution of IncX3 plasmids in the United Arab Emirates.

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 blaNDM-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 blaNDM-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 blaNDM-1 in China, have been widely distributed and are important vehicles of the inter-species spread of the NDM-1 gene.

NDM-2 carbapenemase-producing Acinetobacter baumannii in the United Arab Emirates

Screening 155 carbapenem non-susceptible Acinetobacter baumannii strains recovered in Abu Dhabi hospitals identified two metallo-ß-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.

Plasmid-encoded PER-7 β-lactamase responsible for ceftazidime resistance in Acinetobacter baumannii isolated in the United Arab Emirates

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.

Plasmid-mediated colistin resistance in Escherichia coli from the Arabian Peninsula

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 blaNDM-1 carbapenemase gene. Three strains carried mcr-1 on IncI2 type plasmids, one of them also harboring a blaCTX-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.

Change in meticillin-resistant Staphylococcus aureus clones at a tertiary care hospital in the United Arab Emirates over a 5-year period

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 (SCCmec), 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.

Sequences Intervening between the Core Packaging Determinants Are Dispensable for Maintaining the Packaging Potential and Propagation of Feline Immunodeficiency Virus Transfer Vector RNAs

ABSTRACT The packaging determinants of feline immunodeficiency virus (FIV) consist of two discontinuous core regions, extending from R to ∼150 bp of the 5′ untranslated region and the first ∼100 bp of gag. However, the role of sequences intervening between the core regions in packaging has not been clear. A mutational analysis was conducted to determine whether the intervening sequences played a role in FIV RNA packaging, using an in vivo packaging assay complemented with semiquantitative reverse transcriptase PCR. Our analyses reveal that the intervening sequences are dispensable not only for vector RNA packaging but also for propagation, confirming the discontinuous nature of the FIV packaging signal.

Optimal Packaging of FIV Genomic RNA Depends upon a Conserved Long-range Interaction and a Palindromic Sequence within gag

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.

Characterization of Carbapenem-Resistant Enterobacteriaceae with High Rate of Autochthonous Transmission in the Arabian Peninsula

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.

VIM-4 carbapenemase-producing Enterobacter cloacae in the United Arab Emirates

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.

CHARACTERISTICS OF EPIDEMIC AND SPORADIC STRAINS OF ACINETOBACTER BAUMANNII ISOLATED IN ABU DHABI HOSPITALS

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 blaOXA-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 blaOXA-23 gene, and harboured the int, the blaPER 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 blaOXA-64, blaOXA-66 and blaOXA-69, respectively.