Performance comparison between the mycobacteria growth indicator tube system and L?wenstein-Jensen medium in the routine detection of Mycobacterium tuberculosis at public health care facilities in Rio de Janeiro, Brazil: preliminary results of a pragmatic clinical trial

In view of the fact that the World Health Organization has recommended the use of the mycobacteria growth indicator tube (MGIT) 960 system for the diagnosis of tuberculosis and that there is as yet no evidence regarding the clinical impact of its use in health care systems, we conducted a pragmatic clinical trial to evaluate the clinical performance and cost-effectiveness of the use of MGIT 960 at two health care facilities in the city of Rio de Janeiro, Brazil, where the incidence of tuberculosis is high. Here, we summarize the methodology and preliminary results of the trial. (ISRCTN.org Identifier: ISRCTN79888843 [http://isrctn.org/])     

Mycobacterium tuberculosis typing: usefulness of DRE-PCR to confirm cross-contamination in the mycobacteriology laboratory of a general reference hospital for AIDS.

In this study two molecular typing methods, a simple double repetitive element PCR-based assay and the standardized restriction fragment length polymorphism (RFLP), were used to confirm cross-contamination in the mycobacteriology laboratory. Clinical specimens from 12 patients, submitted for acid-fast bacilli stain smear and processed for culture in Lowenstein-Jensen on the same day, resulted in positive bacterioscopy (+++) and confluent growth only for one of the patients. The specimens from all the other patients but two were smear-negative and culture-positive, with one or two colonies. None of them had clinical symptoms and radiological findings for active tuberculosis (TB). The suspicion of false-positive cultures arose when a health care worker who had had a PPD skin test conversion, claimed to be healthy and had no TB symptoms, was found to have a positive sputum culture. DRE-PCR demonstrated that all nine cultures typed belonged to one cluster, further confirmed by RFLP. Although DRE-PCR has been found to be poorly reproducible, it has enough discriminatory power to be useful for rapid epidemiological investigation in selected settings.     

Spoligotypes of Mycobacterium tuberculosis complex isolates from patients residents of 11 states of Brazil

One of the high tuberculosis (TB) incidence countries in the world, Brazil is characterized by considerable differences in TB incidence on regional and state level. In the present study, we describe Brazilian spoligotypes of 1991 Mycobacterium tuberculosis complex (MTC) clinical isolates from patients residents of 11 states from different regions of the country, diagnosed between 1996 and 2005. By performing spoligotyping on a large number of M. tuberculosis clinical isolates, one of the main objectives of this study was to determine the major genotype families causing TB in Brazil and to verify the region-associated genotype distribution. We observed a total of 577 distinct spoligopatterns, 12.6% of these corresponded to orphan patterns while 87.4% belonged to 326 shared-types (SITs). Among the latter, 86 SITs (isolated from 178 patients) had been observed for the first time in this study, the most frequent being SIT2517 which belonged to the T3-ETH lineage and was exclusively found among patients residents of Belém, the capital of the state of Pará (n=8 isolates). Irrespective of shared-type labeling, a total of 19.5% strains were unique (unclustered) in our study as opposed to 80.5% clustered isolates (189 clusters, size range from 2 to 205 isolates). The three largest clusters were SIT42 of the Latin-America & Mediterranean (LAM) 9 clade (10.3%), SIT53 of the T clade (7.6%), and SIT50 of the Haarlem clade (5.4%). The predominant MTC lineages in Brazil in decreasing order belonged to the LAM (46%); the ill-defined T (18.6%); the Haarlem (12.2%), the X (4.7%), the S (1.9%), and the East African Indian (EAI) (0.85%) families. The rest of clades grouped together as Mycobacterium africanum, Mycobacterium bovis, Beijing, Central Asian (CAS), and the Manu types, represented less than 1% of the strains. Finally, about 15% of the isolates showed spoligotype signatures that were not yet classified among well-defined lineages. In conclusion, we provide hereby a first insight into the population structure of MTC isolates in Brazil, showing the predominance of both LAM and T family and the existence of region-associated genotypes.     

自动BACTEC960系统检测结核分枝杆菌药物敏感性的多中心评估分析

地点：巴西东南部3个分枝杆菌参比实验室。目的：评估自动化分枝杆菌生长指示管（MGIT）检测结核分枝杆菌药物敏感性的能力。设计：用自动BECTECMGIT960（M960）对95份结核分枝杆菌临床分离株进行链霉素（SM），异烟肼（INH）、利福平（RMP）、乙胺丁醇（EMB）的敏感性检测，并与BACTEC460（B460）、比例法（PM）、抗性比率法（RRM）的检测结果进行比较，其中88株以BACTEC460、比例法、抗性比率法中的至少两种方法获得一致结果作为最终结果，并与M960的结果相比较。结果：M960与B460的符合率为95．2％，与PM为96．6%,与RRM为93．4％；其中与RRM在SM上符合率最低，与B460在EMB上符合率最低。M960结果与最终结果的符合率为97．9％，INH与RMP的符合率为99．2%,SM与EMB的符合率分别为96．2%和96．9％，M960报告结果的平均时间为6．9d。结论：与比例法和抗性比率法相比，M960显示出了极大的优势，并且有助于患者的治疗。     

Latent tuberculosis infection among undergraduate medical students in Rio de Janeiro State, Brazil.

SETTING: Five medical schools in three cities with different tuberculosis (TB) incidence rates in Rio de Janeiro State, Brazil. OBJECTIVE: To estimate prevalence of and associated factors for latent tuberculosis infection (LTBI) among medical students. DESIGN: A cross-sectional survey was conducted among undergraduate students in pre-clinical, early and late clinical years from schools in cities with low (28/100,000), intermediate (63/100,000) and high (114/100,000) TB incidence rates. Information on socio-demographic profile, previous BCG vaccination, potential TB exposure, co-morbidity and use of respiratory protective masks was obtained. A tuberculin skin test (TST) was performed using the Mantoux technique by an experienced professional. A positive TST, defined as induration > or = 10 mm, was considered LTBI. RESULTS: LTBI prevalence was 6.9% (95%CI 5.4-8.6). In multivariate analysis, male sex (adjusted odds ratio [aOR] 1.8; 95% CI 1.1-3.0), late clinical years (aOR 1.9; 95% CI 1.01-3.5), intermediate TB incidence (aOR 4.3; 95% CI 1.3-14.6) and high TB incidence in the city of medical school (aOR 5.1; 95% CI 1.6-16.8) were significantly associated with LTBI. CONCLUSIONS: The higher prevalence of LTBI in late clinical years suggests that medical students are at increased risk for nosocomial Mycobacterium tuberculosis infection. The implementation of a TB control program may be necessary in medical schools, particularly in cities with higher TB incidence.     

Yield of sputum induction in the diagnosis of pleural tuberculosis

We prospectively evaluated the diagnostic yield of acid-fast bacilli smear and culture for Mycobacterium tuberculosis using sputum induction (SI) in the workup of patients with suspected pleural tuberculosis (TB) who were unable to produce sputum spontaneously. Of the 113 patients studied, a final diagnosis of pleural TB was made in 84 patients (71 HIV seronegative) and a final diagnosis of another disease in 29 patients. Histopathologic examination of the pleural biopsy tissue had the highest diagnostic yield (78%; 66/84). The bacteriologic yield was 62% (52/84) for the pleural tissue, 12% (10/84) for pleural fluid, and 52% (44/84) for sputum cultures obtained by SI. The yield of SI culture for M. tuberculosis was 55% (35/64) in patients with a normal radiograph (except for the pleural effusion) and 45% (9/20) in those with evidence of parenchymal disease suggestive of pulmonary TB (p = 0.6). The yield of sputum cultures obtained by SI is high in patients suspected of having pleural TB even in those cases with no pulmonary parenchymal abnormalities on the chest radiograph.     

Detection of Mycobacterium tuberculosis by a polymerase chain reaction colorimetric dot-blot assay.

SETTING: A public health laboratory in a tuberculosis-endemic region in Brazil. OBJECTIVE: To evaluate the accuracy of a combined polymerase chain reaction (PCR) colorimetric dot-blot protocol for Mycobacterium tuberculosis detection in clinical samples in a public health laboratory. DESIGN: Eighty clinical samples (13 cerebrospinal fluid, 31 induced sputum, 17 expectorated sputum, eight bronchoalveolar lavage and 11 pleural fluid) were assayed with the developed protocol. The accuracy of polymerase chain reaction (PCR) dot-blot methodology was compared to PCR agarose gel electrophoresis (PCR-AG) using as a gold standard the bacteriological result (culture and biochemical identification) combined with clinical follow-up. One internal region of the IS6110 repetitive element of the M. tuberculosis complex was selected for amplification and the amplified product transferred to nylon membranes to be detected by biotinylated DNA probe. RESULTS: Overall sensitivity and specificity obtained were respectively 90% and 97% for PCR-AG and 95% and 97% for the PCR dot-blot. Among the 56 respiratory specimens, the sensitivity and specificity results for PCR-AG were respectively 88% and 95%, and for PCR dot-blot they were 94% and 95%. Among the 24 non-respiratory specimens the sensitivity and specificity results were respectively 83% and 100% for PCR-AG, and 100% and 100% for the PCR dot-blot protocol. CONCLUSION: The results demonstrated that the PCR dot-blot assay may be helpful in the diagnosis of tuberculosis, and feasible even in resource-poor countries.