Genomic Characterization of MDR Escherichia coli Harboring blaOXA-48 on the IncL/M-type Plasmid Isolated from Blood Stream Infection.

Escherichia coli is responsible for a wide variety of community and hospital acquired extraintestinal infections, and the emergence of ESBL resistant isolates is a major clinical concern. In this study, we characterized the genomic attributes of an OXA-48 and CTX-M-3 producing E. coli EC-IMP153. Whole-genome initial assembly produced 146 contigs with a combined 5,504,170 bp in size and a G+C content of 50.5%. wgSNPs-based phylogenetic comparison with 36 publically available genomes was also performed. Comprehensive genomic analysis showed that EC-IMP153 belonged to sequence type ST-405 and harbored several resistance determinants including the ß-lactam resistance genes blaOXA-48, blaCTX-M-3, blaTEM-1B, blaOXA-1, and blaCMY-70, aminoglycoside fyuA and aac(3)IId, tetracycline tet(A) and tet(R), and fluoroquinolone gyrA, parC, and mfd resistance determinants. Plasmids with the following incompatibility groups were detected in silico and confirmed using PBRT: IncI1-α, IncL, IncW, Col (BS512), and IncF. To our knowledge this is the first in-depth genomic analysis of an OXA-48 producing E. coli ST-405 isolated from a patient in Lebanon and linked to a blood stream infection. Continuous monitoring is necessary to better understand the continued diffusion of such pathogens, especially in view of the population movements triggered by unrest in the Middle East.

Underlying mechanisms of carbapenem resistance in extended-spectrum ß-lactamase-producing Klebsiella pneumoniae and Escherichia coli isolates at a tertiary care centre in Lebanon: role of OXA-48 and NDM-1 carbapenemases.

A recent increase in carbapenem resistance among extended-spectrum ß-lactamase (ESBL)-producing Klebsiella pneumoniae and Escherichia coli isolates at a major tertiary care centre in Lebanon prompted the initiation of this study. Consecutive ESBL-producing isolates were tested for resistance to carbapenems, with initial screening by disk diffusion and Etest using ertapenem. The modified Hodge test was also performed. PCR of ß-lactamase-encoding genes, including bla(NDM-1), bla(KPC), bla(OXA-48), bla(CTX-M), bla(TEM), bla(SHV), bla(CMY-2) and bla(OXA-1), as well as outer membrane porin genes (ompC and ompF) was performed. Sequencing, efflux pump inhibitor tests and random amplified polymorphic DNA (RAPD) analysis were performed. In total, 14 (2.45%) of 572 K. pneumoniae and 24 (1.07%) of 2243 E. coli were ertapenem-non-susceptible [minimum inhibitory concentration (MIC) ≥0.25 µg/mL]. Resistance to other carbapenems was variable. PCR and sequencing analysis revealed that isolates harboured different ß-lactamase genes, including bla(OXA-1), bla(CTX-M-15), bla(TEM-1), bla(CMY-2), bla(OXA-48) and bla(NDM-1). In addition, K. pneumoniae lacked the outer membrane porin-encoding genes, whilst E. coli harboured them with detected mutations. CTX-M-15 was carried on a 90 kb plasmid, whilst OXA-48 was carried on a 70 kb plasmid. Efflux pump inhibition significantly decreased MICs in E. coli. RAPD analysis demonstrated genomic variability. In conclusion, carbapenem resistance in ESBL-producing K. pneumoniae and E. coli is due to the combined effect of ß-lactamases with porin impermeability and/or efflux pump activity observed in these organisms, and in a number of isolates is due to the production of the carbapenemase-encoding genes bla(OXA-48) and the newly emerging bla(NDM-1).

Study of KIR genes in Lebanese patients with tuberculosis.

A total of 103 Lebanese tuberculosis (TB) cases and 38 controls without TB were studied for the killer cell immunoglobulin-like receptors (KIR) genotypic profile using polymerase chain reaction sequence-specific primers. Patients and controls were assigned to the AA, AB or BB genotypes based on their A or B haplotype genetic make-up, and KIR gene frequencies were compared. We found an increase in the KIR A haplotype in TB patients compared to controls, and only KIR 2DL3 was found to be significantly more prevalent among TB patients. This confirms the findings of another unique international study performed in the Mexican population showing a greater repertoire of inhibitory KIR genes among TB patients than controls.

Cutaneous non-tuberculous Mycobacterial infections: a clinical and histopathological study of 17 cases from Lebanon.

BACKGROUND: Only a few studies characterized cutaneous non-tuberculous Mycobacterium (NTM) infections in this region of the world. Objective The aim of this study was to describe the epidemiological, clinical and histological findings of cutaneous NTM infections in Lebanon. PATIENTS/METHODS: Retrospective study of 17 patients (19 histological specimens) diagnosed with cutaneous NTM infections and confirmed by culture-based partial sequencing of the 16S rRNA gene at the American University of Beirut Medical Center between 2005 and 2008. RESULTS: Of 17 cases, 14 were caused by Mycobacterium marinum. All patients were immunocompetent except for one. Clinically, the most common presentation was multiple sporotrichoid lesions over an extremity (8/17). Many patients had peculiar presentations including bruise-like patches, herpetiform lesions, annular ulcerated plaques, symmetrical nodules over the buttocks and locally disseminated lesions with surrounding pale halo. Almost all patients cleared their infection on either minocycline or clarithromycin monotherapies. Histologically, a dermal small vessel proliferation with mixed inflammation (granulation tissue-like changes) was identified in 58% of specimens. The most common type of granulomatous inflammation was the suppurative (47%) followed by the tuberculoid (30%), sarcoidal (11%), and palisading (5%) types. Lichenoid granulomatous dermatitis was noted in 42% of cases. Special staining highlighted mycobacteria in only two specimens. CONCLUSIONS: The incidence of cutaneous NTM infections is high in our area. Many patients had peculiar clinical presentations. Our study is the second to report the common presence of granulation tissue-like changes as a good histological indicator of cutaneous NTM infections. Minocycline and clarithromycin remain the drugs of choice in our area.

First detection of oxacillinase-mediated resistance to carbapenems in Klebsiella pneumoniae from Morocco.

The frequency of carbapenem resistance due to class-D beta-lactamases (i.e. oxacillinases) among the world's Enterobacteriaceae is increasing. Recently, in Morocco, two isolates of carbapenem-resistant Klebsiella pneumoniae were recovered from the same patient, one harbouring plasmid-encoded bla-(OXA-48) and the other the bla-(OXA-1) gene. This represents the first evidence of bla(OXA-48)-mediated carbapenem-resistance in Enterobacteriaceae in Morocco.

Molecular characterization of Staphylococcus aureus in Lebanon.

Staphylococcus aureus is an important human pathogen and is a growing public health concern. In this study, 130 S. aureus, 93 methicillin-resistant S. aureus (MRSA) and 37 methicillin-sensitive S. aureus (MSSA), clinical isolates recovered from Lebanon were typed by protein A gene (spa) sequencing and multi-locus sequence typing (MLST). Forty-eight different spa types were identified and clustered into 30 different groups. MLST revealed 10 sequence types (STs) among the isolates. There were eight major MRSA clones defined as isolates with the same ST and the same SCCmec type. The majority of the PVL-positive isolates (53%) were ST80-MRSA-IVc. Systematic surveillance of both hospital and community isolates in Lebanon together with measures designed to limit the spread are required.

Staphylococcus aureus nasal carriage rate and associated risk factors in individuals in the community.

The increasing prevalence of Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) strains together with their disease impact on hospital patients and individuals in the community has posed a major challenge to healthcare workers. This study examined the prevalence of S. aureus nasal carriage, antimicrobial susceptibility patterns, and possible risk factors in the community. Of 500 studied subjects (aged from 6 to 65 years) in Lebanon, the overall S. aureus nasal carriage rate was 38.4%, the highest (57.1%) being in children aged 6-10 years. Only eight individuals (1.6%) were carriers of MRSA. Risk factors for S. aureus nasal colonization were male gender, young age, contact with healthcare workers, use of needle injections, and having asthma. A significant decrease in colonization rate was associated with nasal wash with water, use of nasal sprays, and the presence of acne. These findings may assist in better understanding of control measures to decrease nasal colonization with S. aureus in Lebanon and elsewhere.

Molecular characterisation of extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella spp. isolates at a tertiary-care centre in Lebanon.

The prevalence of bla CTX-M, bla TEM and bla SHV genes among extended-spectrum beta-lactamase (ESBL)-producing clinical isolates of Escherichia coli (n = 50) and Klebsiella spp. (n = 50) from Lebanon was 96%, 57% and 67%, and 40%, 82% and 84%, respectively. Genotyping revealed that the clonal diversity was unrelated to the presence of bla genes. Sequence analysis of 16 selected isolates identified the bla CTX-M-15, bla TEM-1, bla OXA-1 and six bla SHV genes, as well as the gene encoding the quinolone-modifying enzyme AAC(6')-Ib-cr. The genes encoding CTX-M-15 and AAC(6')-Ib-cr were carried on a 90-kb plasmid of the pC15-1a or pCTX-15 type, which transferred both ESBL production and quinolone resistance from donors to transconjugants.

First detection and sequence analysis of the bla-CTX-M-15 gene in Lebanese isolates of extended-spectrum-beta-lactamase-producing Shigella sonnei.

The emergence in Shigella species of extended-spectrum beta-lactamases (ESBL) that impart resistance to third-generation cephalosporins is a growing concern world-wide. So far, however, ESBL-producing Shigella have only been reported seven times, albeit from seven different countries. In Lebanon, three ESBL-producing clinical isolates of S. sonnei were recovered from 30 cases of shigellosis diagnosed between July 2004 and October 2005. All three were found to be resistant to amoxycillin, cefotaxime, ceftazidime, aztreonam, trimethoprim/sulphamethoxazole, gentamicin, and kanamycin. Each harboured the bla-CTX-M gene, and the results of sequence analysis indicated this to be of the bla-CTX-M-15 type and encoded on a 70-kb plasmid, flanked by an insertion element (ISEcp1). The bla-TEM-1 gene was also detected on the chromosomes of two of the ESBL-producing isolates. Class-2 integrons containing dhfr1, aadA1 and sat1 genes were detected on the chromosomes of all three isolates but not on the plasmids. Fluoroquinolone-modifying factors [QnrA, QnrB, QnrS or AAC(6')-Ib-cr] were not detected. The results of RAPD analysis, combined with data on antimicrobial susceptibility, indicated that each isolate was unique. In conclusion, the emergence of ESBL-producing isolates of S. sonnei has been demonstrated for the first time in Lebanon. The resistance of these isolates to third-generation cephalosporins was mediated by the CTX-M-15 enzyme, which was plasmid-encoded.

Fatal brain infection caused by Aspergillus glaucus in an immunocompetent patient identified by sequencing of the ribosomal 18S-28S internal transcribed spacer.

Cerebral aspergillosis has rarely been reported in immunocompetent patients. We herein describe a unique case of cerebral aspergillosis in a healthy adult that led to his death despite aggressive antifungal therapy. Sequencing of ribosomal 18S-28S internal transcribed spacer identified the organism as Eurotium herbariorum, the teleomorph of Aspergillus glaucus.

Nationwide study of drug resistance among acid-fast bacilli positive pulmonary tuberculosis cases in Lebanon.

OBJECTIVE: To assess the prevalence of drug resistance among smear-positive sputum specimens from pulmonary tuberculosis (TB) cases in Lebanon. DESIGN: Between July 2002 and April 2004, 224 newly diagnosed TB cases and 21 previously treated TB cases were collected nationwide. Mycobacterium tuberculosis isolates were tested against isoniazid (INH), rifampicin (RMP), streptomycin (SM) and ethambutol (EMB) using the BACTEC-TB system. RESULTS: M. tuberculosis and non-tuberculous mycobacteria (NTM) isolates were recovered from 190 and 15 new cases, respectively, and from 16 and 1 previously treated cases, respectively. Overall drug resistance among new TB vs. previously treated TB cases was 19.5% and 75%, and for single drugs it was INH (12% vs. 63%), RMP (3% vs. 56%), SM (12% vs. 44%) and EMB (3% vs. 44%). The overall rate of multidrug resistance (MDR) was 5.8% (1% vs. 62.5%). The male:female ratio was 1.3:1; most were young adults. CONCLUSION: Relatively moderate single drug resistance and very low MDR rates were found among new TB cases, while among previously treated TB cases very high resistance and MDR resistance rates were detected. Such findings underline the need for ongoing stringent control measures to curb the spread of M. tuberculosis and its deleterious effects.

Seroprevalence of toxocariasis in Lebanon: a pilot study.

Toxocariasis is a common helminthic infection that has a worldwide distribution. However, data from Lebanon about the prevalence of this infection are non-existent. We conducted a Toxocara seroprevalence study with 150 subjects attending the outpatient clinics at the American University of Beirut Medical Center between May and June 2004. Serum specimens were tested for anti-Toxocara antibodies by enzyme-linked immunosorbent assay and confirmed by Western blot. Multivariate analysis was performed to identify risk factors for infection. The seroprevalence rate of toxocariasis was 19%. Male gender and below high school education were significantly associated with a positive serological test (odds ratios = 3.1 and 2.8, respectively). Higher numbers of persons in the household, and low family income during childhood, were significant on bivariate analysis only. Toxocariasis is common in Lebanon. A large population-based survey is needed to confirm these results.

Brucella infection in total hip replacement: case report and review of the literature.

A 47-y-old female underwent revision of a left total hip replacement because of loose prosthesis. Routine intraoperative culture of the hip site grew Brucella sp. The patient was treated with a combination of vibramycin and rifampicin for 5 months. At 4-y follow-up, her condition is good. Though prosthetic infection with Brucella spp. is an extremly rare condition (only 1 case each of femur and hip, and 3 cases of knee had been previously reported in the English literature), brucella infection of prosthetic joints should be considered in brucella endemic areas.

Campylobacter jejuni: unusual cause of cholecystitis with lithiasis. Case report and literature review.

A 51-year-old man presented with acute cholecystitis and the routine intraoperative culture of the bile grew Campylobacter jejuni. The patient was cured by laparoscopic cholecystectomy without specific antimicrobial treatment. Cholecystitis owing to Campylobacter spp. could be missed because a culture for Campylobacter is not routinely requested nor is it cost effective to look for it in bile or gallbladder specimens. Moreover, the fastidious nature of these bacteria dictates against their recovery in routine culture. Because this is a rare infection at this site, a review of the literature on this infection is included.

Prevalence of S315T mutation within the katG gene in isoniazid-resistant clinical Mycobacterium tuberculosis isolates from Dubai and Beirut.

OBJECTIVE: To determine the prevalence of S315T mutation within the katG gene that confers clinically significant resistance to isoniazid in isoniazid-resistant Mycobacterium tuberculosis isolates recovered from tuberculosis patients in Dubai and Beirut. METHOD: A total of 28 and 17 isoniazid-resistant and seven and six susceptible clinical M. tuberculosis isolates from Dubai and Beirut, respectively, were tested. The presence of S315T mutation in the katG gene was detected by amplification of the DNA region around codon 315 by polymerase chain reaction followed by restriction digestion with Msp I to generate restriction fragment length polymorphism. The genotyping of the isolates carrying S315T mutation was carried out by touchdown double-repetitive-element PCR (DRE-PCR). RESULTS: The mutation S315T was detected in 18 (64%) of 28 isoniazid-resistant isolates from Dubai and in six (35%) of 17 resistant strains from Beirut. None of the susceptible strains contained this mutation. The genotyping studies showed that the majority of the isolates carrying the S315T mutation exhibited unique DNA banding patterns. CONCLUSION: The data show a varying prevalence of S315T mutation within the katG gene in M. tuberculosis strains isolated from the two geographical locations, Dubai and Beirut.

Ultrashort regimen of lansoprazole-amoxicillin-azithromycin for eradicating Helicobacter pylori.

The efficacy and safety of two ultrashort azithromycin-containing regimens for Helicobacter pylori infection were studied. Patients positive for H. pylori infection were assigned to receive either a three-day drug regimen (group A) or a five-day regimen (group B). In both groups, patients received lansoprazole 30 mg p.o. twice daily on day 1 and, on days 2 and 3, lansoprazole 30 mg p.o. twice daily, amoxicillin 1 g (of anhydrous amoxicillin) p.o. twice daily, and azithromycin 500 mg (of anhydrous azithromycin) p.o. twice daily. Group B patients received lansoprazole 30 mg p.o. twice daily and amoxicillin 1 g p.o. twice daily for two additional days. Gastric biopsy specimens were subjected to culture and susceptibility testing. A minimum of four weeks after the completion of therapy, the patients underwent a 14C-urea breath test to determine whether H. pylori had been eradicated. A total of 28 patients were enrolled (15 in group A and 13 in group B). Treatment was well tolerated. H. pylori was eradicated in 4 (36%) of 11 patients in group A and 2 (22%) of 9 group B patients (26.6% and 15.4%, respectively, in intention-to-treat analysis). None of the isolates of H. pylori showed resistance to amoxicillin or clarithromycin. Regimens consisting of lansoprazole plus two or four days of azithromycin and amoxicillin therapy eradicated H. pylori in 36% and 22% of patients, respectively.