Anne Loughrey

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

BACKGROUND To 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. METHODS Between 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. RESULTS Of 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. CONCLUSIONS The 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.

Antibacterial activity of honey against community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA)

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has now been described globally, as a clinically significant pathogen, particularly associated with skin and soft tissue infections, including abscesses, cellulitis and furunculosis. The recent emergence of CA-MRSA combined with its predominant presentation associated with skin and soft tissue infection, the previous literature indicating honey as an effective treatment of healthcare-associated HA-MRSA-related wound infection, as well as honeys ease of topical application, make the current study timely and of interest to healthcare practitioners involved with wound management. Although previous studies have examined the antimicrobial activity of honey against HA-MRSA, such data are limited regarding the activity of honey against this emerging type of MRSA. CA-MRSA (n=6 isolates), was examined for its susceptibility to natural honey (n=3 honey produced from bees in Northern Ireland and one commercial French honey). Results demonstrated that all honey was able to reduce the cultural count of all CA-MRSA from approximately 10(6) colony-forming units (cfus) (mean = 6.46 log10 cfu/g) to none detectable within 24h of co-culture of separate CA-MRSA organisms individually with all four-honey types examined. Subsequent non-selective enrichment of honey demonstrated that inoculated honey remained positive for CA-MRSA until 72h postinoculation, after which point no culturable organisms could be detected. This study demonstrated that, in vitro, these natural products had an antimicrobial activity against the CA-MRSA organisms tested. Further studies are now required to demonstrate if this antimicrobial activity has any clinical application.

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

OBJECTIVES Between 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. METHODS MICs 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. RESULTS Fifty-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. CONCLUSIONS Gut 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.

Prevalence of Gastrointestinal Bacterial Pathogens in a Population of Zoo Animals

Faecal prevalence of gastrointestinal bacterial pathogens, including Campylobacter, Escherichia coli O157:H7, Salmonella, Shigella, Yersinia, as well as Arcobacter, were examined in 317 faecal specimens from 44 animal species in Belfast Zoological Gardens, during July–September 2006. Thermophilic campylobacters including Campylobacter jejuni, Campylobacter coli and Campylobacter lari, were the most frequently isolated pathogens, where members of this genus were isolated from 11 animal species (11 of 44; 25%). Yersinia spp. were isolated from seven animal species (seven of 44; 15.9%) and included, Yersinia enterocolitica (five of seven isolates; 71.4%) and one isolate each of Yersinia frederiksenii and Yersinia kristensenii. Only one isolate of Salmonella was obtained throughout the entire study, which was an isolate of Salmonella dublin (O 1,9,12: H g, p), originating from tiger faeces after enrichment. None of the animal species found in public contact areas of the zoo were positive for any gastrointestinal bacterial pathogens. Also, water from the lake in the centre of the grounds, was examined for the same bacterial pathogens and was found to contain C. jejuni. This study is the first report on the isolation of a number of important bacterial pathogens from a variety of novel host species, C. jejuni from the red kangaroo (Macropus rufus), C. lari from a maned wolf (Chrysocyon brachyurus), Y. kristensenii from a vicugna (Vicugna vicugna) and Y. enterocolitica from a maned wolf and red panda (Ailurus fulgens). In conclusion, this study demonstrated that the faeces of animals in public contact areas of the zoo were not positive for the bacterial gastrointestinal pathogens examined. This is reassuring for the public health of visitors, particularly children, who enjoy this educational and recreational resource.

Molecular identification of airborne bacteria associated with aerial spraying of bovine slurry waste employing 16S rRNA gene PCR and gene sequencing techniques.

Polymerase chain reaction amplification of the universal 16S ribosomal RNA (rRNA) gene was performed on a collection of 38 bacterial isolates, originating from air sampled immediately adjacent to the agricultural spreading of bovine slurry. A total of 16 bacterial genera were identified including both Gram-positive and Gram-negative genera. Gram-positive organisms accounted for 34/38 (89.5%) of total bacterial numbers consisting of 12 genera and included Staphylococcus (most common genus isolated), Arthrobacter (2nd most common genus isolated), Brachybacterium, Exiguobacterium, Lactococcus, Microbacterium and Sporosarcina (next most common genera isolated) and finally, Bacillus, Brevibacterium, Frigoribacterium, Mycoplana and Pseudoclavibacter. Gram-negative organisms accounted for only 4/38 (10.5%) bacterial isolates and included the following genera, Brevundimonas, Lysobacter, Psychrobacter and Rhizobium. No gastrointestinal pathogens were detected. Although this study demonstrated a high diversity of the microorganisms present, only a few have been shown to be opportunistically pathogenic to humans and none of these organisms described have been described previously as having an inhalational route of infection and therefore we do not believe that the species of organisms identified pose a significant health and safety threat for immunocompetant individuals.

Development of a genus-specific PCR assay for the molecular detection, confirmation and identification of Fusobacterium spp.

Abstract A genus-specific polymerase chain reaction (PCR)-based assay is developed for the detection and identification of clinically relevant Fusobacterium species, including F. nucleatum and F. necrophorum. Two 16S ribosomal DNA (rDNA) primers, FUSO1 (forward primer: 5’-GAG AGA GCT TTG CGT CC-3’ [17-mer]) and FUSO 2 (reverse primer: 5’-TGG GCG CTG AGG TTC GAC -3’ [18-mer]) are designed to target conserved regions of the 16S rDNA gene for Fusobacterium spp. Subsequent proof-of-principle studies employing this assay detected Fusobacterium spp. in the faeces of eight (10%) out of 80 patients with suspected gastrointestinal infection. This assay may be used for the genus-specific detection of Fusobacterium spp. from clinical specimens and for subsequent species identification.

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.

Community-associated MRSA SCCmec type IVd in Irish equids

SIR, – Within the past five years, there has been a marked rise in reports of meticillin-resistant Staphylococcus aureus (MRSA) in animals (Leonard and Markey 2007), including horses (O’Mahony and others 2005), with related discussion regarding the impact of this for animal owners and veterinary personnel (Moore and others 2006). To date, the majority of reports have focused on the differentiation between meticillinsensitive and -resistant organisms, with virtually no attention being given to further characterisation of animal MRSA into health care-associated (HA-MRSA) or community-associated (CA-MRSA) types. In human clinical medicine, the latter organisms differ significantly from HA-MRSA. Although all are S aureus, they have distinct epidemiological and microbiological characteristics, which are summarised in Table 1. Notably, CAMRSA are more likely than HA-MRSA to produce Panton-Valentine leukocidin (PVL) toxin, which is a cytotoxin that causes leucocyte destruction and tissue necrosis, and have appeared as mainly virulent organisms, associated with skin lesions, but have also been reported in fatal cases of necrotising pneumonia (Zetola and others 2005). Given the relatively recent emergence of CA-MRSA in people in the British Isles (Loughrey and others 2007, O’Connell and others 2007), as well as the important clinical and epidemiological differences between these two types of MRSA organisms, we felt it was important to further characterise known equine MRSA isolates from culture collections, in order to further characterise these MRSA organisms. Sixteen previously laboratory-confirmed MRSA isolates were obtained from the Irish Equine Centre, Johnstown, Naas, County Kildare. These isolates had been obtained from equids in 2006 (n=14) and 2007 (n=2), by bacteriological culture from pathological specimens, including wound swabs, joint fluids, lung wash fluids, uterine wash fluids, an umbilical abscess swab, a cervical swab, nasal swabs and skin scrapings. In order to further characterise these isolates to determine whether they were HA-MRSA or CA-MRSA, two genetic loci were examined, namely, presence of the PVL toxin gene, in accordance with the method of Lina and others (1999), and characterisation of the SCCmec gene arrangements, in accordance with the method of Zhang and others (2005). Results indicated that all 16 MRSA isolates were PVL-negative and produced a PCR banding pattern similar to SCCmec IVd. Confirmation of the SCCmec IVd subtype was made by direct automated sequencing of the 746 base pair (bp) PCR amplicons and were confirmed as SCCmec IVd by BLAST analysis. A representative SCCmec IVd DNA sequence (746 bp) from the Irish equids has been deposited in GenBank with accession number EF634484. To date, there has been very little description in the literature of CA-MRSA (PVL-negative, SCCmec IVd subtype) from either humans or animals. One recent report describes 10 isolates of this MRSA type accounting for 10·3 per cent of MRSA from humans, in a Japanese study in hospitals, between 1979/85 (Ma and others 2006). Additionally, to the best of our knowledge, this type of MRSA has not yet been described in isolates from humans either from the Republic of Ireland or Northern Ireland, suggesting that humans are not the primary source of this clonal lineage of MRSA for horses. In conclusion, this report describes the presence of PVL toxin-negative MRSA organisms, belonging to the SCCmec IVd subclass in Irish equids, suggesting a community origin for these organisms, as opposed to a health care origin. Unlike human clinical medicine where reports are numerous, there is little evidence to indicate the importance of such CA-MRSA in animals. Therefore, we urge veterinarians and veterinary microbiologists to further characterise all isolated MRSA organisms from animal populations, as described above, in order to help gain an evidence base to help with our understanding of the potential clinical importance of emerging CA-MRSA in animal populations.

No Evidence of Transmission of Bacteria Between Reptiles and a CF Patient – A Case Report of a Young Adult CF Patient and Reptiles

A microbiological study was undertaken to assess the risk of infection to a CF patient from a collection of pet reptiles, particularly atypical mycobacteria. This study helped to verify that the reptiles under the care of the CF patient did not harbour bacterial organisms that would normally be pathogenic to CF patients. However, the chronic carriage of Pseudomonas aeruginosa and other pathogens in the CF patient may constitute a greater risk of infection to the animals being handled. Therefore, we recommend stringent infection control precautions by CF patients and their pets, particularly adherence to hand washing and disinfection, when handling the animals, their litter or when working with their immediate environment, to potentially minimize the spread of bacterial and other pathogens from animal to human and vice versa. Detailed risk assessments therefore need to be undertaken by clinicians and veterinarians to detail working models that protect both animals and patients from pathogens originating from the other.

Improved cultural selectivity of medically significant fungi by suppression of contaminating bacterial flora employing gallium (III) nitrate.

Incorporation of gallium (III) nitrate into unsupplemented Sabouraud Dextrose Agar to a final concentration of 512 mg/l (2 mM) suppressed bacterial growth, of the following genera Escherichia, Enterococcus, Klebsiella, Listeria, Pseudomonas and Staphylococcus, In contrast growth of Burkholderia cenocepacia, and yeast and filamentous fungi was not affected. Supplementation of selective mycological media with gallium (III) may aid in the selectivity of such media, particularly where clinical specimens are heavily contaminated with bacterial co-habitants and where antibiotic resistance in such bacterial flora may render antibiotic supplements ineffective.