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Biomedical subjects

B Watt

Publications and source records attributed to B Watt.

At least 19 recordsLinked to original sources
PubMed2002

Molecular epidemiology of Mycobacterium malmoense infections in Scotland.

Clinical isolates of Mycobacterium malmoense collected over 5 years from patients across Scotland with a variety of diseases have been characterized by pulsed-field gel electrophoresis (PFGE), ribotyping, and 16S ribosomal DNA gene sequencing. Results indicate that this species harbors little genetic diversity and that the different strain types that were identified by PFGE showed no correlation with geographical origin or date of isolation.

DNA, Bacterial↗
PubMed2002

The efficacy of the heat killing of Mycobacterium tuberculosis.

There is concern that current procedures for the heat inactivation of Mycobacterium tuberculosis may not be adequate. This raises serious safety issues for laboratory staff performing molecular investigations such as IS6110 restriction fragment length polymorphism typing. This paper confirms that the protocol of van Embden et al, as performed routinely in this laboratory, is safe and effective for the heat inactivation of M tuberculosis. This procedure involves complete immersion of a tube containing a suspension of one loopfull of growth in a water bath at 80 degrees C for 20 minutes. Seventy four isolates were included in this investigation. Despite prolonged incubation for 20 weeks, none of the heat killed M tuberculosis suspensions produced visible colonies or gave a positive growth signal from liquid culture. This method did not affect the integrity of the DNA for subsequent molecular investigations.

Bacteriological Techniques↗
PubMed2002

Antibiotic resistant tuberculosis in the United Kingdom: 1993-1999.

BACKGROUND: The re-emergence of tuberculosis as a global health problem over the past two decades, accompanied by an increase in tuberculosis drug resistance, prompted the development of a comprehensive national surveillance system for tuberculosis drug resistance in 1993. METHODS: The UK Mycobacterial Resistance Network (Mycobnet), which includes all mycobacterial reference and regional laboratories in the UK, collects a minimum dataset on all individuals from whom an initial isolate of Mycobacterium tuberculosis complex has been isolated and submitted by source hospital laboratories. Data sought include susceptibility to first line antibiotics, demographic, geographical, and risk factor information. RESULTS: There were 25 217 reports of initial isolates of M tuberculosis complex in the UK between 1993 and 1999. All were tested for sensitivity to isoniazid, rifampicin, and ethambutol and 12 692 of the isolates were also tested for sensitivity to pyrazinamide and streptomycin. A total of 1523 (6.1%) isolates were resistant to one or more drugs, 1397 isolates (5.6%) were resistant to isoniazid with or without resistance to other drugs, and 299 (1.2%) were multidrug resistant. Although the numbers of drug resistant isolates increased over the period, the proportions remained little changed. Certain groups of people were at a higher risk of acquiring drug resistant tuberculosis including younger men, residents of London, foreign born subjects, patients with a previous history of tuberculosis and those infected with HIV. CONCLUSION: Although the proportion of drug resistant tuberculosis cases appears to be stable in the UK at present, more than one in 20 patients has drug resistant disease at diagnosis and more than one in 100 has multidrug resistant disease. Tuberculosis control measures should be strengthened to minimise the emergence of drug resistance through rapid diagnosis, rapid identification of drug resistance, supervised treatment, and maintenance of comprehensive surveillance.

AIDS-Related Opportunistic Infections↗
PubMed2002

A national study of clinical and laboratory factors affecting the survival of patients with multiple drug resistant tuberculosis in the UK.

BACKGROUND: This study aimed to describe the clinical, microbiological, molecular epidemiology and treatment of multidrug resistant tuberculosis (MDRTB) cases in the UK and to determine factors associated with survival. METHODS: Ninety MDRTB cases were identified from 1 January 1996 to 30 June 1997; 69 were DNA fingerprinted. Date of diagnosis was determined and data were collated on key demographic factors, clinical, radiological and treatment details. Variables associated with survival were included in a Cox proportional hazards model. RESULTS: Most of the patients (72.4%) were male, born outside the UK (57.1%), were sputum smear positive (82.2%), and had entered the UK more than 5 years previously (61.9%). Thirty eight of 78 cases (48.7%) had prior TB. Sufficient data on 82 patients were available for survival analysis; 20/27 (74.1%) known to be dead at the end of the observation period had died of tuberculosis. Median survival time overall was 1379 days (95% CI 1336 to 2515) or 3.78 (95% CI 3.66 to 6.89) years (858 days (95% CI 530 to 2515) in immunocompromised individuals (n=32) and 1554 (95% CI 1336 to 2066) days in immunocompetent cases (n=48)). Median survival in patients treated with three drugs to which the bacterium was susceptible on in vitro testing (n=62) was 2066 days (95% CI 1336 to 2515) or 5.66 years, whereas in those not so treated (n=13) survival was 599 days (95% CI 190 to 969) or 1.64 years. CONCLUSIONS: Immunocompromised status, failure to culture the bacterium in 30 days or to apply appropriate three drug treatment, and age were significant factors in mortality. An immunocompromised patient was nearly nine times more likely to die, while application of appropriate treatment reduced the risk (risk ratio 0.06). Increasing age was associated with increasing risk of death (risk ratio 2.079; 95% CI 1.269 to 3.402)-that is, for every 10 year increase in age the risk almost doubled. Overall survival was lower than that reported in previous studies.

Adolescent↗
PubMed2001

Molecular evidence for independent occurrence of IS6110 insertions at the same sites of the genome of Mycobacterium tuberculosis in different clinical isolates.

Several characteristics of Mycobacterium tuberculosis (e.g., conserved genome and low growth rate) have severely restricted the study of the microorganism. The discovery of IS6110 raised hopes of overcoming these obstacles. However, our knowledge of this IS element is relatively limited; even its two basic characteristics (transposition mechanism and target site selection) are far from well understood. In this study, IS6110 insertions in ipl loci (iplA and iplB) in two collections of clinical isolates of M. tuberculosis from different geographic locations, one from Scotland and the other from Thailand, were investigated. Five different IS6110 insertions in the loci were identified: ipl-4::IS6110, ipl-5::IS6110, ipl-11::IS6110, ipl-12::IS6110, and ipl-13::IS6110. An attempt to establish the phylogenetic relationship of the isolates containing these insertions was unsuccessful, suggesting that some of these insertions may have arisen from more than one event. This possibility is further supported by the observation that IS6110 copies existed in the same site but with different orientations in different isolates, and the insertion site of ipl-1::IS6110 harbored IS6110 copies in both iplA and iplB in different strains. All these suggest the independent occurrence of IS6110 insertions at the same sites of the genome of M. tuberculosis in different clinical isolates. The implications of this finding are discussed.

Base Sequence↗
PubMed2000

Comparison of three isolation systems for the culture of mycobacteria from respiratory and non-respiratory samples.

AIMS: To compare the recovery of mycobacteria from clinical samples using the MB/BacT rapid culture system with that obtained using egg medium or the Bactec radiometric method. METHODS: The three methods were compared using 681 clinical samples (462 respiratory and 219 non-respiratory samples) and eight external quality control strains. Culture media were incubated at 35-37 degrees C for six weeks in the MB/BacT system and for 12 weeks in the Bactec system and on egg medium. Solid media were examined macroscopically once a week and the Bactec vials were read six times in the first two weeks, and then weekly for the next 10 weeks (a growth index > 50 indicated a positive vial). The MB/BacT system positive vials were unloaded from the machine as soon as possible after detection. Confirmation of growth for all systems was by Ziehl-Neelson stained smears. Isolates were identified by a combination of phenotypic and molecular methods. RESULTS: Of the 681 clinical samples, 59 (8.7%) were positive on culture, including 23 strains of Mycobacterium tuberculosis. None of the three systems recovered all of the isolates, but each recovered mycobacteria not detected by either of the other two systems. After six weeks incubation, isolation rates were 87%, 78%, and 90%, and mean times to detection were 13, 19, and nine days for the MB/BacT, egg medium, and Bactec systems, respectively. Although the MB/BacT system was slightly slower than the Bactec system, the biomass was greater, allowing earlier use of molecular probes and earlier inoculation of susceptibility tests. CONCLUSIONS: The MB/BacT system provides comparable performance to the Bactec radiometric system, without the problems of disposal of radioactive waste. Optimal recovery is obtained when culture on egg medium is used in conjunction with a rapid culture system.

Bacteriological Techniques↗
PubMed1999

IS6110-mediated deletions of wild-type chromosomes of Mycobacterium tuberculosis.

The ipl locus is a site for the preferential insertion of IS6110 and has been identified as an insertion sequence, IS1547, in its own right. Various deletions around the ipl locus of clinical isolates of Mycobacterium tuberculosis were identified, and these deletions ranged in length from several hundred base pairs up to several kilobase pairs. The most obvious feature shared by these deletions was the presence of an IS6110 copy at the deletion sites, which suggested two possible mechanisms for their occurrence, IS6110 transposition and homologous recombination. To clarify the mechanism, an investigation was conducted; the results suggest that although deletion transpositionally mediated by IS6110 was a possibility, homologous recombination was a more likely one. The implications of such chromosomal rearrangements for the evolution of M. tuberculosis, for IS6110-mediated mutagenesis, and for the development of genetic tools are discussed. The deletion of genomic DNA in isolates of M. tuberculosis has previously been noted at only a few sites. This study examined the deletional loss of genetic material at a new site and suggests that such losses may occur elsewhere too and may be more prevalent than was previously thought. Distinct from the study of laboratory-induced mutations, the detailed analysis of clinical isolates, in combination with knowledge of their evolutionary relationships to each other, gives us the opportunity to study mutational diversity in isolates that have survived in the human host and therefore offers a different perspective on the importance of particular genetic markers in pathogenesis.

Bacterial Proteins↗
PubMed1999

Characterization of IS1547, a new member of the IS900 family in the Mycobacterium tuberculosis complex, and its association with IS6110.

Unlike classically defined insertion sequence (IS) elements, which are delimited by their inverted terminal repeats, some IS elements do not have inverted terminal repeats. Among this group of atypical IS elements, IS116, IS900, IS901, and IS1110 have been proposed as members of the IS900 family of elements, not only because they do not have inverted terminal repeats but also because they share other features such as homologous transposases and particular insertion sites. In this study, we report a newly identified IS sequence, IS1547, which was first identified in a clinical isolate of Mycobacterium tuberculosis. Its structure, insertion site, and putative transposase all conform with the conventions of the IS900 family, suggesting that it is a new member of this family. IS1547 was detected only in isolates of the M. tuberculosis complex, where it had highly polymorphic restriction fragment length polymorphism patterns, suggesting that it may be a useful genetic marker for identifying isolates of the M. tuberculosis complex and for distinguishing different strains of M. tuberculosis. ipl is a preferential locus for IS6110 insertion where there are eight known different insertion sites for IS6110. Surprisingly, the DNA sequence of ipl is now known to be a part of IS1547, meaning that IS1547 is a preferential site for IS6110 insertion.

Amino Acid Sequence↗
PubMed1999

Molecular evidence for heterogeneity of the multiple-drug-resistant Mycobacterium tuberculosis population in Scotland (1990 to 1997).

Multiple-drug-resistant Mycobacterium tuberculosis (MDR-MTB) has been well studied in hospitals or health care institutions and in human immunodeficiency virus-infected populations. However, the characteristics of MDR-MTB in the community have not been well investigated. An understanding of its prevalence and circulation within the community will help to estimate the problem and optimize the strategies for control and prevention of its development and transmission. In this study, MDR-MTB isolates from Scotland collected between 1990 and 1997 were characterized, along with non-drug-resistant isolates. The results showed that they were genetically diverse, suggesting they were unrelated to each other and had probably evolved independently. Several new alleles of rpoB, katG, and ahpC were identified: rpoB codon 525 (ACC-->AAC; Thr525Asn); katG codon 128 (CGG-->CAG; Arg128Gln) and codon 291 (GCT-->CCT; Ala291Pro); and the ahpC synonymous substitution at codon 6 (ATT-->ATC). One of the MDR-MTB isolates from an Asian patient had an IS6110 restriction fragment length polymorphism pattern very similar to that of the MDR-MTB W strain and had the same drug resistance-related alleles but did not have any epidemiological connection with the W strains. Additionally, a cluster of M. tuberculosis isolates was identified in our collection of 715 clinical isolates; the isolates in this cluster had genetic backgrounds very similar to those of the W strains, one of which had already developed multiple drug resistances. The diverse population of MDR-MTB in Scotland, along with a low incidence of drug-resistant M. tuberculosis, has implications for the control of the organism and prevention of its spread.

Alleles↗
PubMed1999

A national audit of the laboratory diagnosis of tuberculosis and other mycobacterial diseases within the United Kingdom.

In order to audit United Kingdom laboratory diagnostic and reference services including novel molecular methods for tuberculosis, a questionnaire was sent to laboratories submitting specimens to the PHLS Mycobacterium Reference Unit (MRU) and regional centres and to the Scottish Mycobacteria Reference Laboratory (SMRL) in 1996-7. Nationally, 67.2% of laboratories responded. Most UK laboratories were fully or conditionally CPA accredited and take part in the NEQAS proficiency scheme. On average only 3.3% of primary samples submitted for mycobacterial diagnosis in 1995 produced a mycobacterial culture from approximately half as many patients (that is, a mean of 1488 specimens producing 49 isolates from 23 patients). Potentially over 380,000 specimens are processed for mycobacteria in the UK each year. The majority of laboratories use 4% NaOH +/- NALC for specimen decontamination. Culture on solid media was used by most laboratories and 62.9% also use liquid media. Most laboratories incubated cultures for eight weeks. Few laboratories use molecular diagnostic methods. Laboratories were most likely to use molecular methods for diagnosing tuberculous meningitis and for specimens from immunocompromised patients, although usage was strongly influenced by cost. Within England and Wales 43.9% (47/107) and 56% (61/109) of laboratories wanted a rapid service for rifampicin resistance detection in M tuberculosis from immunocompetent and immunocompromised patients, respectively. In regard to a tuberculous meningitis service, 80.5% (43/112) and 84.3% (102/121) of laboratories wanted this service for immunocompetent and immunocompromised patients, respectively. The quality of reference services was rated as "very good"/"good" by 85.6% of respondents nationally. Rapid molecular amplification diagnostic services were established at the PHLS MRU for rifampicin drug resistance detection nationally and for tuberculous meningitis at the MRU.

Bacteriological Techniques↗
PubMed1998

The management of pulmonary tuberculosis in adults notified in Scotland in 1993.

The management of pulmonary tuberculosis (TB) in Scotland in 1993 was studied by asking the physicians responsible for all 321 adult cases of the disease notified that year to complete a standardized questionnaire relating to drug treatment and bacteriology. The response rate to the questionnaire was 100%. Isoniazid and rifampicin were used together in initial therapy in 98.4% of cases, while pyrazinamide was prescribed in 90.3% of cases, broadly in keeping with existing treatment guidelines. However, considerable variability was observed both in the drug regimens employed, and in the duration of initial and continuation phases of chemotherapy. Treatment regimens were therefore frequently at variance with published recommendations. Among patients prescribed drug regimens other than those recommended satisfactory completion of therapy was less common. Microbiological confirmation was provided for 84% of cases in which clinical samples were submitted. However, in approximately 11% of cases, no clinical samples were submitted. Closer adherence to existing treatment guidelines and more rigorous pursuit of microbiological confirmation should further improve the overall management of pulmonary TB in Scotland.

Adolescent↗
PubMed1998

IS6110 transposition and evolutionary scenario of the direct repeat locus in a group of closely related Mycobacterium tuberculosis strains.

In recent years, various polymorphic loci and multicopy insertion elements have been discovered in the Mycobacterium tuberculosis genome, such as the direct repeat (DR) locus, the major polymorphic tandem repeats, the polymorphic GC-rich repetitive sequence, IS6110, and IS1081. These, especially IS6110 and the DR locus, have been widely used as genetic markers to differentiate M. tuberculosis isolates and will continue to be so used, due to the conserved nature of the genome of M. tuberculosis. However, little is known about the processes involved in generating these or of their relative rates of change. Without an understanding of the biological characteristics of these genetic markers, it is difficult to use them to their full extent for understanding the population genetics and epidemiology of M. tuberculosis. To address these points, we identified a cluster of 7 isolates in a collection of 101 clinical isolates and investigated them with various polymorphic genetic markers, which indicated that they were highly related to each other. This cluster provided a model system for the study of IS6110 transposition, evolution at the DR locus, and the effects of these on the determination of evolutionary relationships among M. tuberculosis strains. Our results suggest that IS6110 restriction fragment length polymorphism patterns are useful in grouping closely related isolates together; however, they can be misleading if used for making inferences about the evolutionary relationships between closely related isolates. DNA sequence analysis of the DR loci of these isolates revealed an evolutionary scenario, which, complemented with the information from IS6110, allowed a reconstruction of the evolutionary steps and relationships among these closely related isolates. Loss of the IS6110 copy in the DR locus was noted, and the mechanisms of this loss are discussed.

Base Sequence↗
PubMed1998

Palliative care at home: an audit of cancer deaths in Grampian region.

BACKGROUND: Ninety per cent of the last year of life of cancer patients is spent at home. Some studies have suggested that care in this setting is often suboptimal. Information on the standard of palliative care delivered at home by general practitioners (GPs) and their teams is limited, and clarification of the problems faced is needed. AIM: To audit the home-based palliative care of patients dying of cancer. METHOD: Matched postal questionnaires were sent to the GPs and nurses of 1086 successive patients dying of cancer in whatever setting in the Grampian region of Scotland some six weeks after the death to establish the professionals' perception of symptom control, communication problems, use of services, and information given to patients and relatives. RESULTS: Response rates were 88.8% for GPs (964 out of 1086) and 87.1% for nurses (325 out of 375 that were passed on to nurses). Two-thirds of patients received palliative care at home. Pain was poorly controlled in 15.7%, and poor control of other symptoms ranged from 13.8% (nausea and vomiting) to 21% (depression and dyspnoea). Communication difficulties were present in 93.7% of cases, although only 5.2% of these were of a major nature. District nurses were involved in 76.7% of cases and Macmillan nurses in 28.0%. Twenty-six per cent of referrals to district nurses were assessed as being late in the course of the illness. Patients were fully informed about the diagnosis in 66.3% of cases and about the prognosis in 55.4%. General practitioners were more likely to report the presence of communication problems between themselves and the patient (when compared with nurses: 43.9% versus 28.0%), more likely to report that patients were 'not at all informed' about self-help groups (57.5% versus 36.3%), and were less likely to report the involvement of occupational therapists (21.8% versus 39.7%). CONCLUSIONS: Levels of reporting of poor symptom control by professionals was much lower than levels reported by relatives in other studies, but there was no difference between the reporting of GPs and nurses. However, a number of areas were identified where care could be enhanced by improved teamwork and further education and training in symptom control, as well as in communication, use of services, and information provision.

Communication↗