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K J Towner

Publications and source records attributed to K J Towner.

At least 19 recordsLinked to original sources
PubMed2006

Evaluation of the sensitivity of a rapid polymerase chain reaction for detection of group B streptococcus.

The objective of this study was to compare detection of group B streptococcal (GBS) carriage using 'real-time' polymerase chain reaction (PCR) and microbiological standard culture. The study design was a test accuracy study comparing a novel molecular technique against the standard microbiological cultural technique in normal pregnant women. The setting and population consisted of 143 pregnant women with pre-labour rupture of the membranes, recruited from two large teaching hospitals in the UK. The study examined the efficacy of a polymerase chain reaction (PCR) assay for screening pregnant women who presented with term rupture of the membranes. Low vaginal specimens were obtained from the women. The specimens were tested for GBS by conventional culture and with a GBS-specific real-time PCR assay. The main outcome measure was the sensitivity and specificity of the PCR assay with 95% confidence intervals (CI) compared with the standard culture. The length of time to obtain a result was also reported for both methods. Among the 143 women, the results of the culture were positive (at least one colony) for GBS in 20 women (14%). The PCR assay detected GBS carriage in 10 women (7%). As compared with the culture method, the sensitivity and specificity of the PCR assay were 45% and 99%, respectively. The positive and negative predictive values of the PCR assay were 90% and 92%, respectively. The length of time required to obtain results for the majority of women (94%) was <2.5 h for the PCR assay and at least 24 h for culture. While a rapid result (within 3 h) of carriage of GBS can be obtained by the PCR assay, at present, it cannot replace conventional culture without further optimisation of the DNA extraction method. The sensitivity may further be improved by testing both low vaginal and rectal specimens.

Carrier State↗
PubMed2005

Rapid detection of methicillin-resistant Staphylococcus aureus from screening enrichment broths by real-time PCR.

The aim of this study was to evaluate real-time PCR (LightCycler) kits for the detection of methicillin-resistant Staphylococcus aureus in enrichment broth using 200 sets of patient screening swab samples. One hundred sets were processed using a lysostaphin/lysozyme extraction method and a further 100 were tested using a column extraction kit. The PCR kits with lysostaphin/lysozyme lysis showed 95.7% sensitivity, 90.8% specificity, and negative and positive predictive values of 98.6% and 75.9%, respectively, compared with 88%, 95.9%, 84.6% and 95.9%, respectively, with the column extraction kit.

Bacterial Typing Techniques↗
PubMed2005

Prevalence of Acinetobacter baumannii and other Acinetobacter spp. in faecal samples from non-hospitalised individuals.

In total, 226 individuals from the community were investigated for faecal carriage of Acinetobacter spp. by broth enrichment culture, followed by growth on blood agar and/or Leeds Acinetobacter Medium (LAM). Acinetobacter baumannii was isolated on both LAM and blood agar from one of 100 specimens in the UK and one of 126 specimens in The Netherlands. The predominant species were Acinetobactor johnsonii and genomic sp. 11, which were cultured from 22 and five specimens, respectively. A. baumannii did not seem to be widespread in the faecal flora of individuals in the community.

Acinetobacter Infections↗
PubMed2005

Report of the Consensus Conference on Antibiotic Resistance; Prevention and Control (ARPAC).

Antimicrobial resistance is a key public health concern in Europe. It is known that there are significant variations in the prevalence of resistance across Europe, and methods to reduce the problem are also assumed to vary significantly. The 'Antibiotic Resistance; Prevention and Control (ARPAC)' Concerted Action project was funded by the European Commission and conducted by four study groups of the European Society of Clinical Microbiology and Infectious Diseases (ESCMID). The project established a network of European hospitals and collated data on antimicrobial resistance prevalence, antimicrobial susceptibility testing methods, typing methods employed, antimicrobial use, antibiotic policies and practices, and infection control policies and practices. The ARPAC Consensus Conference, entitled 'Control of antibiotic resistance in European hospitals-informing future evidence-based practice', was held in Amsterdam in November 2004. The conference was co-hosted by the European Commission, ESCMID and the Dutch Working Party on Antibiotic Policy (SWAB). Key ARPAC findings were presented and discussed in the context of the worldwide situation. The conference delivered a set of high-priority recommendations likely to have a significant impact on antimicrobial resistance. This report summarises these recommendations.

Anti-Bacterial Agents↗
PubMed2003

Evaluation of a real-time PCR hybridization assay for rapid detection of Legionella pneumophila in hospital and environmental water samples.

A real-time LightCycler assay for Legionella pneumophila was evaluated with 120 water samples potentially contaminated with PCR inhibitors. Results were obtained within five hours, with a detection limit equivalent to 800 cells/L. However, 11 of 22 culture-positive samples containing < 100 CFU/L were also positive by LightCycler assay, indicating the presence of significant numbers of non-viable cells. Following extraction, amplification inhibitors remained in four culture-positive samples, but only one contained > 800 CFU/L. The assay seemed suitable for rapidly screening large sample numbers for heavy contamination with L. pneumophila, but conventional culture should continue to be used to detect low contamination levels.

Legionella pneumophila↗
PubMed2002

Diversity of Staphylococcus aureus strains isolated from two European regions with different prevalences of methicillin resistance.

The genodiversity of Staphylococcus aureus isolates from the Nottingham region of the United Kingdom was compared with isolates from the Freiburg region of Germany. The prevalence of methicillin-resistant Staphylococcus aureus (MRSA) isolates was higher in Nottingham than in Freiburg. In patients from Nottingham hospitals, 80% of MRSA isolates were classical epidemic MRSA-15, but genotypic variants of epidemic MRSA-15 comprised 72% of isolates from Nottingham community-based patients. In contrast, MRSA isolates from Freiburg showed greater diversity, but 47% and 23% of isolates, respectively, belonged to two predominant MRSA genotypes found in isolates from both hospitalised and community-based patients. The results suggest that genodiversity becomes increasingly more confined in settings with a higher frequency and longer duration of MRSA prevalence.

Genetic Variation↗
PubMed2002

Endemic carbapenem resistance associated with OXA-40 carbapenemase among Acinetobacter baumannii isolates from a hospital in northern Spain.

Eighty-two carbapenem-resistant isolates of Acinetobacter baumannii from a single hospital in Bilbao were typed into two major clusters and several subclusters. Disk synergy tests and PCR indicated the production of a zinc-independent OXA-class carbapenemase. Sequencing identified this enzyme, OXA-40, as a variant of the OXA-24-OXA-25-OXA-26 cluster.

Acinetobacter Infections↗
PubMed2000

Use of an automated DNA analysis system (DARAS) for sequence-specific recognition of Neisseria meningitidis DNA.

OBJECTIVES: To combine use of the polymerase chain reaction (PCR) for rapid diagnosis of meningococcal meningitis with a novel automated detection system for sequence-specific recognition of PCR products. METHODS: DNA was extracted from cerebrospinal fluid (CSF) by a quick boil-lysis method, followed by PCR with primers specific for Neisseria meningitidis. Sequence-specific recognition of N. meningitidis DNA was performed with an automated DNA analysis system (DARAS) and the data were compared with results following agarose gel electrophoresis or conventional microbiological culture. RESULTS: The DARAS system had a sensitive detection limit of 102 meningococci/mL with spiked samples, compared with a detection limit of 104 meningococci/mL following agarose gel electrophoresis. When the system was used to examine 74 CSF samples, the 19 CSF samples positive for N. meningitidis by conventional microbiological methods were also all positive in the DARAS system and the 55 samples negative by DARAS for meningococci were also negative by conventional microbiological methods. CONCLUSION: The sensitivity and specificity of the DARAS system makes it a useful tool for the diagnosis of meningococcal meningitis. The system is user-friendly, requires minimal hands-on time and generates data in an informative numerical format.

Colony Count, Microbial↗
PubMed2000

Use of RAPD-ALF analysis for investigating the frequency of bacterial cross-transmission in an adult intensive care unit.

Bacterial cross-transmission was investigated during a 12-month period in an adult intensive care unit (ICU) by the generation of random amplified polymorphic DNA (RAPD) fingerprinting profiles, combined with automated laser fluorescence (ALF) analysis. The potential episodes of cross-transmission identified, were compared with those detected by the conventional first-line screen of antibiogram typing. Over the year, 215 primary gram-negative bacterial isolates were obtained from 160 patients. In total, 22 possible episodes of cross-transmission, involving 70 (44%) of the 160 patients, were identified by RAPD-ALF analysis, and 19 of these were substantiated with epidemiological evidence. Conversely, 31 possible episodes were identified on the basis of antibiogram data, but only three of these episodes, two involving Acinetobacter baumannii and one involving Serratia marcescens, correlated with those identified by RAPD-ALF analysis. It was concluded that analysis of antibiogram data alone is an unreliable method for assessing bacterial cross-transmission, unless the organism involved has a particularly stable or unusual resistance pattern. In contrast, the technique of RAPD-ALF analysis may provide a rapid and simple technique for obtaining an insight into the population dynamics of gram-negative bacteria in adult ICUs.

Acinetobacter↗
PubMed2000

Evaluation of a PCR-immunoassay technique for detection of Neisseria meningitidis in cerebrospinal fluid and peripheral blood.

A multiplex PCR-immunoassay for the rapid diagnosis of bacterial meningitis from cerebrospinal fluid (CSF) or peripheral blood was compared with conventional microbiological techniques used in the routine diagnostic laboratory. The multiplex PCR was designed to detect simultaneously a universal conserved sequence coding for bacterial 16S rRNA and the Neisseria meningitidis porA gene. The PCR-immunoassay had a detection limit of (3-5) x 10(2) cfu/ml (equivalent to 1-3 cfu/PCR) with spiked CSF or blood samples, compared with (3-5) x 10(3) cfu/ml for PCR followed by conventional agarose gel electrophoresis for detection of PCR products. Of 294 CSF samples from patients suspected on clinical grounds of suffering from meningitis, the PCR-immunoassay detected bacterial DNA in 29 CSF samples, 15 of which were also positive for N. meningitidis DNA. The 29 positive CSF samples comprised 25 samples that were also reported positive and four that were reported negative by conventional methodology; the latter four were all positive for N. meningitidis by the PCR-immunoassay. Of 173 peripheral blood samples examined, the PCR-immunoassay detected bacterial DNA in 18 samples, 14 of which were also positive for N. meningitidis DNA. In comparison, only 10 samples were reported positive for N. meningitidis by conventional methodology. All negative PCR-immunoassay results correlated with those obtained by conventional methodology for both CSF and blood samples. The sensitivity and speed of the PCR-immunoassay system indicated that it could be used as a routine diagnostic test for meningococcal meningitis, enabling a diagnosis to be made within 4 h of receipt of the specimen.

Bacteremia↗
PubMed2000

Detection of Legionella pneumophila using a real-time PCR hybridization assay.

A real-time PCR hybridization assay for Legionella pneumophila is described; the assay uses LightCycler (Idaho Technology) methodology to specifically detect 2.5 CFU/reaction, equivalent to 1,000 CFU/liter of starting water sample. The assay, including DNA extraction and confirmation of product identity, is completed within 90 min of receipt of a sample.

DNA, Bacterial↗
PubMed1999

Molecular and antibiogram relationships of Acinetobacter isolates from two contrasting hospitals in the United Kingdom and South Africa.

The aim of this study was to compare the molecular relationships and antibiograms of nosocomial isolates of Acinetobacter spp. from two acute-care hospitals in Nottingham, UK, and Soweto, South Africa, with different hospital infection control problems and procedures. In contrast to Nottingham, where randomly amplified polymorphic DNA fingerprinting demonstrated that a single multiresistant strain of Acinetobacter baumannii has predominated in the hospital intensive care unit over an 11-year period, the Soweto isolates formed a heterogeneous group of unrelated molecular clusters of different antibiograms, with numerous different strains of Acinetobacter baumannii, Acinetobacter sp. 3 and Acinetobacter sp. 13TU apparently being endemic throughout the Soweto hospital. The contrasting results illustrate the need to maintain exemplary infection control procedures in hospitals where high standards have been achieved and warn of what might result if such measures are diminished.

Acinetobacter↗
PubMed1999

Evaluation of the mecA femB duplex polymerase chain reaction for detection of methicillin-resistant Staphylococcus aureus.

This study systematically evaluated a recently described duplex polymerase chain reaction test for methicillin-resistant Staphylococcus aureus with 25 different German epidemic strains of methicillin-resistant Staphylococcus aureus and 66 staphylococci other than methicillin-resistant Staphylococcus aureus, including 17 different coagulase-negative staphylococcal species and subspecies, that were either oxacillin susceptible or oxacillin resistant. The results were compared with those of conventional cultural identification and susceptibility testing. Of the 91 isolates tested, all 25 confirmed strains of methicillin-resistant Staphylococcus aureus were identified correctly. None of the remaining strains of methicillin-susceptible Staphylococcus aureus was misidentified as methicillin-resistant Staphylococcus aureus. It was concluded that the duplex polymerase chain reaction appears to offer a time-saving and accurate method of detection of methicillin-resistant Staphylococcus aureus.

Bacterial Proteins↗
PubMed1999

Increase in incidence of resistance to ampicillin, chloramphenicol and trimethoprim in clinical isolates of Salmonella serotype Typhimurium with investigation of molecular epidemiology and mechanisms of resistance.

Antimicrobial resistance patterns of Salmonella serotype Typhimurium isolates obtained during the period 1987-1994 were examined and the molecular epidemiology and the mechanisms of resistance to ampicillin, chloramphenicol and trimethoprim were investigated in 24 strains isolated during 1994. Resistance to ampicillin increased from 18% to 78%, to chloramphenicol from 15% to 78%, to tetracycline from 53% to 89% and to co-trimoxazole from 3% to 37%, whereas resistance to norfloxacin remained at 0%. Of Salmonella serotype Typhimurium strains isolated during 1994, all ampicillin-resistant strains had an MIC > 256 mg/L, except one strain in which the MIC was 64 mg/L. Twelve strains (52%) had a TEM-type beta-lactamase, nine (39%) a CARB-type beta-lactamase and two strains (8%) had an OXA-type beta-lactamase. Chloramphenicol acetyl-transferase activity was detected in only nine (47%) of 19 chloramphenicol resistant strains, whereas all eight trimethoprim-resistant strains produced a dihydrofolate reductase type Ia enzyme. Three different epidemiological groups were defined by either low-frequency restriction analysis of chromosomal DNA and pulsed-field gel electrophoresis or repetitive extragenic palindromic-PCR. The latter technique provided an alternative, rapid and powerful genotyping method for S. Typhimurium. Although quinolones provide a good therapeutic alternative, the multiresistance of S. Typhimurium is of public health concern and it is important to continue surveillance of resistance levels and their mechanisms.

Ampicillin Resistance↗
PubMed1998

Surveillance of an adult intensive care unit for long-term persistence of a multi-resistant strain of Acinetobacter baumannii.

Sporadic infections with Acinetobacter spp., punctuated with prolonged outbreaks of infection involving larger numbers of patients and a particular epidemic strain of Acinetobacter baumannii, have occurred in the adult intensive care unit (ICU) of Nottingham University Hospital since 1985. The aim of this study was to screen patients admitted to the ICU for three or more days during a non-outbreak period in 1994-1995 and to use DNA fingerprinting techniques to compare any isolates of Acinetobacter spp. with isolates obtained from the same ICU during the previous ten years. In the present study, almost 20% of the ICU patients screened during 1994-1995 became colonized with Acinetobacter spp. The commonest species isolated from patients was Acinetobacter baumannii; five different strains were identified by random amplified polymorphic DNA fingerprinting, including the epidemic strain responsible for outbreaks of infection in 1985-1986 and 1992-1993. Environmental sampling yielded Acinetobacter spp. from one or more samples on four occasions; Acinetobacter radioresistens was the commonest species isolated, and Acinetobacter baumannii (not the epidemic strain) was isolated on only one occasion from the environment. The long-term persistence of a potentially epidemic strain in the ICU, even during a non-outbreak period, indicates a need for continued vigilance. Consequently, periodic patient and environmental surveillance, combined with typing of isolates, is recommended for ICUs where significant outbreaks of Acinetobacter infection have occurred in the past.

Acinetobacter↗
PubMed1998

Molecular epidemiology of aminoglycoside resistance in Acinetobacter spp.

Most aminoglycoside resistance in Acinetobacter spp. involves production of aminoglycoside-modifying enzymes. Previous studies have shown that the genes encoding these enzymes can be present on plasmids, transposons or within integron-type structures. To determine whether particular mechanisms of aminoglycoside resistance have developed in strains from specific geographical locations (with subsequent clonal spread), or whether common mechanisms have been acquired by genotypically distinct clinical isolates of Acinetobacter spp. throughout the world, a genotypically heterogeneous collection of 24 multiresistant clinical isolates of Acinetobacter spp. from 15 hospitals in 11 countries worldwide was studied. All were resistant to two or more aminoglycoside antibiotics. The full aminoglycoside resistance profile was determined for each isolate, allowing a putative enzyme content to be inferred, with subsequent confirmation of enzyme content and genetic location by polymerase chain reaction (PCR) and hybridisation techniques. All produced at least one aminoglycoside-modifying enzyme, most commonly AAC(3)-I and ANT(3'')-I in various combinations. Other enzymes found were AAC(3)-II, AAC(6')-I, ANT(2''), APH(3')-I and APH(3')-VI. None was confined to strains from a particular geographical area. Nine isolates transferred resistance mediated by AAC(3)-I, ANT(2'')-I, APH(3')-I or APH(3)'-VI by conjugation to a sensitive strain of A. baumannii, but most resistance was non-transferable. PCR mapping revealed an integron location in six isolates for the aac(3)-Ia gene and in three isolates for the ant(3'')-Ia gene. Overall, the study demonstrated that similar aminoglycoside-modifying enzymes are found in unrelated isolates of Acinetobacter spp., and that particular genes are not restricted to specific areas of the world. The demonstration of certain genes on plasmids and integrons emphasises the probable importance of these structures in the dissemination of certain types of aminoglycoside resistance in Acinetobacter spp.

Acinetobacter↗
PubMed1998

Development and evaluation of a PCR-based immunoassay for the rapid detection of methicillin-resistant Staphylococcus aureus.

A multiplex polymerase chain reaction (PCR), involving detection of the mecA and femB genes, was combined with a novel immunoassay system capable of detecting specific PCR products. The resulting PCR-immunoassay was evaluated in comparison with conventional microbiological techniques used in the routine diagnostic laboratory for the rapid identification of methicillin-resistant Staphylococcus aureus (MRSA), either in pure culture or in overnight broth cultures obtained following enrichment of patient screening swabs. Among the 480 purified isolates of staphylococci and 246 enrichment broths examined, only one 'false-negative' result was obtained by PCR, compared with 18 'false-negative' results obtained by conventional methodology and demonstrated by further conventional examination. Five demonstrable 'false-positive' results were obtained by conventional methodology, compared with a possible 10 by the PCR-immunoassay, although it was not certain that these 10 PCR results were true 'false positives' as, by definition, MRSA could not be isolated by conventional methodology. The results indicated that the routine diagnostic laboratory was encountering difficulties in identifying MRSA correctly, and that the conventional microbiological techniques lacked sensitivity. Overall, the PCR technique was more accurate and sensitive than conventional methodology in detecting MRSA, and results were available within 24 h of screening swabs arriving in the laboratory, compared with a minimum of 48-72 h by conventional techniques. The immunoassay system added to the usefulness of the method by allowing the detection of specific PCR products within 5 min of completing the PCR, without the normal additional step of agarose gel electrophoresis.

Bacterial Proteins↗