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T L Pitt

Publications and source records attributed to T L Pitt.

At least 73 records · Page 4Linked to original sources
PubMed1993

Lipopolysaccharide profile typing as a technique for comparative typing of gram-negative bacteria.

We have applied the technique of lipopolysaccharide (LPS) profiling in sodium dodecyl sulfate-polyacrylamide gel electrophoresis to the typing of 124 isolates of 12 gram-negative species from suspected outbreaks of infection. LPS was prepared by proteinase K digestion or micro-phenol-water extraction. A total of 11 of the 12 species gave clear ladder band profiles, the exception being Acinetobacter baumannii. When compared with conventional typing for Enterobacter cloacae, Pseudomonas aeruginosa, and Serratia marcescens, LPS profile type alone was sufficient to allow relatedness or distinguishability of isolates to be established, and this was corroborated by serotype and phage type data. Serologically nontypeable isolates invariably lacked O repeating units and thus could not be classified by their silver stain profile. We conclude that LPS profiling is useful for the epidemiological investigation of small clusters of isolates in order to determine whether or not cross-infection between patients has occurred.

Bacterial Typing Techniques↗
PubMed1993

Relationships among Pseudomonas pseudomallei isolates from patients with recurrent melioidosis.

Patients with melioidosis may present with recurrent infections after clinical resolution of their primary illness. Because there has been no satisfactory typing scheme for Pseudomonas pseudomallei, recrudescence could not be distinguished from reinfection. We determined the strain identity of primary and relapse isolates of P. pseudomallei from 25 patients with culture-proven melioidosis to answer whether secondary infections were due to the initial infecting strain or to the acquisition of a new strain. Fifty-four isolates were compared by the patterns of BamHI restriction digests produced after hybridization with a cDNA copy of Escherichia coli rRNA. Twenty-three patients had primary and relapse isolates with identical or highly similar ribotype patterns. The patterns of isolates from two patients were different; the primary and relapse isolates differed by a single fragment for one, and the other had identical primary and first-relapse isolates while the second-relapse isolate was markedly different. The results indicated that recurrent infection probably resulted from endogenous relapse in most of the melioidosis patients studied, although reinfection from an exogenous source was also possible in two cases.

Bacterial Typing Techniques↗
PubMed1993

Latex agglutination test for identification of Pseudomonas pseudomallei.

A latex agglutination test was developed and evaluated for the rapid presumptive identification of Pseudomonas pseudomallei, the causative organism of melioidosis. The test was 100% sensitive for 52 isolates of Ps pseudomallei and 100% specific when tested with other medically important Pseudomonas species and Enterobacteriaceae. A subsequent field trial, with clinical specimens from patients with suspected melioidosis, confirmed the sensitivity and specificity of the test.

Burkholderia pseudomallei↗
PubMed1993

Adult cystic fibrosis: association of acute pulmonary exacerbations and increasing severity of lung disease with auxotrophic mutants of Pseudomonas aeruginosa.

BACKGROUND: Pseudomonas aeruginosa has been located in the endobronchiolar spaces of patients with cystic fibrosis where nutrients may be limited. In these sites it is thought that adaptation of the pathogen might occur and growth factors, present in relative excess, may thus promote survival of the organism. Auxotrophy of pulmonary isolates of P aeruginosa has previously been shown to be a feature of cystic fibrosis and chronic lung sepsis; auxotrophic isolates have additional nutritional requirements to the prototrophic "wild types" of the species. A study was therefore carried out to determine whether the proportion of auxotrophs differs between stable and acutely ill patients, or correlates with the extent of underlying disease. METHODS: Sputum samples were cultured for P aeruginosa and tested for auxotrophy by spreading serial dilutions of homogenised sputum on to a minimal medium which supports only prototrophs, and a complete medium which supports both nutritional types. The proportion of auxotrophs to prototrophs was determined and growth factors of confirmed auxotrophs were identified. RESULTS: Thirty two (86%) of 37 adults with cystic fibrosis infected with P aeruginosa harboured auxotrophs; methionine dependent mutants were isolated from seven of 16 patients tested (44%). More than 50% of the total number of colonies were auxotrophic in 19 of 26 samples (73%) from patients with acute exacerbations and in only six of 15 samples (40%) from clinically stable patients. In four patients from whom samples in both the acute and stable states were available, the proportion of auxotrophs fell in the sample taken when stable. Auxotrophs predominated in all samples from 11 of those patients with very severe underlying lung disease, in contrast to 13 of 30 samples from patients with less severe disease. There was no association between the percentage of auxotrophs and the presence of other respiratory pathogens. CONCLUSIONS: The majority of adults with cystic fibrosis infected with P aeruginosa harbour auxotrophs in the sputum. A significant proportion of acutely ill patients and those with severe underlying disease have a preponderance of auxotrophs in the sputum compared with stable patients and those with less severe disease.

Acute Disease↗
PubMed1992

An outbreak of melioidosis in imported primates in Britain.

An outbreak of melioidosis, a bacterial infection caused by Pseudomonas pseudomallei, was identified in a batch of feral cynomolgus monkeys (Macaca fascicularis) imported to Britain from the Philippines. Thirteen confirmed or possible cases occurred among a batch of 50 animals. Subsequent investigations revealed that the infection was uncommon among imported primates from a variety of sources, although three other cases were identified in monkeys imported from Indonesia. The majority of the affected monkeys had splenic abscesses, and hepatic abscesses and infections of the soft tissues and skin were also frequently observed. Most of the infected animals had no clinical signs despite extensive abscesses, and the presence of infection was only suspected when they were shown to have serum antibodies to P pseudomallei by an enzyme-linked immunosorbent assay. Although there was no evidence of cross infection of other animals or human handlers, this outbreak is a reminder of the dangers of working with wild-caught primates and the potential for the establishment of environmental foci of melioidosis.

Abscess↗
PubMed1992

Auxotrophy of Pseudomonas aeruginosa in cystic fibrosis.

Seventy-four of 403 (18.4%) sputum isolates of Pseudomonas aeruginosa from 49 of 136 (36.0%) adults with cystic fibrosis (CF) were auxotrophic mutants. Two of 11 (18.2%) isolates of P. aeruginosa taken from patients with non-CF bronchiectasis were also auxotrophic. All 99 strains taken from non-bronchiectatic sources were prototrophic. Forty-six of 55 (83.6%) CF auxotrophs required one or more of 36 growth factors tested; the requirements for the remaining 9 isolates were not identified. Methionine was the sole factor required by 17 of 22 (77.3%) isolated which depended on a single factor. We conclude that auxotrophy is a feature of P. aeruginosa infection in cystic fibrosis.

Amino Acids↗
PubMed1992

Diversity of aminoglycoside resistance in Enterobacter cloacae in Greece.

Ninety Enterobacter cloacae strains isolated from 12 Greek hospitals were examined in terms of epidemiological types and resistance mechanisms. Using O serotyping 69% of the strains were assigned to a specific serotype and overall 16 different serotypes were identified. The combination of serotyping, phagetyping and biotyping efficiently discriminated most of the strains, indicating that single epidemic strains were not prevalent, although serotypes 3, 7, and group II predominated. Eight representative strains, all resistant to gentamicin, tobramycin, amikacin and netilmicin, were further examined for transferability and mechanisms of resistance. Aminoglycoside resistance was found to be transferable in most strains, and 13 R plasmids of 40-120 MDa molecular weight were detected. The enzymes detected consisted of three enzymes active against gentamicin [ANT(2h'), AAC(3)-I and AAC(3)-V]; three active against tobramycin [ANT(2"), AAC(3)-V and AAC(6')-I]; two active against netilmicin [AAC(3)-V and AAC(6')-I]; and one active against amikacin [AAC(6')-I]. APH(3') and ANT (3"), which modify neomycin and streptomycin plus spectinomycin respectively, were also found. Overall up to five aminoglycoside modifying enzymes were detected on the same R plasmid, AAC(6')-I plus ANT(2") being the most prevalent. The high incidence of multiresistance in Enterobacter cloacae and the fact that resistance is due to enzymatic inactivation of the antibiotics, indicate that in Greece this species might act as a gene pool for the spread of resistance to other bacteria of clinical relevance.

Aminoglycosides↗
PubMed1992

Homogeneity of lipopolysaccharide antigens in Pseudomonas pseudomallei.

An antiserum raised against a single strain of Pseudomonas pseudomallei reacted equally in a whole cell agglutination test, an indirect haemagglutination (IHA) test and an ELISA with a panel of 12 strains of the species which had been isolated from human beings and animals in various parts of the Far East and Australia between 1923 and 1990. Absorption of the serum with either of two strains removed all reactivity of the serum with other strains. Phenol-water extracted lipopolysaccharide (LPS) from a single strain blocked the reactivity of the serum with red cells sensitised with crude extracts of any of the panel of strains, thereby suggesting that the 'common' antigen was LPS. This antigen was not detected in other Pseudomonas species with the exception of Pseudomonas mallei. Protease K-digested extracts of the 12 strains gave highly similar silver-stained LPS banding patterns in gel electrophoresis. Furthermore, immunoblots of LPS with either rabbit or a patient's serum showed identical ladder profiles for each strain. The results suggest that the LPS antigen is highly conserved throughout P. pseudomallei and that this antigen is detected by the IHA test.

Agglutination Tests↗
PubMed1992

Pseudomonas cepacia pulmonary infection in adults with cystic fibrosis: is nosocomial acquisition occurring?

Ribotyping of 25 isolates of Pseudomonas cepacia taken from the sputum of 21 adults with cystic fibrosis (CF) who were registered at the Royal Brompton Hospital between 1987 and 1991, revealed that seven patients (33.3%) shared strains of a similar ribotype pattern with others, while 14 patients (63.7%) harboured strains unique to each individual. Constancy of sputum strain carriage was seen in two of three patients sampled twice over a 3-month period. Although no evidence for patient-to-patient transmission of P. cepacia within this group of patients was found, the fact that one third of CF patients shared strains of the same ribotype with others, suggests that nosocomial acquisition of this organism may have occurred.

Adolescent↗
PubMed1992

Prevalence of a plasmid-mediated type II dihydrofolate reductase gene among trimethoprim-resistant urinary pathogens in Greek hospitals.

The genetic basis of trimethoprim resistance was examined in 24 Klebsiella pneumoniae, 27 Enterobacter cloacae, five Enterobacter aerogenes and nine Serratia marcescens urinary isolates from five hospitals in Greece. Analysis of the 65 isolates by serotyping and phage-typing identified 53 distinct strains. Thirty-eight isolates (15 K. pneumoniae, 19 E. cloacae, two E. aerogenes and two S. marcescens) hybridized with a probe specific for a gene encoding type II dihydrofolate reductase (DHFR). Three of the K. pneumoniae and four of the E. cloacae isolates which reacted with this probe also hybridized with probes specific for type I DHFR and transposon Tn7. Two E. cloacae isolates hybridized only with the probe for type I DHFR, while a further three isolates hybridized only with the type I DHFR and Tn7 probes. None of the isolates hybridized with a probe for type V DHFR. The plasmids in transconjugants derived from 40 isolates were analysed by digestion with restriction enzymes and Southern blotting. Eighteen (45%) of the donors (12 K. pneumoniae and 6 E. cloacae) produced transconjugants containing plasmids of about 95 kb in size, while transconjugants from the other donors had plasmids in the range 100-185 kb. Of the 18 transconjugants containing a 95 kb plasmid, 15 had similar restriction endonuclease digest patterns, although they varied in terms of the range of antimicrobial resistances which they encoded. When EcoRI digests of these 15 plasmids were hybridized with the type II DHFR probe, a 23 kb common band reacted with the probe.(ABSTRACT TRUNCATED AT 250 WORDS)

Cross Infection↗
PubMed1992

2-Heptyl-4-hydroxyquinoline N-oxide, an antistaphylococcal agent produced by Pseudomonas aeruginosa.

Clinical isolates of Pseudomonas aeruginosa produce low molecular weight, hydrophobic substances which inhibit the growth of Staphylococcus aureus and other Gram-positive organisms. The active substances have been purified from a culture of P. aeruginosa and characterized as a mixture of 2-heptyl-4-hydroxyquinoline N-oxide and its homologues. The alkyl-hydroxyquinolines (derived through reduction of the N-oxide) were also produced by P. aeruginosa cultures but, in general, were less active against S. aureus. 2-Heptyl-4-hydroxyquinoline was identified in a sample of human bronchial secretions from a patient with cystic fibrosis who was heavily colonized with P. aeruginosa. Production of antibacterial alkyl-hydroxyquinoline N-oxides in human lung may explain the observation that the presence of P. aeruginosa in the sputum of patients with cystic fibrosis is correlated with the absence of S. aureus.

Chromatography, High Pressure Liquid↗
PubMed1992

Extrapulmonary sites of Pseudomonas aeruginosa in adults with cystic fibrosis.

BACKGROUND: Pseudomonas aeruginosa infection is seldom eradicated in patients with cystic fibrosis despite intensive antipseudomonal treatment. Upper airway sites of infection may contribute to perpetuation of lower airways infection. This study was designed to find out which extrapulmonary sites are infected and whether the strains at these sites are identical to those in the lungs. METHODS: Sputum and upper airway samples from 42 patients were cultured for P aeruginosa and stool samples from 20 patients were also tested. Nineteen isolates from sputum and extrapulmonary sites from four patients were genotyped with the pCM tox probe. RESULTS: P aeruginosa was isolated from the sputum of 36 patients, 34 of whom had infection in the upper airways. Six of the 20 patients tested were positive for P aeruginosa in the stool. The nasopharynx was colonised in 30 patients, the oropharynx in 29, the middle meatus in 13, the external nares in six, and the inferior turbinate in four. Three of four patients tested had the same strain of P aeruginosa (a different one in each individual) in the sputum and the upper airways, and in two of the three the stool isolate was a different strain. CONCLUSION: Most adults with cystic fibrosis and P aeruginosa pulmonary infection have upper airway reservoirs of the organism and strains from these sites are identical to those in the lungs.

Adolescent↗
PubMed1991

Serological relationships of the O antigens of Klebsiella pneumoniae O5, Escherichia coli O8 and a new O serotype of Serratia marcescens.

Klebsiella pneumoniae O5, Escherichia coli O8 and Serratia marcescens 3255 were shown to cross-react in both ELISA and immunoblotting. The cross-reaction appeared to be due to the O antigen of their lipopolysaccharide (LPS). In addition, there was evidence that the reactions of these strains with their homologous antisera were due, in part, to determinants other than O polysaccharide.

Antigens, Bacterial↗
PubMed1991

Human monoclonal antibodies specific for capsular polysaccharides of Klebsiella recognize clusters of multiple serotypes.

We report the generation and the characterization of a set of human monoclonal antibodies (HmAb) specific for Gram-negative bacteria of Klebsiella pneumoniae. The eight human hybridomas secrete either IgM kappa, IgA1 kappa, or IgA2 kappa antibodies. One HmAb binds bacteria of only one serotype. Five HmAb recognize non-overlapping clusters of 2, 3, or 10 different serotypes. The remaining two HmAb both recognize three serotypes. Two serotypes are recognized by both HmAb, and in addition both HmAb bind one more nonidentical serotype. These results suggest that in man, epitopes are immunodominant, different from serotype-specific determinants detected by conventional rabbit antisera. Screening of clinical isolates revealed that the HmAb recognize not only representative typing strains but also most isolates of the corresponding serotype. In addition, most of the isolates that were non-typable by polyclonal antisera were recognized by one of the HmAb. Fine specificity analyses revealed that all HmAb are highly specific for the isolated capsular polysaccharides (CPS) of bacteria within the corresponding cluster of serotypes. However, the avidity of a HmAb for the different CPS can differ significantly. Taken together, our results suggest that the unequivocal interactions between HmAb and CPS offer the basis for an alternative, better defined classification system, and that passive immunization with a limited number of HmAb may provide a feasible strategy for the protection against the majority of fatal, nosocomial infections with multidrug-resistant strains of K. pneumoniae.

Antibodies, Bacterial↗
PubMed1991

Interaction between Pseudomonas aeruginosa and Staphylococcus aureus: description of an anti-staphylococcal substance.

The presence of Pseudomonas aeruginosa in the sputum of 191 patients with cystic fibrosis was significantly related (p less than 0.0001) to the absence of Staphylococcus aureus. Cross-streaking tests showed that 40 of 50 clinical strains of P. aeruginosa produced substances that inhibited the growth of S. aureus. When incorporated into agar plates, this antibacterial substance(s) inhibited the growth of 177 of 189 strains of nine staphylococcal species, all of 16 methicillin-resistant S. aureus and 27 of 39 strains of six other gram-positive genera. The substance(s) did not inhibit 23 strains of seven gram-negative genera tested. The antibacterial activity was heat stable and could be extracted into chloroform; activity was retained on Sephadex G-15 (V/Vo approximately 2, Mr less than 500) and eluted as a single peak from high performance liquid chromatography, well separated from pseudomonic acid, pyocyanin and a number of other phenazines.

Anti-Bacterial Agents↗
PubMed1991

Comparison of ribotyping with conventional methods for the type identification of Enterobacter cloacae.

An Escherichia coli rRNA probe was compared with a combination of O serotyping, phage susceptibility, and biotype pattern for the type identification of strains of Enterobacter cloacae. Forty-five isolates of E. cloacae from 36 patients in nine hospitals were examined. By conventional typing, only 26 (57.7%) could be assigned to a specific serotype and 6 (13.3%) were autoagglutinating owing to rough lipopolysaccharide antigens. All isolates could be assigned to one of three biotypes, but many phage sensitivity patterns were evident. Twenty-nine distinct strains were identified by combined typing. Probing of EcoRI and BamHI digests of chromosomal DNA with a cDNA copy of E. coli rRNA proved to be highly discriminating between strains. Thirty different ribotypes based on 28 bands were recorded. Overall, agreement between the ribotyping and combined typing methods was good (84.4%), and discrepancies were generally confined to serologically unclassifiable strains and variability in biotype codes. Ribotyping was reproducible, and five of six pairs of isolates from the same and different patients gave identical hybridization profiles on separate occasions. We conclude that ribotyping is a highly discriminatory and reproducible method for the typing of E. cloacae, but in most outbreaks it offers little increase in discrimination over traditional methods.

Bacterial Typing Techniques↗