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M D Cave

Publications and source records attributed to M D Cave.

At least 37 records · Page 2Linked to original sources
PubMed1999

Evidence from molecular fingerprinting of limited spread of drug-resistant tuberculosis in Texas.

To determine the contribution of recent transmission to spread of drug-resistant tuberculosis in Texas, we performed IS6110-based and pTBN12-based restriction fragment length polymorphism (RFLP) analyses on Mycobacterium tuberculosis isolates. Isolates collected from 201 patients in Texas between 1992 and 1994 were studied. The distribution of cases was strikingly focal. All cases were reported from 35 of the 254 counties in Texas, and 74% (148 of 201) were reported from only 9 counties. One hundred sixty-one (80%) of the patients had M. tuberculosis isolates with unique RFLP patterns, and 41 (20%) patients were in 20 clusters, each comprising 2 to 3 patients. The largest number of cases of drug-resistant tuberculosis were reported in counties bordering Mexico, but the percentage of clustered cases was highest in northeast Texas and in counties that included the cities of Dallas, Fort Worth, and Houston. Compared to nonclustered patients, clustered patients were more likely to be African American and to have been born in the United States. Clustered patients were significantly more likely to be from the same geographic area, and clustered patients from the same geographic area were more likely to have isolates with identical drug susceptibility patterns, suggesting that they were linked by recent transmission. In 11 of 20 clusters, clustered patients were from geographically separate regions, and most isolates did not have identical drug susceptibility patterns, suggesting that tuberculosis was contracted from a common source in the remote past. Based on the low percentage of clustered cases and the small cluster size, we conclude that there is no evidence for the extensive transmission of drug-resistant tuberculosis in Texas.

Adolescent↗
PubMed1999

Quantitative analysis of mRNA as a marker for viability of Mycobacterium tuberculosis.

Numerous assays which use conserved DNA or rRNA sequences as targets for amplification have been described for the diagnosis of tuberculosis. However, these techniques have not been applied successfully to the monitoring of therapeutic efficacy owing to the persistence of amplifiable nucleic acid beyond the point at which smears and cultures become negative. Semiquantitative analysis of rRNA has been used to reduce the time required for antimicrobial susceptibility testing of Mycobacterium tuberculosis, although growth for up to 5 days in the presence of some drugs is still required to discriminate resistant strains. The purpose of the present study was to determine whether quantitative analysis of M. tuberculosis mRNA could be used to assess bacterial viability and to illustrate the application of this technique to rapid determination of drug susceptibility. Levels of mRNA encoding the 85B protein (alpha-antigen), IS6110 DNA, and 16S rRNA were compared in parallel cultures of M. tuberculosis that were treated with either no drug, 0. 2 microg of isoniazid per ml, or 1 microg of rifampin per ml. Exposure of sensitive strains to isoniazid or rifampin for 24 h reduced the levels of 85B mRNA to <4 and <0.01%, respectively, of those present in control cultures without drug. In contrast, the levels of IS6110 DNA and 16S rRNA did not diminish over the same period. Strains which were resistant to either isoniazid or rifampin demonstrated no reduction in 85B mRNA in the presence of the drug to which they were nonresponsive. Quantitative analysis of 85B mRNA offers a potentially useful tool for the rapid determination of M. tuberculosis drug susceptibility and for the monitoring of therapeutic efficacy.

Acyltransferases↗
PubMed1999

Detection of viable Mycobacterium tuberculosis by reverse transcriptase-strand displacement amplification of mRNA.

Numerous assays have been described for the detection of DNA and rRNA sequences that are specific for the Mycobacterium tuberculosis complex. Although beneficial to initial diagnosis, such assays have proven unsuitable for monitoring therapeutic efficacy owing to the persistence of these nucleic acid targets long after conversion of smears and cultures to negative. However, prokaryotic mRNA has a typical half-life of only a few minutes and we have previously shown that the presence of mRNA is a good indicator of bacterial viability. The purpose of the present study was to develop a novel reverse transcriptase-strand displacement amplification system for the detection of M. tuberculosis alpha-antigen (85B protein) mRNA and to demonstrate the use of this assay in assessing chemotherapeutic efficacy in patients with pulmonary tuberculosis. The assay was applied to sequential, noninduced sputum specimens collected from four patients: 10 of 11 samples (91%) collected prior to the start of therapy were positive for alpha-antigen mRNA, compared with 1 of 8 (13%), 2 of 8 (25%), 2 of 8 (25%), and 0 of 8 collected on days 2, 4, 7, and 14 of treatment, respectively. In contrast, 39 of 44 samples (89%) collected on or before day 14 were positive for alpha-antigen DNA. The loss of detectable mRNA corresponded to a rapid drop over the first 4 days of treatment in the number of viable organisms present in each sputum sample, equivalent to a mean fall of 0.43 log10 CFU/ml/day. Analysis of mRNA is a potentially useful method for monitoring therapeutic efficacy and for rapid in vitro determination of drug susceptibility.

Acyltransferases↗
PubMed1999

Foci of tuberculosis transmission in central Los Angeles.

To identify sites of tuberculosis transmission and to determine the contribution of HIV-infected patients to tuberculosis morbidity in an urban area, we prospectively evaluated 249 patients with culture-proven tuberculosis in central Los Angeles. Restriction fragment length polymorphism (RFLP) analysis was performed on Mycobacterium tuberculosis isolates to identify patients infected with the same strain. Using RFLP and clinical and epidemiologic data, we identified the most likely source case and site of transmission for 79 patients. Homelessness, birth in the United States and Native American ethnicity were independent predictors of being a source case, but HIV infection was not. Three homeless shelters were sites of tuberculosis transmission for 55 (70%) of the 79 patients. HIV-infected patients constituted 27% (66/249) of the study population, but only 17% (13/79) of patients were infected by an HIV-infected source case. We conclude that transmission of tuberculosis in central Los Angeles was highly focal, and that the major transmission sites were three homeless shelters. HIV- infected tuberculosis patients did not play a major role in spread of tuberculosis. Tuberculosis control measures targeted at specific homeless shelters can reduce tuberculosis morbidity in urban areas where homelessness is common and the incidence of tuberculosis is high.

AIDS-Related Opportunistic Infections↗
PubMed1999

Measurement of sputum Mycobacterium tuberculosis messenger RNA as a surrogate for response to chemotherapy.

Effective treatment regimens for pulmonary tuberculosis are difficult to assess because of the slow growth rate of Mycobacterium tuberculosis in culture and its protracted clearance from sputum. A rapid method that reflects effective antimicrobial activity would markedly advance evaluation of treatment and promote the assessment of new antituberculosis drugs. Conventional methods measure the progressive reduction of numbers of acid-fast bacilli in the sputum smear and the clearance of organisms in sputum culture. In this study, we measured levels of M. tuberculosis 85B (alpha antigen) messenger RNA (mRNA), 16S ribosomal RNA (rRNA), and IS6110 DNA in patients' sputa to ascertain whether they could serve as potential surrogate markers of response to chemotherapy. Sputum specimens were sequentially collected for up to a year from 19 smear-positive pulmonary tuberculosis patients receiving an optimal drug treatment regimen. Nucleic acids were isolated from these specimens, and two M. tuberculosis molecular targets (mRNA, DNA) were quantified, using the ABI Prism 7700 Sequence Detection System. The Mycobacterium genus-specific 16S rRNA was quantified with a limiting dilution RT-PCR assay. Results show that levels of 85B mRNA declined after initiation of therapy, as did viable M. tuberculosis colony counts, with 90% of patients becoming negative for both markers after 2 mo of treatment. The rapid disappearance of M. tuberculosis mRNA from sputum suggests that it is a good indicator of microbial viability and a useful marker for rapid assessment of response to chemotherapy.

Adult↗
PubMed1999

The home environment and salmonellosis in children.

OBJECTIVE: To explore the role of foods and the home environment in the development of Salmonella infections in infants and children. METHODS: Home investigations were conducted of patients younger than 4 years of age infected with Salmonella. Cultures were obtained from foods, persons residing in the home, animals/pets/insects, and environmental sources. Like serotypes encountered in the index patients and isolates from the home underwent typing with pulsed-field gel electrophoresis. RESULTS: Home inspections were conducted in approximately 66% of eligible homes on the average of 3.4 days after the confirmation of the Salmonella isolate. A total of 526 cultures from 50 homes were obtained from foods (120), household members (73), refrigerators (52), water (47), countertops (46), soil (42), can-openers (36), vacuum cleaners (34), animals/pets/insects (26), and others (50). Isolates with a serotype identical to those in the index patient were found in 16 homes, 3 of which included an isolate of a second serotype, and an isolate of a different serotype was recovered in 3 homes. The pulsed-field gel electrophoresis patterns of the isolates of identical serotypes from the subjects and from their environment were indistinguishable in all but 2 patients. Among isolates of the same serotype encountered in different homes, all patterns were different. The identical serotype was found in multiple locations (4), dirt surrounding front doors (4), household members (3), vacuum cleaner (1), animals/pets/insects (1), and a refrigerator shelf (1). CONCLUSIONS: These data illustrate the importance of the child's environment in the development of salmonellosis. Clinicians should concentrate on educating the parents about the environmental spread of Salmonella. Contaminated foods in the home play a less significant role in the infection of infants and children.

Arkansas↗
PubMed1998

Epidemiology and molecular identification of Salmonella infections in children.

OBJECTIVE: To explore the role of foods and the environment in the development of infections with Salmonella in infants and children. DESIGN: Case-controlled survey and the use of pulsed-field gel electrophoresis to establish DNA fingerprint patterns. SETTING: Ambulatory and hospitalized patients at a children's hospital. PATIENTS OR OTHER PARTICIPANTS: A consecutive sample of children younger than 4 years old who were infected with Salmonella and 3 age-matched controls per patient were to be surveyed. Of the 103 eligible cases of salmonellosis, 90 cases and 264 controls were included in the study. DATA ANALYSIS: Univariate analysis was done using the Mantel-Haenszel chi2 test or the Fisher exact test. The Bonferroni correction was used for multiple comparisons. DNA fingerprints were inspected for identical banding. RESULTS: Results demonstrated similar diets between cases and controls with the exception of more potato or macaroni salad or coleslaw consumption in the control group (P<.001). DNA fingerprints of Salmonella newport and Salmonella typhimurium demonstrated that all cases were due to unique isolates except in 5 instances involving 12 patients. Seven of these patients could be connected geographically. CONCLUSIONS: Most of the cases of salmonellosis in children younger than 4 years are of a sporadic nature and the major source of infection remains unidentified. For patients infected with identical isolates of Salmonella, a common food source could not be incriminated with the methods used. Environmental contamination or other sources of Salmonella are suggested by these epidemiological data.

Analysis of Variance↗
PubMed1998

Dissemination of Mycobacterium tuberculosis across the San Francisco Bay Area.

The propensity of Mycobacterium tuberculosis genotypes to spread across geographic boundaries was investigated by comparing the IS6110 and polymorphic GC-rich sequence patterns of M. tuberculosis isolates from San Francisco and the East Bay, two distinct regions separated by San Francisco Bay. Of 724 isolates from incident tuberculosis patients during 1992 and 1993, only 53 (7.3%) had patterns matching > or = 1 isolates from the other region. In the multivariable analysis of patient risk factors, an AIDS diagnosis (odds ratio [OR], 1.89; 95% confidence interval [CI], 1.00-3.57) and non-Asian race (OR, 3.43; 95% CI, 1.59-7.42) were associated with having an isolate with a matching pattern. Of 375 unique IS6110 patterns among San Francisco isolates, only 9 (2.4%) matched patterns of East Bay isolates. These population-based data suggest that in the San Francisco Bay Area, M. tuberculosis does not rapidly spread across geographic boundaries, and tuberculosis control efforts should focus on transmission within defined areas.

Acquired Immunodeficiency Syndrome↗
PubMed1998

Diversity of DNA fingerprints of Mycobacterium tuberculosis isolates in the United States.

To investigate the diversity of IS6110 fingerprints of Mycobacterium tuberculosis isolates in the United States and to determine if matching IS6110 fingerprints represent recent interstate tuberculosis transmission, we performed restriction fragment length polymorphism analysis of M. tuberculosis isolates from 1,326 patients in three geographically separated states. Seven hundred ninety-five different IS6110 fingerprint patterns were generated, and pattern diversity was similar in each state. Ninety-six percent of the fingerprint patterns were observed in only one state, demonstrating that most IS6110 fingerprint patterns are confined to a single geographic location. Of the IS6110 fingerprint patterns that were shared by isolates from more than one state, most isolates with 1 to 5 IS6110 copies were separable by pTBN12 fingerprinting whereas those with > 15 copies were not. One high-copy-number M. tuberculosis strain had identical IS6110 and pTBN12 fingerprints and included 57 isolates from three states. Epidemiological data demonstrated significant recent transmission of tuberculosis within each city but not among the states. This suggests that identical fingerprints of isolates from geographically separate locations most likely reflect interstate tuberculosis transmission in the past, with subsequent intrastate spread of disease. Further evaluation of M. tuberculosis strains that cause outbreaks in different geographic locations will provide insight into the epidemiological and bacteriological factors that facilitate the spread of tuberculosis.

DNA Fingerprinting↗
PubMed1998

Comparison of the ABI 7700 system (TaqMan) and competitive PCR for quantification of IS6110 DNA in sputum during treatment of tuberculosis.

Mycobacterium tuberculosis can persist in sputum for long periods of time after the initiation of antituberculosis chemotherapy. The purpose of this study was to determine whether quantitative estimates of M. tuberculosis DNA in sputum correlate with the numbers of viable bacilli and thus measure the therapeutic response of patients during treatment. Two methods of M. tuberculosis DNA quantification were examined by using DNA isolated from sputum specimens serially collected during the course of chemotherapy. A competitive PCR assay was compared to an automated system of real-time quantification with the ABI Prism 7700 Sequence Detection System (TaqMan). The ABI 7700 system uses standard PCR in conjunction with a fluorogenic probe in which the intensity of fluorescence is proportional to the amount of target DNA present. The results showed that both PCR systems are reproducible and accurate. The amounts of M. tuberculosis DNA quantified in sputum corresponded well with the numbers of acid-fast bacilli (AFB) counted by microscopy. Before initiation of antituberculosis therapy, measures of AFB, M. tuberculosis DNA, and cultivable bacilli were similar, suggesting that quantification of DNA is a good method for measuring the initial bacillary load. However, the rate of disappearance of both AFB and M. tuberculosis DNA did not correlate with the decline in cultivable bacilli in the specimen; therefore, these tests are not appropriate for monitoring treatment efficacy.

Antitubercular Agents↗
PubMed1998

Polymerase chain reaction to detect Mycobacterium tuberculosis in histologic specimens.

There is a need for rapid and sensitive detection of Mycobacterium tuberculosis in tissue specimens. A polymerase chain reaction (PCR)-based assay for the diagnosis of tuberculosis was evaluated in 60 formalin-fixed tissue specimens, the target for the amplification being a segment of IS6110 in the M. tuberculosis chromosome. Of the 60 formalin-fixed, paraffin-embedded tissue specimens studied, 57 showed granulomatous inflammation and 53 had been cultured for mycobacteria; 10 were positive for M. tuberculosis and three were positive for other mycobacteria. Of 60 samples, 15 showed acid-fast bacilli on special staining. When done comparatively on a positive culture for M. tuberculosis, PCR for M. tuberculosis DNA in 60 tissue samples was 100% sensitive and 93% specific, having a positive predictive value of 76.9% and negative predictive value of 100%. PCR for M. tuberculosis DNA done on tissue samples was positive for 14 of 19 patients who had a clinical diagnosis of tuberculosis, negative for all six patients with nontuberculous mycobacterial infections, and negative for all 33 patients who had a diagnosis of a disease other than mycobacterial infection. When compared with the clinical diagnosis of tuberculosis, PCR for M. tuberculosis DNA in these patients' tissues was 73.6% sensitive and 100% specific, having a positive predictive value of 100% and negative predictive value of 88.6%. These data indicate that PCR amplification is useful for detecting M. tuberculosis DNA in formalin-fixed tissue specimens, and that it can be used to increase diagnostic accuracy in patients who have perplexing diagnostic problems associated with a granulomatous tissue response.

Bacteriological Techniques↗
PubMed1997

Patterns of tuberculosis transmission in Central Los Angeles.

CONTEXT: Recent studies suggest that many tuberculosis cases in urban areas result from recent transmission. Delineation of the epidemiologic links between patients is important to optimize strategies to reduce tuberculosis transmission. OBJECTIVE: To identify epidemiologic links among recently infected urban patients with tuberculosis. DESIGN: Prospective evaluation of patients with tuberculosis. SETTING: Central Los Angeles, Calif. PATIENTS: A total of 162 patients who had culture-proven tuberculosis. INTERVENTIONS: Patients were prospectively interviewed to identify their contacts and whereabouts. The IS6110-based and pTBN12-based restriction fragment length polymorphism analyses were performed on Mycobacterium tuberculosis isolates. Patients whose isolates had identical or closely related restriction fragment length polymorphism patterns were considered a cluster. Unconditional logistic regression was used to identify independent predictors of clustering. MAIN OUTCOME MEASURES: Relationship of clinical and epidemiologic variables to clustering. RESULTS: A total of 96 (59%) of 162 patients were in 8 clusters. Only 2 of the 96 clustered patients named others in the cluster as contacts. The degree of homelessness was an independent predictor of clustering. Compared with nonclustered patients, patients in 6 clusters were significantly more likely to have spent time at 3 shelters and other locations when at least 1 patient in the cluster was contagious, and these locations were independent predictors of clustering. Among nonhomeless persons, clustered patients were significantly more likely than nonclustered patients to have used daytime services at 3 shelters. CONCLUSIONS: (1) Traditional contact investigation does not reliably identify patients infected with the same M tuberculosis strain, and (2) locations at which the homeless congregate are important sites of tuberculosis transmission for homeless and nonhomeless persons. Measures that reduce tuberculosis transmission should be based on locations rather than on personal contacts.

Adolescent↗
PubMed1997

Differences in the prevalence of IS6110 insertion sites in Mycobacterium tuberculosis strains: low and high copy number of IS6110.

SETTING: Mycobacterium tuberculosis (M. tuberculosis) isolates from various parts of the USA which have few copies of the insertion sequence IS6110. OBJECTIVES: To characterize the sites of insertion of IS6110 among M. tuberculosis isolates that have one to six copies of the insertion sequence. DESIGN: The mixed-linker polymerase chain reaction (ML-PCR) procedure was used to amplify the terminal repeats on the ends of IS6110 and adjacent flanking sequences. From the ML-PCR products, sequences flanking 14 copies of IS6110 in strains containing less than seven copies of the insertion were determined. Sequence information from the flanking deoxyribonucleic acid was used to construct flanking primers that can be used to indicate the presence of IS6110 at a particular site when paired with outbound IS6110 primers in a PCR. Over 200 strains of diverse origin were screened for the insertion of IS6110 at several distinct sites using this procedure. RESULTS: The direct repeat (DR) locus has been described as a highly preferred site for insertion of IS6110 in strains of M. tuberculosis. Another highly preferred site of insertion of IS6100, DK1, is herein described. Insertions at DK1 are highly prevalent in M. tuberculosis strains harboring two to six copies of IS6110. The prevalence of insertions at this site decreases in strains with more than six copies of IS6110, even though the sequence itself is present in strains lacking a copy of IS6110 at this site. CONCLUSION: In addition to the DR locus there are other conserved sites of insertion among M. tuberculosis strains. The data further suggest a separate lineage for the high copy and the low copy strains, and a possible sequential insertion of IS6110 in strains of M. tuberculosis with less than seven copies.

Base Sequence↗
PubMed1997

Interpretation of restriction fragment length polymorphism analysis of Mycobacterium tuberculosis isolates from a state with a large rural population.

Epidemiologic relatedness of Mycobacterium tuberculosis isolates from Arkansas residents diagnosed with tuberculosis in 1992-1993 was assessed using IS6110- and pTBN12-based restriction fragment length polymorphism (RFLP) and epidemiologic investigation. Patients with isolates having similar IS6110 patterns had medical records reviewed and were interviewed to identify epidemiologic links. Complete RFLP analyses were obtained for isolates of 235 patients; 78 (33%) matched the pattern of > or = 1 other isolate, forming 24 clusters. Epidemiologic connections were found for 33 (42%) of 78 patients in 11 clusters. Transmission of M. tuberculosis likely occurred many years in the past for 5 patients in 2 clusters. Of clusters based only on IS6110 analyses, those with > or = 6 IS6110 copies had both a significantly greater proportion of isolates that matched by pTBN12 analysis and patients with epidemiologic connections, indicating IS6110 patterns with few bands lack strain specificity. Secondary RFLP analysis increased specificity, but most clustered patients still did not appear to be epidemiologically related. RFLP clustering in rural areas may not represent recent transmission.

Adolescent↗
PubMed1997

Retrospective detection of laboratory cross-contamination of Mycobacterium tuberculosis cultures with use of DNA fingerprint analysis.

In 1992-1993, we investigated possible cross-contamination of Mycobacterium tuberculosis cultures as part of a study of tuberculosis in Arkansas by using DNA fingerprint analysis. Of patients whose isolates were matched, those for whom smears were negative and only one culture was positive were identified from laboratory records. Clinical, laboratory, DNA fingerprint, and epidemiological data were reviewed. Of 259 patients, nine (3.5%) were judged to be due to cross-contamination. None of these patients had a clinical course consistent with tuberculosis. All nine specimens were processed with another isolate with a matching DNA fingerprint, and epidemiological connections were not identified among any of the patients. To avoid erroneous diagnoses and unnecessary therapy and public health investigations, specimens from patients with tuberculosis whose smears are negative and only one culture is positive should be investigated for cross-contamination. An inconsistent clinical course and a DNA fingerprint that matches those of other culture-positive specimens processed concurrently, coupled with the lack of an epidemiological connection, suggest cross-contamination.

Adult↗
PubMed1997

The incidence of false-positive cultures for Mycobacterium tuberculosis.

The frequency of false-positive cultures for Mycobacterium tuberculosis due to cross-contamination has been difficult to determine because of the lack of specific strain markers. Isolates collected prospectively over 5 yr from a municipal health department laboratory underwent DNA fingerprinting using the IS6110 and pTBN12 sequences. We reviewed the clinical and laboratory records of all isolates that had matching DNA fingerprints and were processed within 42 d of each other; 8 isolates were classified as probable or definite false-positives, representing 4.0% (8/199) of the culture-positive patients. A convenience sample of 42 isolates from three other mycobacterial laboratories also underwent DNA fingerprinting, and five (12%) were found to be definite or probable false-positives. Cross-contamination during initial processing of specimens was the most common source of false-positive cultures. The source of cross-contamination for three false-positive cultures was a laboratory proficiency survey specimen containing strain H37Ra. Ten of the 13 patients were misdiagnosed as having tuberculosis, and seven received unnecessary multidrug treatment. Clinicians should be aware of the potential for false-positive cultures for M. tuberculosis, and mycobacteriology laboratories need to carefully review procedures to minimize this occurrence. DNA fingerprinting provides a valuable tool for the study of false-positive cultures.

Bacteriological Techniques↗
PubMed1997

A longitudinal study of transmission of tuberculosis in a large prison population.

The purpose of this study was to determine the extent of transmission of tuberculosis in a large prison population over an 18-mo period. Restriction-fragment-length polymorphism (RFLP) analysis of isolates of Mycobacterium tuberculosis was performed, using the insertion sequence IS6110 and the plasmid pTBN12. Patients infected with strains having the same fingerprint were grouped in clusters. Medical records were reviewed and movement of inmates among prisons was examined for selected patients. Tuberculosis was diagnosed in 216 inmates (case rate = 2,283 per 100,000 per year). Isolates from 210 (97%) patients were fingerprinted, 155 (74%) were grouped in 25 clusters, and 55 (26%) showed a unique fingerprint. Recent infection was inferred in 62% of these patients. Eighty-four percent (161 of 192) of patients tested were human immunodeficiency virus (HIV)-positive, of whom 121 were in clusters and 40 were not (p = 0.74). Patients in clusters were less adherent with tuberculosis treatment than those not in clusters (p < 0.05), and prison transmission of resistant strains was observed. It is crucial that infection control guidelines be fully implemented in the prison setting to prevent tuberculosis transmission.

Adult↗