Action against antibiotic resistance: no time to lose.
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Anti-Bacterial Agents↗
Biomedical subjects
P M Hawkey
Publications and source records attributed to P M Hawkey.
Exceptional desiccation tolerance of Acinetobacter radioresistens.
The taxonomy of the genus Acinetobacter, which includes several important nosocomial pathogens, has been confused due to a lack of discriminatory phenotypic characteristics for identification. Molecular methods such as amplified ribosomal DNA restriction analysis (ARDRA) now enable the accurate identification of species. Ten clinical isolates of Acinetobacter radioresistens had genospecies confirmed by ARDRA but the APJ 20NE system, commonly used in clinical microbiology laboratories, mis-identified them as Acinetobacter lwoffii. Desiccation resistance of Acinetobacter spp. is an important attribute for their survival in the clinical environment. We investigated the ability of A. radioresistens to survive desiccation using an established glass surface model and compared the results to A. lwoffii and Acinetobacter baumannii. The 10 strains of A. radioresistens were extremely resistant to desiccation and survived for an average of 157 days at 31% relative humidity (RH). In contrast, two strains of A. lwoffii and three strains of A. baumannii survived for an average of three and 20 days respectively, at 31% RH, which was used as an approximation to climatic conditions in UK hospitals. A. radioresistens is thus well adapted for survival in the hospital environment and carriage on human skin and yet it is reported less frequently than A. lwoffii amongst clinical isolates. Cases of A. radioresistens infection may be under-reported due to mis-identification as A. lwoffii and further studies that use molecular identification methods are required to elucidate the role of A. radioresistens in human disease.
Acinetobacter↗
Ventilation grilles as a potential source of methicillin-resistant Staphylococcus aureus causing an outbreak in an orthopaedic ward at a district general hospital.
The spread of methicillin-resistant Staphylococcus aureus (MRSA) in a hospital is thought to be mainly by direct contact. Environmental sources such as exhaust ducting systems have been increasingly recognized as a source for MRSA outbreaks in intensive therapy units. We describe an outbreak of MRSA related to ventilation grilles in an orthopaedic ward. Six patients and one nurse were involved in an outbreak with EMRSA-15 during March 1996. The index case was transferred from a large university hospital in Leeds. One of the patients had shared the same bay with the index case. The rest of the patients were in another bay of the same ward and had no direct contact with the index patient. An environmental source was suspected and the ventilation grilles in boys 1 and 2 were found to be harbouring EMRSA-15. The ventilation system at that time was working on an intermittent cycle from 4 p.m.-8 a.m. Daily shut-down of the system temporarily created a negative pressure, sucking air in from the ward environment into the ventilation system and probably contaminating the outlet grilles. It is likely that contaminated air was blown back into the ward when the ventilation system was started. The system was thoroughly cleaned, appropriate infection control measures were instituted and the ventilation system was put back on a continuous running cycle and the outbreak terminated. Six months after the outbreak no isolates of EMRSA-15 had been made on the ward.
Air Microbiology↗
Prospective study of the risk of Clostridium difficile diarrhoea in elderly patients following treatment with cefotaxime or piperacillin-tazobactam.
BACKGROUND: Rates of Clostridium difficile diarrhoea have recently been rising, with the elderly being at highest risk. AIM: To compare the incidence of C. difficile colonization and diarrhoea in elderly patients treated for presumed infection with either empirical cefotaxime (CTX) or piperacillin-tazobactam (PT). METHODS: A prospective, ward-based, crossover study was carried out on two well-matched care of the elderly wards at a UK tertiary care hospital, in patients requiring empirical broad-spectrum antibiotic treatment. RESULTS: There was a highly significant increased incidence of C. difficile colonization (26/34 vs. 3/14, P=0.001) and diarrhoea (18/34 vs. 1/14, P=0.006) in patients who received CTX as opposed to PT. DNA fingerprinting suggested that most infections arose from strains acquired from the hospital environment. CONCLUSIONS: Elderly patients are significantly less likely to develop C. difficile diarrhoea after treatment with PT than after CTX. The source of C. difficile appears to be predominantly from the ward environment.
Aged↗
PCR single strand conformational polymorphism can be used to detect the gene encoding SHV-7 extended-spectrum beta-lactamase and to identify different SHV genes within the same strain.
The PCR single strand conformational polymorphism (PCR-SSCP) technique described to identify mutants of the SHV beta-lactamases was extended to identify an SHV-7 type beta-lactamase. This was found in a strain of Klebsiella pneumoniae, the first recorded isolate in the UK to produce this type of enzyme. We also demonstrate that PCR-SSCP can be used to identify more than one SHV beta-lactamase gene in a single strain.
Anti-Bacterial Agents↗
Survival of Acinetobacter baumannii on dry surfaces: comparison of outbreak and sporadic isolates.
Acinetobacter spp. are important nosocomial pathogens reported with increasing frequency in outbreaks of cross-infection during the past 2 decades. The majority of such outbreaks are caused by Acinetobacter baumannii. To investigate whether desiccation tolerance may be involved in the ability of certain strains of A. baumannii to cause hospital outbreaks, a blind study was carried out with 39 epidemiologically well-characterized clinical isolates of A. baumannii for which survival times were determined under simulated hospital conditions. The survival times on glass coverslips of 22 strains isolated from eight well-defined hospital outbreaks in a German metropolitan area were compared with the survival times of 17 sporadic strains not involved in outbreaks but rather isolated from inpatients in the same geographic area. All sporadic isolates have been shown by pulsed-field gel electrophoresis to represent different strain types. There was no statistically significant difference between the survival times of sporadic strains of A. baumannii and outbreak strains (27.2 versus 26.5 days, respectively; P < or = 0.44) by the Wilcoxon-Mann-Whitney test. All investigated A. baumannii strains, irrespective of their areas of endemicity or epidemic occurrence, have the ability to survive for a long time on dry surfaces. Antimicrobial susceptibility testing showed that A. baumannii outbreak strains were significantly more resistant to various broad-spectrum antimicrobial agents than sporadic strains. Both desiccation tolerance and multidrug resistance may contribute to their maintenance in the hospital setting and may explain in part their propensity to cause prolonged outbreaks of nosocomial infection.
Acinetobacter↗
Molecular epidemiology of Stenotrophomonas maltophilia isolated from clinical specimens from patients with cystic fibrosis and associated environmental samples.
Stenotrophomonas maltophilia was isolated from the respiratory tracts of 41 (25%) of 163 children attending our pediatric cystic fibrosis unit between September 1993 and December 1995. The extents of S. maltophilia contamination of environmental sites frequented by these patients were investigated with a selective medium incorporating vancomycin, imipenem, and amphotericin B. Eighty-two isolates of S. maltophilia were cultured from 67 different environmental sites sampled between January and July 1996. The organism was widespread in the home environment, with 20 (36%) and 25 (42%) of sampled sites positive in the homes of colonized and noncolonized patients, respectively. In the nosocomial setting, it was isolated from 18 (32%) sites in the hospital ward and from 4 (17%) sites in the outpatient clinic area. The most common sites of contamination were sink drains, faucets, and other items frequently in contact with water. All environmental and clinical isolates were genotyped with enterobacterial repetitive intergenic consensus sequences as primers. A total of 33 of the 41 patients were colonized with unique strains, and four pairs of patients shared strains. Further characterization by pulsed-field gel electrophoresis after digestion with XbaI found that there was no evidence of patient-to-patient transmission; however, there was some evidence that a small number of patients may have acquired the organism from the hospital environment. Resampling of environmental sites in the hospital ward in January 1997 revealed evidence of genetic drift, complicating the accurate determination of environmental sources for clinical strains. The source of the majority of S. maltophilia strains colonizing the respiratory tracts of these patients with cystic fibrosis remained uncertain but may have represented multiple, independent acquisitions from a variety of environmental sites both within and outside the hospital.
Adolescent↗
Quinolones in sweat and quinolone resistance.
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Anti-Infective Agents↗
The tet(M) gene of a Tn1545-like transposon and Neisseria gonorrhoeae.
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DNA Transposable Elements↗
Prevalence and mechanism of resistance to 'third-generation' cephalosporins in clinically relevant isolates of Enterobacteriaceae from 43 hospitals in the UK, 1990-1991.
In a UK survey of the occurrence of extended spectrum beta-lactamases, 96 hospitals submitted a total of 3951 non-selected, non-duplicate isolates of Enterobacteriaceae from 100 patients in each hospital, 206 of these cultures being mixed and, therefore, discarded. These isolates were initially screened for strains likely to produce extended-spectrum beta-lactamases (ESBLs) by MIC determination of beta-lactams followed by a bioassay, then disc approximation test and isoelectric focusing (IEF). Isolates were further examined using two pairs of PCR primers for both blaTEM and blaSHV genes. The ability of isolates to transfer resistance to both cefotaxime and ceftazidime by conjugation and transformation were examined. Four hundred and nine cefotaxime/ceftazidime-resistant isolates (10.9%) were identified from the 3745 submitted isolates, of which 338 (9.0%) were Enterobacteriaceae, 29 Escherichia coli, 35 Klebsiella spp. and seven Hafnia alveii. IEF suggested that 17 isolates produced an ESBL, which was confirmed in most cases by PCR and hydrolysis, five isolates produced an SHV enzyme by IEF, but not confirmed by PCR, and 11 had isoelectric points in the range 8-9 suggesting a possible AmpC enzyme. Only two isolates transferred the determinants. In the case of the Klebsiella spp., 19 of the 24 ceftazidime-resistant/clavulanate-sensitive isolates were positive by PCR for a blaSHV gene. No isolates were identified as carrying blaTEM, although eight isolates had isoelectric points of 5-6.3, suggesting the presence of a possible TEM beta-lactamase. The results for the H. alveii isolates suggest that either an AmpC-like enzyme or a transferable beta-lactamase which is not TEM/SHV is present. This study shows that a wide range of genotypically and phenotypically different isolates of Enterobacteriaceae producing ESBL-like enzymes is present throughout the UK at a frequency of about 1% of unselected isolates. It is important that surveillance of resistance to these clinically important antibiotics is maintained as the occurrence of localized or more widespread outbreaks caused by bacteria producing ESBLs is to be expected.
Cephalosporins↗
Molecular evolution of the tet(M) gene in Gardnerella vaginalis.
Five clinical isolates of Gardnerella vaginalis known to carry the tetracycline resistance determinant Tet M were examined by hybridization and nucleotide sequencing. Four of the strains carried tet(M) genes with identical sequences. The two versions of the tet(M) gene found in G. vaginalis did not show complete identity with other published tet(M) sequences, but showed mosaic structures with regions of homology to tet(M) gene sequences from Tn916, Tn1545 and the American type plasmid found in Neisseria gonorrhoeae. Hybridization studies showed that all isolates carried the tet(M) gene on a single HindII restriction fragment of variable length. No evidence was found for the presence of sequences homologous to the transposition functions of Tn916.
Base Sequence↗
Transcontinental importation into the UK of Escherichia coli expressing a plasmid-mediated AmpC-type beta-lactamase exposed during an outbreak of SHV-5 extended-spectrum beta-lactamase in a Leeds hospital.
Sixteen strains of Escherichia coli with high-level resistance to extended-spectrum cephalosporins and other classes of antibiotic have been isolated at St James' University Hospital, Leeds. They produce up to three separate beta-lactamases: TEM-1, SHV-5 and, in five isolates, a plasmid-mediated AmpC-type enzyme. With the exception of carbapenems, the isolates reported in this study were resistant to all beta-lactam antibiotics including extended-spectrum cephalosporins and the monobactam aztreonam. There was evidence of the spread of a plasmid encoding SHV-5, particularly amongst patients on the liver transplant unit. Sensitivity to beta-lactam antibiotics in five isolates expressing the AmpC-type beta-lactamase was not restored by the beta-lactamase inhibitor clavulanic acid. These bacteria also carried blaSHV-5 on a large plasmid. PCR-amplification of the structural gene and digestion with restriction endonucleases demonstrated that the plasmid-mediated blaAmpC probably identified as BIL-1 using the criteria available. Four of the five patients carrying isolates that carried the plasmid-located blaAmpC gene had recently visited the Indian subcontinent and we presume that they returned carrying these bacteria. Restriction fragment length polymorphism analysis using pulsed field gel electrophoresis (PFGE) suggests that at least four distinct strains existed amongst these five isolates. The two isolates that had very similar PFGE patterns had different plasmid profiles and were isolated from different locations in the hospital and at different times. This study demonstrates the ease with which highly resistant bacteria can be imported into the UK and spread within hospitals.
Cross Infection↗
Comparison and application of ribosome spacer DNA amplicon polymorphisms and pulsed-field gel electrophoresis for differentiation of methicillin-resistant Staphylococcus aureus strains.
Analysis of sequences in the fragments of the 16S-23S rRNA intergenic spacer region by the ribosome spacer PCR (RS-PCR) can differentiate strains of methicillin-resistant Staphylococcus aureus (MRSA). We compared this technique with pulsed-field gel electrophoresis (PFGE) for typing MRSA strains and its application during an investigation of an outbreak. A total of 180 isolates of MRSA collected from various hospital laboratories within the United Kingdom and elsewhere were typed by PFGE and RS-PCR. PFGE identified 17 different types among the 180 strains examined, and RS-PCR generated 13 different types. PFGE could detect minor genetic variations among the isolates and could identify the variants which were not discriminated by RS-PCR. Four unique strain types detected by PFGE were not detected by RS-PCR. When applied to typing the outbreak-related strains from the vascular surgery unit at the General Infirmary at Leeds, the results of RS-PCR were identical to those of PFGE. Our results have shown that RS-PCR is a rapid, inexpensive technique that is highly reproducible and almost as discriminatory as PFGE for typing MRSA isolates and should be useful in the local investigation of MRSA outbreaks.
DNA, Bacterial↗
A new selective differential medium for isolation of Stenotrophomonas maltophilia.
A new selective differential medium for the isolation of Stenotrophomonas (formerly Xanthomonas) maltophilia was developed. The medium, VIA agar, contained vancomycin, imipenem, and amphotericin B as selective agents and incorporated a mannitol/bromothymol blue indicator system. Compared with Xanthomonas maltophilia Selective Medium (XMSM), VIA agar was less inhibitory to Stenotrophomonas maltophilia and was more selective than XMSM in preventing the growth of unwanted bacteria from contaminated specimens. Although vancomycin-resistant strains of Enterococcus faecium may grow on VIA agar, these can be easily distinguished from Stenotrophomonas maltophilia because of mannitol fermentation.
Amphotericin B↗
Detection of mutations conferring extended-spectrum activity on SHV beta-lactamases using polymerase chain reaction single strand conformational polymorphism (PCR-SSCP).
Single strand conformational polymorphism (SSCP) is a recently developed technique used to detect single base mutations in short PCR-generated amplimers. The method has been adapted and applied to differentiation of beta-lactamase genes. Each of the five standard SHV strains used produced a unique SSCP pattern, allowing the possibility of rapid identification of the SHV genes of other isolates. A clinical isolate that phenotypically produced SHV-5 yielded a pattern of major bands indistinguishable from that of the SHV-5 standard strain, illustrating the applicability of this technique. We therefore report a reliable and reproducible technique that can be applied to the characterisation of the SHV beta-lactamases.
Base Sequence↗
Different effects of zinc ions on in vitro susceptibilities of Stenotrophomonas maltophilia to imipenem and meropenem.
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Cations, Divalent↗
Influence of relative humidity and suspending menstrua on survival of Acinetobacter spp. on dry surfaces.
Acinetobacter spp. are being reported with increasing frequency as a cause of nosocomial infection and have been isolated from the skin of healthy individuals, patients, hospital staff, dry nonbiotic objects, and different pieces of medical equipment. Factors affecting the survival of Acinetobacter spp. under conditions closely similar to those found in the hospital environment were investigated in the present study to help us understand the epidemiology of nosocomial Acinetobacter infection. Bacterial cells were suspended in distilled water or bovine serum albumin and were dried onto glass coverslips and kept at different relative humidities. Cells washed from coverslips were used to determined viable counts. Freshly isolated strains of Acinetobacter spp. belonging to the clinically important Acinetobacter calcoaceticus-Acinetobacter baumannii complex were found to be more resistant to drying conditions (e.g., 30 days for A. baumannii 16/49) than American Type Culture Collection strains (e.g., 2 days for A. baumannii ATCC 9955). The majority of strains belonging to the Acb complex had survival times similar to those observed for the gram-positive organism Staphylococcus aureus tested in the experiment. Survival times were prolonged for almost all the strains tested when they were suspended in bovine serum albumin (e.g., 60 days for A. baumannii R 447) compared with those for strains suspended in distilled water (11 days for R 447). The survival times for strains at higher relative humidity (31 or 93%) were longer than those for strains of Acinetobacter kept at a relative humidity of 10% (11 days at 31% relative humidity and 4 days at 10% relative humidity for R447). These findings are consistent with the observed tendency of Acinetobacter spp. to survive on dry surfaces, and they can be transferred not only by moist vectors but also under dry conditions in a hospital environment during nosocomial infection outbreaks. The results obtained in the experiment support the previously suggested airborne spread of Acinetobacter spp. in hospital wards and repeated outbreaks after incomplete disinfection of contaminated dry surfaces.
Acinetobacter↗