Paul J. Rooney

Nursing homes as a reservoir of extended-spectrum β-lactamase (ESBL)-producing ciprofloxacin-resistant Escherichia coli

BACKGROUNDnTo assess the prevalence and risk factors for faecal carriage of fluoroquinolone-resistant, extended-spectrum beta-lactamase (ESBL)-producing, Escherichia coli (MDR E. coli) among residents in nursing homes in Northern Ireland.nnnMETHODSnBetween January 2004 and May 2006, retrospective histories of hospital admissions, antimicrobial treatment and co-morbidities were collected. Faecal samples were cultured for MDR E. coli. These isolates and their ESBL genes were typed by a reference laboratory.nnnRESULTSnOf the 294 patients included in the study, faecal samples from 119 (40.5%) grew MDR E. coli. The proportion of carriers in the different homes ranged from 0% to 75%. Epidemic strain A belonging to the ST131, O25:H4 lineage with the CTX-M-15 enzyme accounted for 58 (49%) of all isolates; its proportion varied from 0% to 100% among homes. Fifty-one percent of carriers had no history of recent hospital admission and only 13.5% had a known history of ESBL E. coli colonization or infection. In a multivariate logistic regression model, days of fluoroquinolone use [odds ratio (OR) = 1.33, 95% confidence interval (CI) 1.04-1.69, P = 0.02] and a history of urinary tract infection (OR = 2.56, 95% CI 1.37-4.78, P = 0.003) were the only variables independently associated with the risk of carrying MDR E. coli.nnnCONCLUSIONSnThe high level of faecal carriage of MDR E. coli in nursing home residents demonstrates their importance as a reservoir population. Public health measures to combat spread of these organisms should address the needs of this group.

Molecular epidemiology of fluoroquinolone-resistant ST131 Escherichia coli producing CTX-M extended-spectrum β-lactamases in nursing homes in Belfast, UK

OBJECTIVESnBetween January 2004 and May 2006 Escherichia coli producing extended-spectrum β-lactamases (ESBLs) were isolated from the faeces of 118/294 residents from 16 nursing homes in Belfast. Of these, 58 isolates belonged to UK strain A, a variant of the international ST131 clone. Here we investigated the remaining 60 ESBL producers.nnnMETHODSnMICs were determined and interpreted using BSAC methodology. Isolates were characterized by phylogenetic typing, real-time PCR and PFGE. Plasmids were rep typed by PCR and their similarity to IncI1 reference plasmid pEK204 was investigated by restriction fragment length polymorphism analysis. The molecular environments surrounding bla(CTX-M) were determined by DNA sequencing and PCR.nnnRESULTSnFifty-nine of 60 isolates belonged to the B2, ST131 lineage; of these 28 belonged to the previously defined UK strain C, while the other 31 were clustered into five groups by PFGE. Forty-nine isolates harboured bla(CTX-M-3) on plasmids of five different rep types (I1, FIA, FIA-FIB, N and Y) and 11 harboured bla(CTX-M-15) on F-type plasmids (FIA and FIA-FIB). All CTX-M-3 ESBL producers and three with CTX-M-15 ESBL had an intact copy of ISEcp1 immediately upstream of bla(CTX-M); the remaining eight with CTX-M-15 ESBL had a truncated ISEcp1.nnnCONCLUSIONSnGut colonization among nursing home residents in Belfast with ciprofloxacin-resistant E. coli producing ESBLs almost entirely involves clonal spread of ST131 variants, with similar genetic environments for bla(CTX-M-3) or bla(CTX-M-15) as in pEK204 and pEK499. Such diversity indicates dissemination of both plasmids and ESBL genes among a single commonly multiresistant clone.

Distribution of serotypes and antibiotic resistance among pneumococci in Northern Ireland

Altogether, 488 consecutive strains of Streptococcus pneumoniae isolated from clinical specimens were serotyped and their antibiotic susceptibility determined. Of all strains isolated, 89.7% (90.6% for strains isolated from patients with serious infection) were of types present in the new polyvalent (23-valent) pneumococcal vaccine. Four strains showed reduced susceptibility to penicillin (minimum inhibitory concentration 0.1-1.0 mg/l). Two of those strains (both serotype 23) were also of intermediate susceptibility to other antibiotics (ampicillin, cephradine, chloramphenicol and tetracycline) but were sensitive to erythromycin. A significant proportion (12%) was resistant to tetracycline.

Determination of total antibiotic resistance in waterborne bacteria in rivers and streams in Northern Ireland: Can antibiotic-resistant bacteria be an indicator of ecological change?

Antibiotic resistance (ABR) has now become a major public health issue. Relatively little studies have been published on the incidence of ABR in environmental isolates circulating within the community. A study was performed to determine the diversity of total ABR (intrinsicxa0+xa0acquired resistance) in waterborne bacteria. Surface water from 12 waterways, including 11 rivers/steams and 1 lake, were examined for the presence of ABR phenotypes, using a direct antibiotic susceptibility assay and demonstrated the presence of ABR (in increasing order of resistance), to the following 19 agents: amikacin (17%), gentamicin (17%), ciprofloxacin (33%), colistin (42%), linezolid (42%), tobramycin (42%), vancomycin (42%), ertapenem (67%), erythromycin (75%), meropenem (75%), rifampin (75%), teichoplanin (75%), tetracycline (75%), trimethoprim (75%), fusidic acid (83%), aztreonam (92%), clindamycin (92%), penicillin (92%) and cefoxitin (100%). Multiple resistance to the major classes of antibiotics was noted, which varied from one to six classes, with a mean resistance to 3.7 major antibiotics classes, with diminishing antibacterial effectiveness in the following order: aminoglycosidesxa0>xa0fluoroquinolonesxa0>xa0glycopeptidesxa0>xa0macrolidesxa0>xa0tetracyclinesxa0>xa0β-lactams. Overall, these data indicate that waterborne bacteria are an important source of ABR determinants and contribute to the mass balance of ABR in the environment, and may be used as an indicator of ecological change in water ecosystems. The waterborne ABR organisms may potentially act as donors in pathogens, which may acquire these through horizontal gene transfer or other genetic exchange events, thus leading to clinically significant cases in both animal and human health. Therefore, environmental bacteria should not be regarded as being devoid of ABR determinants, simply because they are physically removed from clinical settings. Such bacteria have natural intrinsic resistance, as well as having the ability to acquire determinants from agricultural run-off and human wastewater discharge, which may contain ABR organisms, as well as sublethal concentrations of metabolically active antibiotic. The tracking of such organisms to their source may help determine the source of fecal pollution in aquatic ecosystems.

Identification of airborne bacterial and fungal species in the clinical microbiology laboratory of a university teaching hospital employing ribosomal DNA (rDNA) PCR and gene sequencing techniques

Universal or “broad-range” PCR-based ribosomal DNA (rDNA) was performed on a collection of 58 isolates (n = 30 bacteria + 28 fungi), originating from environmental air from several locations within a busy clinical microbiology laboratory, supporting a university teaching hospital. A total of 10 bacterial genera were identified including both Gram-positive and Gram-negative genera. Gram-positive organisms accounted for 27/30 (90%) of total bacterial species, consisting of seven genera and included (in descending order of frequency) Staphylococcus, Micrococcus, Corynebacterium, Paenibacillus, Arthrobacter, Janibacter and Rothia. Gram-negative organisms were less frequently isolated 3/30 (10%) and comprised three genera, including Moraxella, Psychrobacter and Haloanella. Eight fungal genera were identified among the 28 fungal organisms isolated, including (in descending order of frequency) Cladosporium, Penicillium, Aspergillus, Thanatephorus, Absidia, Eurotium, Paraphaeosphaeria and Tritirachium, with Cladosporium accounting for 10/28 (35.7%) of the total fungal isolates. In conclusion, this study identified the presence of 10 bacterial and eight fungal genera in the air within the laboratory sampled. Although this reflected diversity of the microorganisms present, none of these organisms have been described previously as having an inhalational route of laboratory-acquired infection. Therefore, we believe that the species of organisms identified and the concentration levels of these airborne contaminants determined, do not pose a significant health and safety threat for immunocompotent laboratory personnel and visitors.

Evaluation of the VITEK®2 AST N-054 test card for the detection of extended-spectrum β-lactamase production in Escherichia coli with CTX-M phenotypes

OBJECTIVESnA new VITEK 2 antibiotic susceptibility testing (AST) card, AST N-054, was introduced for aerobic gram-negative bacilli in 2007 and has been widely adopted for routine use in the UK. We evaluated its performance for detecting extended-spectrum beta-lactamase (ESBL) production in Escherichia coli.nnnMETHODSnESBL-producing faecal isolates of E. coli (n = 137) from residents in nursing homes were tested using the AST N-054 card on VITEK 2 and with MASTDISCS ID ESBL detection disc diffusion tests (Mast Diagnostics, Bootle, UK). The susceptibility result recommended by the VITEK 2 software was also recorded.nnnRESULTSnThe AST N-054 card detected ESBL production in 93 of the 137 isolates tested [test sensitivity 67.9% (95% CI, 59.7-75.1)]. E. coli strain A, a widespread lineage in the UK with a low-level CTX-M enzyme production, accounted for most of the detection failures, with 35/73 strain A isolates incorrectly reported versus 9/64 non-strain A isolates (P < 0.0001). The MASTDISCS correctly detected ESBL in 135/137 isolates [test sensitivity 98.5% (95% CI, 94.5-99.9)]. Of the 44 isolates found to be negative for ESBL production by VITEK 2, the Advanced Expert System misreported 29 as susceptible to cefotaxime and all as susceptible to ceftazidime and aztreonam.nnnCONCLUSIONSnThese data suggest that the AST N-054 card for the VITEK 2 system is less reliable than other previously reported cards for the detection of CTX-M beta-lactamase-producing E. coli circulating in the UK, particularly strain A isolates.

Speciation of Bacillus spp. in honey produced in Northern Ireland by employment of 16S rDNA PCR and automated DNA sequencing techniques

Phenotypic speciation of foodborne Bacillus spp. remains problematic in terms of obtaining a reliable identification. In this study, we wished to identify several bacterial isolates from honey produced in Northern Ireland, and which belonged to the genus Bacillus, through employment of a molecular identification scheme based on PCR amplification of universal regions of the 16S rRNA operon in combination with direct automated sequencing of the resulting amplicons. Seven samples of honey and related materials (propolis) were examined microbiologically and were demonstrated to have total viable counts (TVC) ranging from <100 to 1700 colony-forming units/g. No yeasts or filamentous fungi were isolated from the honey materials. Several bacterial isolates were identified using this method, yielding two different genera (Paenibacillus and Bacillus), as well as four Bacillus species, namely Bacillus pumilus, B.xa0licheniformis, B.xa0subtilis and B.xa0fusiformis, with B.xa0pumilus the most frequently identified species present. When the use of molecular identification methods is justified, employment of partial 16S rDNA PCR and sequencing provides a valuable and reliable method of identification of Bacillus spp. from foodstuffs and negates associated problems of conventional laboratory and phenotypic identification.

Rapid discrimination of Enterococcus faecium strains using phenotypic analytical techniques

Clinical isolates of glycopeptide resistant enterococci (GRE) were used to compare three rapid phenotyping and analytical techniques. Fourier transform infrared (FT-IR) spectroscopy, Raman spectroscopy and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) were used to classify 35 isolates of Enterococcus faecium representing 12 distinct pulsed-field gel electrophoresis (PFGE) types. The results show that the three analytical techniques provide clear discrimination among enterococci at both the strain and isolate levels. FT-IR and Raman spectroscopic data produced very similar bacterial discrimination, reflected in the Procrustes distance between the datasets (0.2125–0.2411, p < 0.001); however, FT-IR data provided superior prediction accuracy to Raman data with correct classification rates (CCR) of 89% and 69% at the strain level, respectively. MALDI-TOF-MS produced slightly different classification of these enterococci strains also with high CCR (78%). Classification data from the three analytical techniques were consistent with PFGE data especially in the case of isolates identified as unique by PFGE. This study presents phenotypic techniques as a complementary approach to current methods with a potential for high-throughput point-of-care screening enabling rapid and reproducible classification of clinically relevant enterococci.

Phenotypic diversity of Campylobacter isolates from sporadic cases of acute human gastroenteritis in Northern Ireland.

J. E. MOORE, B. C. MILLAR, M. A. S. McMAHON, D. A. McDOWELL and P. J. ROONEY *Northern Ireland Public Health Laboratory, Department of Bacteriology, Belfast City Hospital, Belfast; School of Biomedical Science, Centre for Molecular Biosciences, University of Ulster, Cromore Road, Coleraine, Co. Londonderry; and Food Microbiology Research Group, University of Ulster, Shore Road, Jordanstown, Newtownabbey, Co. Antrim, Northern Ireland, UK