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P M Hawkey

Publications and source records attributed to P M Hawkey.

At least 73 records · Page 4Linked to original sources
PubMed1996

Validation of use of whole-cell repetitive extragenic palindromic sequence-based PCR (REP-PCR) for typing strains belonging to the Acinetobacter calcoaceticus-Acinetobacter baumannii complex and application of the method to the investigation of a hospital outbreak.

Acinetobacter spp. are being reported with increasing frequency as causes of nosocomial infection. In order to identify reservoirs of infection as quickly as possible, a rapid typing method that can differentiate epidemic strains from environmental and nonepidemic strains is needed. In 1993, a cluster of Acinetobacter baumannii isolates from five patients in the adult intensive therapy unit of our tertiary-care teaching hospital led us to develop and optimize a rapid repetitive extragenic palindromic sequence-based PCR (REP-PCR) typing protocol for members of the Acinetobacter calcoaceticus-A. baumannii complex that uses boiled colonies and consensus primers aimed at repetitive extragenic palindromic sequences. Four of the five patient isolates gave the same REP-PCR typing pattern as isolates of A. baumannii obtained from the temperature probe of a Bennett humidifier; the fifth isolate had a unique profile. Disinfection of the probe with 70% ethanol, as recommended by the manufacturer, proved ineffective, as A. baumannii with the same REP-PCR pattern was isolated from it 10 days after cleaning, necessitating a change in our decontamination procedure. Results obtained with REP-PCR were subsequently confirmed by ribotyping. To evaluate the discriminatory power (D) of REP-PCR for typing members of the A. calcoaceticus-A. baumannii complex, compared with that of ribotyping, we have applied both methods to a collection of 85 strains that included representatives of six DNA groups within the complex. Ribotyping using EcoRI digests yielded 53 patterns (D = 0.98), whereas 68 different REP-PCR patterns were observed (D = 0.99). By computer-assisted analysis of gel images, 74 patterns were observed with REP-PCR (D = 1.0). Overall, REP-PCR typing proved to be slightly more discriminatory than ribotyping. Our results indicate that REP-PCR typing used boiled colonies is a simple, rapid, and effective means of typing members of the A. calcoaceticus-A. baumannii complex.

Acinetobacter↗
PubMed1995

Molecular epidemiology of ceftazidime resistant Enterobacteriaceae from patients on a paediatric oncology ward.

Between the autumn of 1989 and January 1990, 21 of the 44 children on the paediatric oncology ward of St. James's University Hospital, Leeds, UK were infected or colonised with Enterobacteriaceae producing extended-spectrum beta-lactamases. This represents 48% of the patients on the ward. Only six patients (14%) had microbiologically proven septicaemia caused by such bacteria during this period. Eighty-one isolates of Enterobacteriaceae producing extended-spectrum beta-lactamases derived from blood culture (7 isolates from 6 patients) or faecal samples (74 isolates) were available for examination. These comprised 28 Escherichia coli, 28 Klebsiella oxytoca, 11 Klebsiella pneumoniae, 10 Citrobacter freundii, 3 Enterobacter spp. and 1 Serratia marcescens. Clinical isolates were resistant to penicillins and to ceftazidime. Strains isolated in this study also showed multiple resistance to a range of antimicrobial agents. Transfer to a nalidixic acid resistant laboratory strain of E. coli UB5201 was attempted, but transfer of the ceftazidime resistance determinant was only successful in 25 isolates (31%). Examination of plasmid DNA revealed sequences in each isolate that hybridised with the TEM beta-lactamase gene probe used on a variety of plasmids ranging in size from 2.5- > 150 kb, sometimes found on several replicons in a single isolate. The TEM gene probe also hybridised with chromosomal DNA in a large number of isolates. Nucleotide sequence analysis demonstrated the presence of three extended-spectrum beta-lactamases: TEM-10B produced by two isolates, TEM-12B produced by 37 isolates and TEM-26B produced by 40 isolates. In two cases, isolates produced two beta-lactamases, and it proved impossible to identify these enzymes unequivocally. The genes encoding TEM-10B and TEM-26B both differ from TEM-12B by single nucleotide substitutions. Analysis of the ribotype patterns derived from the clinical isolates provided evidence for cross-colonisation between patients, and this was confirmed by analysis of the plasmid profiles. Four years after discontinuing ceftazidime and other extended-spectrum cephalosporins on this ward, patients were still colonised with bacteria that produced extended-spectrum beta-lactamases.

Base Sequence↗
PubMed1995

Replacement of NCTC 4175, the current type strain of Proteus vulgaris, with ATCC 29905. Request for an opinion.

The current type strain of Proteus vulgaris, NCTC 4175 (= ATCC 13315), differs substantially from typical strains of this species both biochemically and chemotaxonomically. DNA relatedness studies revealed that strains previously classified as P. vulgaris belong to six genomospecies. One of these genomospecies contains strains that are negative in indole, salicin, and esculin reactions (biogroup 1) and has been named Proteus penneri. A second genomospecies, which is most frequently isolated from human urine, contains typical P. vulgaris strains that are positive in indole, salicin, and esculin reactions (biogroup 2). The members of the remaining four genomospecies are indole positive and negative in salicin and esculin reactions (biogroup 3). Of 36 biogroup 3 strains studied, only strain NCTC 4175T (T = type strain) and one other strain, CDC 1732-80, belong to genomospecies 3. To retain NCTC 4175 as the type strain of P. vulgaris would restrict this species to these two strains, whose origins are unknown. This would mean that hundreds of strains for which the description of P. vulgaris was written and which have been representatives of this species for the past 50 years would have to be renamed as members of a new species. To prevent this confusion, we request that biogroup 2 reference strain ATCC 29905 (= CDC PR1) replace NCTC 4175 as the type strain of P. vulgaris.

DNA, Bacterial↗
PubMed1995

Campylobacter jejuni in the stomach.

Campylobacter jejuni is the commonest cause of acute bacterial enteritis in the UK. However, in this case a 74-year-old lady underwent gastroscopy for an upper gastrointestinal haemorrhage and was noted to have a gastric ulcer. Gastric biopsy revealed spiral gram-negative bacteria and culture yielded a moderate growth of C. jejuni. Identification was confirmed by growth characteristics, biochemical tests and PCR amplification of the species-specific flagellin gene--fla A. To prevent misidentification, it is important that laboratories routinely culturing gastric biopsies for Helicobacter pylori should perform a rapid urease test and not rely solely on microscopic morphology.

Aged↗
PubMed1994

Convergent evolution of TEM-26, a beta-lactamase with extended-spectrum activity.

TEM-26, an extended-spectrum beta-lactamase has been characterized in clinical isolates of Klebsiella pneumoniae and Escherichia coli derived from patients on the Paediatric Oncology Unit of St James's University Hospital, Leeds. The nucleotide sequence of this beta-lactamase gene (blaTEM26b) was determined, and compared with the nucleotide sequences of other TEM-type beta-lactamases. The blaTEM26b gene was found to differ from blaTEM12b by a single nucleotide. This difference causes the substitution of glutamic acid in blaTEM12b for lysine in blaTEM26b at position 102 in the predicted amino acid sequence. The blaTEM12b gene was first described in an isolate of Klebsiella oxytoca from a patient nursed on the same unit that yielded the strains that carry blaTEM26b. However, the blaTEM26b gene differs at no less than six nucleotides from the nucleotide sequence encoding the TEM-26 beta-lactamase that was first described in isolates from cancer patients nursed in the Children's Hospital, Stanford, California, USA. This indicates that the genes encoding TEM-26 have evolved from different progenitors.

Amino Acid Sequence↗
PubMed1994

Description of Leeds Acinetobacter Medium, a new selective and differential medium for isolation of clinically important Acinetobacter spp., and comparison with Herellea agar and Holton's agar.

Acinetobacter spp. are responsible for an increasing number of opportunistic, nosocomial infections. They have been isolated from diverse inanimate objects in the hospital environment and are resistant to most of the commonly used antibiotics. Existing media for the isolation of Acinetobacter spp. are either nonselective, allowing the growth of unwanted bacteria, or too inhibitory, inhibiting the growth of many Acinetobacter strains. For the rapid isolation and effective control of Acinetobacter infection, a new selective and differential medium, Leeds Acinetobacter Medium (LAM), has been developed to isolate Acinetobacter spp. from clinical and environmental sources. The concentration of antibiotics and other ingredients in this medium have been determined according to the results of MIC and viable counts performed for these ingredients. LAM was compared with other selective and differential media for the isolation of Acinetobacter spp. from a local hospital environment and proved to be better in terms of recovery and selectivity.

Acinetobacter↗
PubMed1993

Gastroduodenal dysfunction and bacterial colonisation of the ventilated lung.

The source of ventilator-associated pneumonia (gastric or oropharyngeal flora) remains controversial. We investigated the source of bacterial colonisation of the ventilated lung in 100 consecutive intensive-care patients. Gram-negative bacilli were isolated from the lower respiratory tract in 19 patients. Bacteria isolated from the stomach contents either previously or at the same time were identical to lower respiratory isolates in 11 patients. No gram-negative oropharyngeal isolate was identical to a lower respiratory tract isolate. Gastric bacterial overgrowth with gram-negative bacilli was associated with the presence of bilirubin in the stomach contents. Detectable bilirubin was also associated with subsequent acquisition of gram-negative bacilli in the lower respiratory tract. Only 5 gastric aspirate specimens with pH < 3.5 contained gram-negative bacilli. These results establish a relation between duodenal reflux and subsequent bacterial colonisation of the lower respiratory tract. Restoration of normal gastroduodenal motility might help prevent pneumonia in intensive-care patients.

Adolescent↗
PubMed1993

Staphylococcal pneumonia in ventilated patients: a twelve-month review of cases in an intensive care unit.

We reviewed staphylococcal lower respiratory tract infections in our intensive care unit over a 12-month period. Staphylococcus aureus was isolated from tracheal aspirates more commonly in patients with intracranial trauma (P < 0.001), between one and six days (mean = 3) after admission to the intensive care unit. Bacteriophage typing of all S. aureus lower respiratory tract isolates from the 17 patients with head injury did not provide evidence for a common external source of infection or patient-to-patient transmission.

Bacteriophage Typing↗
PubMed1993

Co-existent cross-infection with Streptococcus pneumoniae and group B streptococci on an adult oncology unit.

An outbreak of respiratory infection in an adult oncology unit is described where there was evidence of co-existent cross-infection with Streptococcus pneumoniae serotype 14 and Lancefield group B streptococcus type Ia. The presumed route of transmission was droplet spread from patient to patient. No further cases arose after the ward had been closed to new admissions and all symptomatic patients were treated with erythromycin. We suggest that patients with pneumococcal pneumonia on units housing elderly debilitated patients should be isolated. Infection control teams should be aware of the ability of Lancefield group B streptococci to spread by cross-infection among adult patients.

Adult↗
PubMed1993

Effect of divalent cations in bacteriological media on the susceptibility of Xanthomonas maltophilia to imipenem, with special reference to zinc ions.

The susceptibility of Xanthomonas maltophilia strains to beta-lactams is known to vary according to the type of growth media used. We have assayed the divalent cation content of various susceptibility testing media and correlated this with the susceptibility to imipenem of 30 strains of X. maltophilia, by calculating the IC50. No correlation was found with Ca++ and Mg++ content (r = 0.005, P = 0.99), but a highly significant correlation with the Zn++ content (7.7-42.7 mumol/L) of the medium was found (r = 0.93, P = 0.003). The effect of Zn++ on the susceptibility of X. maltophilia to imipenem was further investigated by adding varying amounts of zinc sulphate to Oxoid Mueller-Hinton agar which has a low Zn++ content (14.2 mumol/L). A highly significant correlation between the Zn++ content and the IC50 was observed (r = 0.95, P = 0.001). Some variability was seen from one series of IC50 determinations to another and samples of ultra-pure water were processed in exactly the same fashion as the agar media and then assayed for cation content. No significant increase in Ca++ or Mg++ content of the water were observed but water autoclaved in universal containers with metal caps and rubber washers acquired up to 40 mumol/L of Zn++. Studies of the correlation between in-vitro sensitivity tests and the clinical performance of beta-lactams used against X. maltophilia will need to take account of the Zn++ content of the susceptibility testing media used.

Anti-Bacterial Agents↗
PubMed1993

The use of a DNA probe and PCR to examine the distribution of the aac(6')-Ic gene in Serratia marcescens and other gram-negative bacteria.

A nucleotide probe for the chromosomal aminoglycoside 6' acetyltransferase gene (aac(6')-Ic) of Serratia marcescens was used in non-radioactive dot-blot hybridization experiments with 186 strains belonging to ten different species of Serratia. The gene was only detected in strains of S. marcescens (all strains tested), and positive hybridization was seen irrespective of whether or not strains were kanamycin-resistant. An additional 180 strains belonging to 28 Gram-negative bacterial species other than in the genus Serratia did not hybridize with the probe. A rapid PCR test for the aac(6')-Ic gene was developed and used to confirm that the aac(6')-Ic gene is only found in S. marcescens. Southern hybridization analysis of S. marcescens chromosomal DNA demonstrated that the gene was usually located on a PvuII fragment with a putative isoleucine tRNA-2 gene, but polymorphisms with respect to the size of this fragment were observed.

Acetyltransferases↗
PubMed1993

Nucleotide sequences of the tet(M) genes from the American and Dutch type tetracycline resistance plasmids of Neisseria gonorrhoeae.

High-level tetracycline-resistant Neisseria gonorrhoeae (TRNG) has been associated with the presence of a plasmid approximately 25.2 MDa in size which carries a Tet M tetracycline resistance determinant. Two different plasmid types, American and Dutch, have previously been described, based on the restriction endonuclease digestion pattern. In this study, the tet(M) genes from the two plasmid types have been amplified by the polymerase chain reaction (PCR) and then sequenced. The gene sequences from the two plasmids shared 96.8% identity, and showed similarities with different segments of the tet(M) gene sequences from Tn1545, Tn916 and Ureaplasma urealyticum. The data suggest that it is highly likely that the Tet M determinant found in the American type plasmid has a different origin from that present in the Dutch plasmid.

Base Sequence↗
PubMed1993

Identification and typing of Proteus penneri and Proteus vulgaris biogroups 2 and 3, from clinical sources, by computerized analysis of electrophoretic protein patterns.

Seventy-six strains of the Proteus vulgaris complex (Pr. penneri and Pr. vulgaris biogroups 2 and 3) were characterized by one-dimensional SDS-PAGE of cellular proteins. The protein patterns were highly reproducible. The strains came from various countries and were mainly of human origin: urine (28), respiratory tract (13), wounds (8), faeces (7), blood (3), miscellaneous sources (6) and unknown sources (11). The patterns of these strains, together with those of the type strains of seven Morganella, Proteus and Providencia species were subjected to two numerical analyses. In the first, in which the principal protein bands (in the 35.0-42.0 kDa range) were excluded, the strains of the Pr. vulgaris complex formed four clusters at the 83% similarity level. These corresponded to Pr. penneri, Pr. vulgaris biogroup 2, and two clusters (3a and 3b) represented biogroup 3. Each of these clusters was distinct from the Morganella, Proteus and Providencia reference strains. In the second analysis, which included all the protein bands, the 41 Pr. penneri strains showed little heterogeneity but 17 subphenons could be recognized among the 35 strains of Pr. vulgaris biogroups 2 and 3. These results support the division of biogroup 3 strains into at least two separate taxa. Other results indicate that biogroup 3 is heterogeneous and may contain further genomic groups. The method also provides a basis for typing clinical strains of Pr. vulgaris biogroups 2 and 3.

Bacterial Proteins↗