Evaluation of the 2007 WHO guideline to improve the diagnosis of tuberculosis in ambulatory HIV-positive adults

Background

In 2007 WHO issued a guideline to improve the diagnosis of smear-negative and extrapulmonary tuberculosis (EPTB) in HIV-positive patients. This guideline relies heavily on the acceptance of HIV-testing and availability of chest X-rays.

Methods and Findings

Cohort study of TB suspects in four tuberculosis (TB) clinics in Phnom Penh, Cambodia. We assessed the operational performance of the guideline, the incremental yield of investigations, and the diagnostic accuracy for smear-negative tuberculosis in HIV-positive patients using culture positivity as reference standard. 1,147 (68.9%) of 1,665 TB suspects presented with unknown HIV status, 1,124 (98.0%) agreed to be tested, 79 (7.0%) were HIV-positive. Compliance with the guideline for chest X-rays and sputum culture requests was 97.1% and 98.3% respectively. Only 35 of 79 HIV-positive patients (44.3%) with a chest X-ray suggestive of TB started TB treatment within 10 days. 105 of 442 HIV-positive TB suspects started TB treatment (56.2% smear-negative pulmonary TB (PTB), 28.6% smear-positive PTB, 15.2% EPTB). The median time to TB treatment initiation was 5 days (IQR: 2–13 days), ranging from 2 days (IQR: 1–11.5 days) for EPTB, over 2.5 days (IQR: 1–4 days) for smear-positive PTB to 9 days (IQR: 3–17 days) for smear-negative PTB. Among the 34 smear-negative TB patients with a confirmed diagnosis, the incremental yield of chest X-ray, clinical suspicion or abdominal ultrasound, and culture was 41.2%, 17.6% and 41.2% respectively. The sensitivity and specificity of the algorithm to diagnose smear-negative TB in HIV-positive TB suspects was 58.8% (95%CI: 42.2%–73.6%) and 79.4% (95%CI: 74.8%–82.4%) respectively.

Conclusions

Pending point-of-care rapid diagnostic tests for TB disease, diagnostic algorithms are needed. The diagnostic accuracy of the 2007 WHO guideline to diagnose smear-negative TB is acceptable. There is, however, reluctance to comply with the guideline in terms of immediate treatment initiation.     

Nucleic acid amplification tests for diagnosis of smear-negative TB in a high HIV-prevalence setting: a prospective cohort study

Background

Nucleic acid amplification tests are sensitive for identifying Mycobacterium tuberculosis in populations with positive sputum smears for acid-fast bacilli, but less sensitive in sputum-smear-negative populations. Few studies have evaluated the clinical impact of these tests in low-income countries with high burdens of TB and HIV.

Methods

We prospectively enrolled 211 consecutive adults with cough ≥2 weeks and negative sputum smears at Mulago Hospital in Kampala, Uganda. We tested a single early-morning sputum specimen for Mycobacterium tuberculosis DNA using two nucleic acid amplification tests: a novel in-house polymerase chain reaction targeting the mycobacterial secA1 gene, and the commercial Amplified® Mycobacterium tuberculosis Direct (MTD) test (Gen-Probe Inc, San Diego, CA). We calculated the diagnostic accuracy of these index tests in reference to a primary microbiologic gold standard (positive mycobacterial culture of sputum or bronchoalveolar lavage fluid), and measured their likely clinical impact on additional tuberculosis cases detected among those not prescribed initial TB treatment.

Results

Of 211 patients enrolled, 170 (81%) were HIV-seropositive, with median CD4+ T-cell count 78 cells/µL (interquartile range 29-203). Among HIV-seropositive patients, 94 (55%) reported taking co-trimoxazole prophylaxis and 29 (17%) reported taking antiretroviral therapy. Seventy-five patients (36%) had culture-confirmed TB. Sensitivity of MTD was 39% (95% CI 28–51) and that of secA1 was 24% (95% CI 15–35). Both tests had specificities of 95% (95% CI 90–98). The MTD test correctly identified 18 (24%) TB patients not treated at discharge and led to a 72% relative increase in the smear-negative case detection rate.

Conclusions

The secA1 and MTD nucleic acid amplification tests had moderate sensitivity and high specificity for TB in a predominantly HIV-seropositive population with negative sputum smears. Although newer, more sensitive nucleic acid assays may enhance detection of Mycobacterium tuberculosis in sputum, even currently available tests can provide substantial clinical impact in smear-negative populations.     

Evaluation of a novel biphasic culture medium for recovery of mycobacteria: a multi-center study

Background

Mycobacterial culture and identification provide a definitive diagnosis of TB. Culture on Löwenstein-Jensen (L-J) medium is invariably delayed because of the slow growth of M. tuberculosis on L-J slants. Automated liquid culture systems are expensive. A low-cost culturing medium capable of rapidly indicating the presence of mycobacteria is needed. The aim of this study was to develop and evaluate a novel biphasic culture medium for the recovery of mycobacteria from clinical sputum specimens from suspected pulmonary tuberculosis patients.

Methods and Findings

The biphasic medium consisted of 7 ml units of L-J slant medium, 3 ml units of liquid culture medium, growth indicator and a mixture of antimicrobial agents. The decontamination sediments of sputum specimens were incubated in the biphasic culture medium at 37°C. Mycobacterial growth was determined based on the appearance of red granule sediments and the examination using acid-fast bacilli (AFB). The clinical sputum specimens were cultured in the biphasic medium, on L-J slants and in the Bactec MGIT 960 culture system. Among smear-positive specimens, the mycobacteria recovery rate of the biphasic medium was higher than that of the L-J slants (P<0.001) and similar to that of MGIT 960 (P>0.05). Among smear-negative specimens, the mycobacterial recovery rate of the biphasic medium was higher than that of L-J slants (P<0.001) and lower than that of MGIT 960 (P<0.05). The median times to detection of mycobacteria were 14 days, 20 days and 30 days for cultures grown in MGIT, in biphasic medium, on L-J slants for smear negative specimens, respectively (P<0.001).

Conclusions

The biphasic culture medium developed in this study is low-cost and suitable for mycobacterial recovery. It does not require any expensive detection instrumentation, decreases the time required for detection of M. tuberculosis complex, and increases the detection rate of M. tuberculosis complex.     

Baseline predictors of sputum culture conversion in pulmonary tuberculosis: importance of cavities, smoking, time to detection and W-Beijing genotype

Background

Time to detection (TTD) on automated liquid mycobacterial cultures is an emerging biomarker of tuberculosis outcomes. The M. tuberculosis W-Beijing genotype is spreading globally, indicating a selective advantage. There is a paucity of data on the association between baseline TTD and W-Beijing genotype and tuberculosis outcomes.

Aim

To assess baseline predictors of failure of sputum culture conversion, within the first 2 months of antitubercular therapy, in participants with pulmonary tuberculosis.

Design

Between May 2005 and August 2008 we conducted a prospective cohort study of time to sputum culture conversion in ambulatory participants with first episodes of smear and culture positive pulmonary tuberculosis attending two primary care clinics in Cape Town, South Africa. Rifampicin resistance (diagnosed on phenotypic susceptibility testing) was an exclusion criterion. Sputum was collected weekly for 8 weeks for mycobacterial culture on liquid media (BACTEC MGIT 960). Due to missing data, multiple imputation was performed. Time to sputum culture conversion was analysed using a Cox-proportional hazards model. Bayesian model averaging determined the posterior effect probability for each variable.

Results

113 participants were enrolled (30.1% female, 10.5% HIV-infected, 44.2% W-Beijing genotype, and 89% cavities). On Kaplan Meier analysis 50.4% of participants underwent sputum culture conversion by 8 weeks. The following baseline factors were associated with slower sputum culture conversion: TTD (adjusted hazard ratio (aHR) = 1.11, 95% CI 1.02; 1.2), lung cavities (aHR = 0.13, 95% CI 0.02; 0.95), ever smoking (aHR = 0.32, 95% CI 0.1; 1.02) and the W-Beijing genotype (aHR = 0.51, 95% CI 0.25; 1.07). On Bayesian model averaging, posterior probability effects were strong for TTD, lung cavitation and smoking and moderate for W-Beijing genotype.

Conclusion

We found that baseline TTD, smoking, cavities and W-Beijing genotype were associated with delayed 2 month sputum culture. Larger studies are needed to confirm the relationship between the W-Beijing genotype and sputum culture conversion.     

Characteristics and programme-defined treatment outcomes among childhood tuberculosis (TB) patients under the national TB programme in Delhi

Background

Childhood tuberculosis (TB) patients under India''s Revised National TB Control Programme (RNTCP) are managed using diagnostic algorithms and directly observed treatment with intermittent thrice-weekly short-course treatment regimens for 6–8 months. The assignment into pre-treatment weight bands leads to drug doses (milligram per kilogram) that are lower than current World Health Organization (WHO) guidelines for some patients.

Objectives

The main aim of our study was to describe the baseline characteristics and treatment outcomes reported under RNTCP for registered childhood (age <15 years) TB patients in Delhi. Additionally, we compared the reported programmatic treatment completion rates between children treated as per WHO recommended anti-TB drug doses with those children treated with anti-TB drug doses below that recommended in WHO guidelines.

Methods

For this cross-sectional retrospective study, we reviewed programme records of all 1089 TB patients aged <15 years registered for TB treatment from January to June, 2008 in 6 randomly selected districts of Delhi. WHO disease classification and treatment outcome definitions are used by RNTCP, and these were extracted as reported in programme records.

Results and Conclusions

Among 1074 patients with records available, 651 (61%) were females, 122 (11%) were <5 years of age, 1000 (93%) were new cases, and 680 (63%) had extra-pulmonary TB (EP-TB)—most commonly peripheral lymph node disease [310 (46%)]. Among 394 pulmonary TB (PTB) cases, 165 (42%) were sputum smear-positive. The overall reported treatment completion rate was 95%. Similar reported treatment completion rates were found in all subgroups assessed, including those patients whose drug dosages were lower than that currently recommended by WHO. Further studies are needed to assess the reasons for the low proportion of under-5 years of age TB case notifications, address challenges in reaching all childhood TB patients by RNTCP, the accuracy of diagnosis, and the clinical validity of reported programme defined treatment completion.     

Predictors on delay of initial health-seeking in new pulmonary tuberculosis cases among migrants population in East China

Objectives

To determine the length of delay in initial health-seeking in new pulmonary tuberculosis (PTB) cases among migrant population in the eastern part of China, and factors associated with it.

Methods

A cross-sectional study was conducted using a structured questionnaire in six counties in Shanghai, Guangdong and Jiangsu from May to October, 2008, to estimate the extent and factors responsible for delayed initial health-seeking of the new PTB cases. The interval between self-reported onset of TB symptoms and date of first attendance at any medical institution was determined. More than the median duration was defined as delayed health-seeking.

Results

A total of 323 new migrant PTB patients participated in the study. Only 6.5% had medical insurance. The median and mean durations to initial health-seeking were respectively 10 and 31 days. There was no significant association between socio-demographic factors and delayed initial health-seeking. Average monthly working days >24 (AOR, 1.61; 95% CI, 1.03–2.51), and hemoptysis or bloody sputum (AOR, 0.48; 95% CI, 0.28–0.85) were significantly associated with delayed initial health-seeking.

Conclusions

Interventions to improve health seeking behavior among the migrant population in China must focus on strengthening their labor, medical security and health education.     

Performance of the 2007 WHO Algorithm to Diagnose Smear-Negative Pulmonary Tuberculosis in a HIV Prevalent Setting

Background

The 2007 WHO algorithm for diagnosis of smear-negative pulmonary tuberculosis (PTB) including Mycobacterium tuberculosis (MTB) culture was evaluated in a HIV prevalent area of Kenya.

Methods

PTB smear-negative adult suspects were included in a prospective diagnostic study (2009–2011). In addition, program data (2008–2009) were retrospectively analysed. At the first consultation, clinical examination, chest X-ray, and sputum culture (Thin-Layer-Agar and Lowenstein-Jensen) were performed. Patients not started on TB treatment were clinically re-assessed after antibiotic course. The algorithm performance was calculated using culture as reference standard.

Results

380 patients were included prospectively and 406 analyzed retrospectively. Culture was positive for MTB in 17.5% (61/348) and 21.8% (72/330) of cases. Sensitivity of the clinical-radiological algorithm was 55.0% and 31.9% in the prospective study and the program data analysis, respectively. Specificity, positive and negative predictive values were 72.9%, 29.7% and 88.6% in the prospective study and 79.8%, 30.7% and 80.8% in the program data analysis. Performing culture increased the number of confirmed TB patients started on treatment by 43.3% in the prospective study and by 44.4% in the program data analysis. Median time to treatment of confirmed TB patients was 6 days in the prospective study and 27 days in the retrospective study. Inter-reader agreement for X-ray interpretation between the study clinician and a radiologist was low (Kappa coefficient = 0.11, 95%CI: 0.09–0.12). In a multivariate logistic analysis, past TB history, number of symptoms and signs at the clinical exam were independently associated with risk of overtreatment.

Conclusion

The clinical-radiological algorithm is suboptimal to diagnose smear-negative PTB. Culture increases significantly the proportion of confirmed TB cases started on treatment. Better access to rapid MTB culture and development of new diagnostic tests is necessary.     

Use of a molecular diagnostic test in AFB smear positive tuberculosis suspects greatly reduces time to detection of multidrug resistant tuberculosis

Background

The WHO has recommended the implementation of rapid diagnostic tests to detect and help combat M/XDR tuberculosis (TB). There are limited data on the performance and impact of these tests in field settings.

Methods

The performance of the commercially available Genotype MTBDRplus molecular assay was compared to conventional methods including AFB smear, culture and drug susceptibility testing (DST) using both an absolute concentration method on Löwenstein-Jensen media and broth-based method using the MGIT 960 system. Sputum specimens were obtained from TB suspects in the country of Georgia who received care through the National TB Program.

Results

Among 500 AFB smear-positive sputum specimens, 458 (91.6%) had both a positive sputum culture for Mycobacterium tuberculosis and a valid MTBDRplus assay result. The MTBDRplus assay detected isoniazid (INH) resistance directly from the sputum specimen in 159 (89.8%) of 177 specimens and MDR-TB in 109 (95.6%) of 114 specimens compared to conventional methods. There was high agreement between the MTBDRplus assay and conventional DST results in detecting MDR-TB (kappa = 0.95, p<0.01). The most prevalent INH resistance mutation was S315T (78%) in the katG codon and the most common rifampicin resistance mutation was S531L (68%) in the rpoB codon. Among 13 specimens from TB suspects with negative sputum cultures, 7 had a positive MTBDRplus assay (3 with MDR-TB). The time to detection of MDR-TB was significantly less using the MTBDRplus assay (4.2 days) compared to the use of standard phenotypic tests (67.3 days with solid media and 21.6 days with broth-based media).

Conclusions

Compared to conventional methods, the MTBDRplus assay had high accuracy and significantly reduced time to detection of MDR-TB in an area with high MDR-TB prevalence. The use of rapid molecular diagnostic tests for TB and drug resistance should increase the proportion of patients promptly placed on appropriate therapy.     

Comparison of Xpert MTB/RIF with other nucleic acid technologies for diagnosing pulmonary tuberculosis in a high HIV prevalence setting: a prospective study

Background

The Xpert MTB/RIF (Cepheid) non-laboratory-based molecular assay has potential to improve the diagnosis of tuberculosis (TB), especially in HIV-infected populations, through increased sensitivity, reduced turnaround time (2 h), and immediate identification of rifampicin (RIF) resistance. In a prospective clinical validation study we compared the performance of Xpert MTB/RIF, MTBDRplus (Hain Lifescience), LightCycler Mycobacterium Detection (LCTB) (Roche), with acid fast bacilli (AFB) smear microscopy and liquid culture on a single sputum specimen.

Methods and Findings

Consecutive adults with suspected TB attending a primary health care clinic in Johannesburg, South Africa, were prospectively enrolled and evaluated for TB according to the guidelines of the National TB Control Programme, including assessment for smear-negative TB by chest X-ray, clinical evaluation, and HIV testing. A single sputum sample underwent routine decontamination, AFB smear microscopy, liquid culture, and phenotypic drug susceptibility testing. Residual sample was batched for molecular testing. For the 311 participants, the HIV prevalence was 70% (n = 215), with 120 (38.5%) culture-positive TB cases. Compared to liquid culture, the sensitivities of all the test methodologies, determined with a limited and potentially underpowered sample size (n = 177), were 59% (47%–71%) for smear microscopy, 76% (64%–85%) for MTBDRplus, 76% (64%–85%) for LCTB, and 86% (76%–93%) for Xpert MTB/RIF, with specificities all >97%. Among HIV+ individuals, the sensitivity of the Xpert MTB/RIF test was 84% (69%–93%), while the other molecular tests had sensitivities reduced by 6%. TB detection among smear-negative, culture-positive samples was 28% (5/18) for MTBDRplus, 22% (4/18) for LCTB, and 61% (11/18) for Xpert MTB/RIF. A few (n = 5) RIF-resistant cases were detected using the phenotypic drug susceptibility testing methodology. Xpert MTB/RIF detected four of these five cases (fifth case not tested) and two additional phenotypically sensitive cases.

Conclusions

The Xpert MTB/RIF test has superior performance for rapid diagnosis of Mycobacterium tuberculosis over existing AFB smear microscopy and other molecular methodologies in an HIV- and TB-endemic region. Its place in the clinical diagnostic algorithm in national health programs needs exploration. Please see later in the article for the Editors'' Summary     

Prevalence of Pulmonary Tuberculosis among Adults in a Rural Sub-District of South India

Background

We conducted a survey to estimate point prevalence of bacteriologically positive pulmonary TB (PTB) in a rural area in South India, implementing TB program DOTS strategy since 2002.

Methods

Survey was conducted among persons ≥15 years of age in fifteen clusters selected by simple random sampling; each consisting of 5–12 villages. Persons having symptoms suggestive of PTB or history of anti-TB treatment (ATT) were eligible for sputum examination by smear microscopy for Acid Fast Bacilli and culture for Mycobacterium tuberculosis; two sputum samples were collected from each eligible person.Persons with one or both sputum specimen positive on microscopy and/or culture were labeled suffering from PTB. Prevalence was estimated after imputing missing values to correct for bias introduced by incompleteness of data.In six clusters, registered persons were also screened by X-ray chest. Persons with any abnormal shadow on X-ray were eligible for sputum examination in addition to those with symptoms and ATT. Multiplication factor calculated as ratio of prevalence while using both screening tools to prevalence using symptoms screening alone was applied to entire study population to estimate prevalence corrected for non-screening by X-ray.

Results

Of 71,874 residents ≥15 years of age, 63,362 (88.2%) were screened for symptoms and ATT. Of them, 5120 (8.1%) - 4681 (7.4%) with symptoms and an additional 439 (0.7%) with ATT were eligible for sputum examination. Spot specimen were collected from 4850 (94.7%) and early morning sputum specimens from 4719 (92.2%). Using symptom screening alone, prevalence of smear, culture and bacteriologically positive PTB in persons ≥15 years of age was 83 (CI: 57–109), 152 (CI: 108–197) and 196 (CI :145–246) per 100,000 population respectively. Prevalence corrected for non-screening by X-ray was 108 (CI: 82–134), 198 (CI: 153–243) and 254 (CI: 204–301) respectively.

Conclusion

Observed prevalence suggests further strengthening of TB control program.     

Tuberculosis contact screening and isoniazid preventive therapy in a South Indian district: operational issues for programmatic consideration

Background

Under India''s Revised National Tuberculosis Control Programme (RNTCP), all household contacts of sputum smear positive Pulmonary Tuberculosis (PTB) patients are screened for TB. In the absence of active TB disease, household contacts aged <6 years are eligible for Isoniazid Preventive Therapy (IPT) (5 milligrams/kilogram body weight/day) for 6 months.

Objectives

To estimate the number of household contacts aged <6 years, of sputum smear positive PTB patients registered for treatment under RNTCP from April to June''2008 in Krishna District, to assess the extent to which they are screened for TB disease and in its absence initiated on IPT.

Methods

A cross sectional study was conducted. Households of all smear positive PTB cases (n = 848) registered for treatment from April to June''2008 were included. Data on the number of household contacts aged <6 years, the extent to which they were screened for TB disease, and the status of initiation of IPT, was collected.

Results

Households of 825 (97%) patients were visited, and 172 household contacts aged <6 years were identified. Of them, 116 (67%) were evaluated for TB disease; none were found to be TB diseased and 97 (84%) contacts were initiated on IPT and 19 (16%) contacts were not initiated on IPT due to shortage of INH tablets in peripheral health centers. The reasons for non-evaluation of the remaining eligible children (n = 56, 33%) include no home visit by the health staff in 25 contacts, home visit done but not evaluated in 31 contacts. House-hold contacts in rural areas were less likely to be evaluated and initiated on IPT [risk ratio 6.65 (95% CI; 3.06–14.42)].

Conclusion

Contact screening and IPT implementation under routine programmatic conditions is sub-optimal. There is an urgent need to sensitize all concerned programme staff on its importance and establishment of mechanisms for rigorous monitoring.     

Rapid diagnosis of smear-negative tuberculosis using immunology and microbiology with induced sputum in HIV-infected and uninfected individuals

Rationale and Objectives

Blood-based studies have demonstrated the potential of immunological assays to detect tuberculosis. However lung fluid sampling may prove superior as it enables simultaneous microbiological detection of mycobacteria to be performed. Until now this has only been possible using the expensive and invasive technique of broncho-alveolar lavage. We sought to evaluate an immunoassay using non-invasive induced-sputum to diagnose active tuberculosis.

Methods and Results

Prospective cohort study of forty-two spontaneous sputum smear-negative or sputum non-producing adults under investigation for tuberculosis. CD4 lymphocytes specific to purified-protein-derivative of Mycobacterium tuberculosis actively synthesising interferon-gamma were measured by flow cytometry and final diagnosis compared to immunoassay using a cut-off of 0.5%. Sixteen subjects (38%) were HIV-infected (median CD4 count [range] = 332 cells/µl [103–748]). Thirty-eight (90%) were BCG-vaccinated. In 27 subjects diagnosed with active tuberculosis, the median [range] percentage of interferon-gamma synthetic CD4+ lymphocytes was 2.77% [0–23.93%] versus 0% [0–2.10%] in 15 negative for active infection (p<0.0001). Sensitivity and specificity of the immunoassay versus final diagnosis of active tuberculosis were 89% (24 of 27) and 80% (12 of 15) respectively. The 3 positive assays in the latter group occurred in subjects diagnosed with quiescent/latent tuberculosis. Assay performance was unaffected by HIV-status, BCG-vaccination or disease site. Combining this approach with traditional microbiological methods increased the diagnostic yield to 93% (25 of 27) alongside acid-fast bacilli smear and 96% (26 of 27) alongside tuberculosis culture.

Conclusions

These data demonstrate for the first time that a rapid immunological assay to diagnose active tuberculosis can be performed successfully in combination with microbiological methods on a single induced-sputum sample.     

Association of BMI category change with TB treatment mortality in HIV-positive smear-negative and extrapulmonary TB patients in Myanmar and Zimbabwe

Objective

The HIV epidemic has increased the proportion of patients with smear-negative and extrapulmonary tuberculosis (TB) diagnoses, with related higher rates of poor TB treatment outcomes. Unlike in smear-positive pulmonary TB, no interim markers of TB treatment progress are systematically used to identify individuals most at risk of mortality. The objective of this study was to assess the association of body mass index (BMI) change at 1 month (±15 days) from TB treatment start with mortality among HIV-positive individuals with smear-negative and extrapulmonary TB.

Methods and Findings

A retrospective cohort study of adult HIV-positive new TB patients in Médecins Sans Frontières (MSF) treatment programmes in Myanmar and Zimbabwe was conducted using Cox proportional hazards regression to estimate the association between BMI category change and mortality. A cohort of 1090 TB patients (605 smear-negative and 485 extrapulmonary) was followed during TB treatment with mortality rate of 28.9 per 100 person-years. In multivariable analyses, remaining severely underweight or moving to a lower BMI category increased mortality (adjusted hazard ratio 4.05, 95% confidence interval 2.77–5.91, p<0.001) compared with remaining in the same or moving to a higher BMI category.

Conclusions

We found a strong association between BMI category change during the first month of TB treatment and mortality. BMI category change could be used to identify individuals most at risk of mortality during TB treatment among smear-negative and extrapulmonary patients.     

Factors associated with negative direct sputum examination in Asian and African HIV-infected patients with tuberculosis (ANRS 1260)

Objective

To identify factors associated with negative direct sputum examination among African and Cambodian patients co-infected by Mycobacterium tuberculosis and HIV.

Design

Prospective multicenter study (ANRS1260) conducted in Cambodia, Senegal and Central African Republic.

Methods

Univariate and multivariate analyses (logistic regression) were used to identify clinical and radiological features associated with negative direct sputum examination in HIV-infected patients with positive M. tuberculosis culture on Lowenstein-Jensen medium.

Results

Between September 2002 and December 2005, 175 co-infected patients were hospitalized with at least one respiratory symptom and pulmonary radiographic anomaly. Acid-fast bacillus (AFB) examination was positive in sputum samples from 110 subjects (63%) and negative in 65 patients (37%). Most patients were at an advanced stage of HIV disease (92% at stage III or IV of the WHO classification) with a median CD4 cell count of 36/mm³. In this context, we found that sputum AFB negativity was more frequent in co-infected subjects with associated respiratory tract infections (OR = 2.8 [95%CI:1.1–7.0]), dyspnea (OR = 2.5 [95%CI:1.1–5.6]), and localized interstitial opacities (OR = 3.1 [95%CI:1.3–7.6]), but was less frequent with CD4≤50/mm³ (OR = 0.4 [95%CI:0.2–0.90), adenopathies (OR = 0.4 [95%CI:0.2–0.93]) and cavitation (OR = 0.1 [95%CI:0.03–0.6]).

Conclusions

One novel finding of this study is the association between concomitant respiratory tract infection and negative sputum AFB, particularly in Cambodia. This finding suggests that repeating AFB testing in AFB-negative patients should be conducted when broad spectrum antibiotic treatment does not lead to complete recovery from respiratory symptoms. In HIV-infected patients with a CD4 cell count below 50/mm3 without an identified cause of pneumonia, systematic AFB direct sputum examination is justified because of atypical clinical features (without cavitation) and high pulmonary mycobacterial burden.     

Validation of a clinical-radiographic score to assess the probability of pulmonary tuberculosis in suspect patients with negative sputum smears

Background

Clinical suspects of pulmonary tuberculosis in which the sputum smears are negative for acid fast bacilli represent a diagnostic challenge in resource constrained settings. Our objective was to validate an existing clinical-radiographic score that assessed the probability of smear-negative pulmonary tuberculosis (SNPT) in high incidence settings in Peru.

Methodology/Principal Findings

We included in two referral hospitals in Lima patients with clinical suspicion of pulmonary tuberculosis and two or more negative sputum smears. Using a published but not externally validated score, patients were classified as having low, intermediate or high probability of pulmonary tuberculosis. The reference standard for the diagnosis of tuberculosis was a positive sputum culture in at least one of 2 liquid (MGIT or Middlebrook 7H9) and 1 solid (Ogawa) media. Prevalence of tuberculosis was calculated in each of the three probability groups.684 patients were included. 184 (27.8%) had a diagnosis of pulmonary tuberculosis. The score did not perform well in patients with a previous history of pulmonary tuberculosis. In patients without, the prevalence of tuberculosis was 5.1%, 31.7% and 72% in the low, intermediate and high probability group respectively. The area under de ROC curve was 0.76 (95% CI 0.72–0.80) and scores ≥6 had a positive LR of 10.9.

Conclusions/Significance

In smear negative suspects without previous history of tuberculosis, the clinical-radiographic score can be used as a tool to assess the probability of pulmonary tuberculosis and to guide the decision to initiate or defer treatment or to requesting additional tests.     

The Risk and Timing of Tuberculosis Diagnosed in Smear-Negative TB Suspects: A 12 Month Cohort Study in Harare,Zimbabwe

Background

Cases of smear-negative TB have increased dramatically in high prevalence HIV settings and pose considerable diagnostic and management challenges.

Methods and Findings

Between February 2006 and July 2007, a cohort study nested within a cluster-randomised trial of community-based case finding strategies for TB in Harare, Zimbabwe was undertaken. Participants who had negative sputum smears and remained symptomatic of TB were follow-up for one year with standardised investigations including HIV testing, repeat sputum smears, TB culture and chest radiography. Defaulters were actively traced to the community. The objectives were to investigate the incidence and risk factors for TB. TB was diagnosed in 218 (18.2%) participants, of which 39.4% was bacteriologically confirmed. Most cases (84.2%) were diagnosed within 3 months, but TB incidence remained high thereafter (111.3 per 1000 person-years, 95% CI: 86.6 to 146.3). HIV prevalence was 63.3%, and HIV-infected individuals had a 3.5-fold higher risk of tuberculosis than HIV-negative individuals.

Conclusion

We found that diagnosis of TB was insensitive and slow, even with early radiography and culture. Until more sensitive and rapid diagnostic tests become widely available, a much more proactive and integrated approach towards prompt initiation of ART, ideally from within TB clinics and without waiting for TB to be excluded, is needed to minimise the risk and consequences of diagnostic delay.     

Evaluation of combined LED-fluorescence microscopy and bleach sedimentation for diagnosis of tuberculosis at peripheral health service level

Background

Sputum microscopy is the only diagnostic for tuberculosis (TB) available at peripheral levels of health service in resource-poor countries. Its sensitivity is reduced in high HIV-prevalence settings. Sodium hypochlorite (NaOCl) specimen sedimentation prior microscopy and light-emitting diode (LED)-fluorescence microscopy (FM) can individually improve performance of microscopy. This study aimed to evaluate the performance of combined LED-FM and NaOCl sputum sedimentation for TB detection at peripheral level of health services.

Methods

A prospective study was conducted in an urban health clinic in Nairobi, Kenya. Three sputum specimens were collected over 2 days from consecutive TB suspects. Smears were prepared and stained with auramine O and Ziehl-Neelsen (ZN) methods. Bleach (3.5%) was added to the remaining specimen before overnight sedimentation at room temperature. Auramine O staining was performed on smears of sediment. A 4^th specimen was collected for TB culture. Auramine smears were read under the same microscope as used for ZN smears, but equipped with the LED FluoLED™ fluorescence illuminator.

Results

497 patients were included, and 1394 specimens collected. The yield of positive specimen was significantly increased after NaOCl sedimentation (24.9%) compared to direct LED-FM (20.6%) and direct ZN (20.3%). In detecting smear-positive patients, sensitivity was 78.5% for LED-FM after NaOCl sedimentation compared to 73.2% and 72.0% for direct LED-FM (P = 0.06) and direct ZN (P = 0.06), respectively. Specificity was 87.8% for LED-FM after NaOCl sedimentation compared to 96.7% and 95.9% for direct LED-FM (P<0.01) and direct ZN (P<0.01), respectively. Inter-reading agreement (kappa = 0.7) and technicians'' acceptability were good.

Conclusion

NaOCl sedimentation did not improve the performance of LED-FM in the diagnosis of pulmonary TB at peripheral health service level.     

Screening for HIV-associated tuberculosis and rifampicin resistance before antiretroviral therapy using the Xpert MTB/RIF assay: a prospective study

Background

The World Health Organization has endorsed the Xpert MTB/RIF assay for investigation of patients suspected of having tuberculosis (TB). However, its utility for routine TB screening and detection of rifampicin resistance among HIV-infected patients with advanced immunodeficiency enrolling in antiretroviral therapy (ART) services is unknown.

Methods and Findings

Consecutive adult HIV-infected patients with no current TB diagnosis enrolling in an ART clinic in a South African township were recruited regardless of symptoms. They were clinically characterised and invited to provide two sputum samples at a single visit. The accuracy of the Xpert MTB/RIF assay for diagnosing TB and drug resistance was assessed in comparison with other tests, including fluorescence smear microscopy and automated liquid culture (gold standard) and drug susceptibility testing. Of 515 patients enrolled, 468 patients (median CD4 cell count, 171 cells/µl; interquartile range, 102–236) produced at least one sputum sample, yielding complete sets of results from 839 samples. Mycobacterium tuberculosis was cultured from 81 patients (TB prevalence, 17.3%). The overall sensitivity of the Xpert MTB/RIF assay for culture-positive TB was 73.3% (specificity, 99.2%) compared to 28.0% (specificity, 100%) using smear microscopy. All smear-positive, culture-positive disease was detected by Xpert MTB/RIF from a single sample (sensitivity, 100%), whereas the sensitivity for smear-negative, culture-positive TB was 43.4% from one sputum sample and 62.3% from two samples. Xpert correctly identified rifampicin resistance in all four cases of multidrug-resistant TB but incorrectly identified resistance in three other patients whose disease was confirmed to be drug sensitive by gene sequencing (specificity, 94.1%; positive predictive value, 57%).

Conclusions

In this population of individuals at high risk of TB, intensive screening using the Xpert MTB/RIF assay increased case detection by 45% compared with smear microscopy, strongly supporting replacement of microscopy for this indication. However, despite the ability of the assay to rapidly detect rifampicin-resistant disease, the specificity for drug-resistant TB was sub-optimal. Please see later in the article for the Editors'' Summary     

Clinical Utility of a Commercial LAM-ELISA Assay for TB Diagnosis in HIV-Infected Patients Using Urine and Sputum Samples

Background

The accurate diagnosis of TB in HIV-infected patients, particularly with advanced immunosuppression, is difficult. Recent studies indicate that a lipoarabinomannan (LAM) assay (Clearview-TB®-ELISA) may have some utility for the diagnosis of TB in HIV-infected patients; however, the precise subgroup that may benefit from this technology requires clarification. The utility of LAM in sputum samples has, hitherto, not been evaluated.

Methods

LAM was measured in sputum and urine samples obtained from 500 consecutively recruited ambulant patients, with suspected TB, from 2 primary care clinics in South Africa. Culture positivity for M. tuberculosis was used as the reference standard for TB diagnosis.

Results

Of 440 evaluable patients 120/387 (31%) were HIV-infected. Urine-LAM positivity was associated with HIV positivity (p = 0.007) and test sensitivity, although low, was significantly higher in HIV-infected compared to uninfected patients (21% versus 6%; p<0.001), and also in HIV-infected participants with a CD4 <200 versus >200 cells/mm³ (37% versus 0%; p = 0.003). Urine-LAM remained highly specific in all 3 subgroups (95%–100%). 25% of smear-negative but culture-positive HIV-infected patients with a CD4 <200 cells/mm³ were positive for urine-LAM. Sputum-LAM had good sensitivity (86%) but poor specificity (15%) likely due to test cross-reactivity with several mouth-residing organisms including actinomycetes and nocardia species.

Conclusions

These preliminary data indicate that in a high burden primary care setting the diagnostic usefulness of urine-LAM is limited, as a rule-in test, to a specific patient subgroup i.e. smear-negative HIV-infected TB patients with a CD4 count <200 cells/mm³, who would otherwise have required further investigation. However, even in this group sensitivity was modest. Future and adequately powered studies in a primary care setting should now specifically target patients with suspected TB who have advanced HIV infection.     

Poor performance of serological tests in the diagnosis of pulmonary tuberculosis: evidence from a contact tracing field study

Background

Delayed or missed diagnosis of TB continues to fuel the global TB epidemic, especially in resource limited settings. Use of serology for the diagnosis of tuberculosis, commonly used in India, is another factor. In the present study a commercially available serodiagnostic assay was assessed for its diagnostic value in combination with smear, culture and clinical manifestations.

Methodology/Principal Findings

A total of 2300 subjects were recruited for the study, but 1041 subjects were excluded for various reasons. Thus 1259 subjects were included in the study of which 470 were pulmonary tuberculosis cases (440 of 470 were culture-positive) and 789 were their asymptomatic contacts. A house-to-house survey method was used. Blood samples were tested for IgM, IgA, and IgG antibodies using the Pathozyme Myco M (IgM), Myco A (IgA) and Myco G (IgG) enzyme immunoassay (EIA). Out of 470 PTB cases, BCG scar was positive in 82.34%. The Mantoux test and smear positivity rates in PTB cases were 94.3% (430/456), and 65.32% (307/470), respectively. Among the asymptomatic contacts, BCG scar was positive in 95.3% and Mantoux test was positive in 80.66% (442/548) contacts. No contact was found falsely smear positive. The sensitivity of IgM, IgA, and IgG EIA tests was 48.7%, 25.7% and 24.4%, respectively, while the specificity was 71.5%, 80.5%, 76.6%, respectively. Performance of EIAs was not affected by the previous BCG vaccination. However, prior BCG vaccination was statistically significantly (p = 0.005) associated with Mantoux test positivity in PTB cases but not in contacts (p = 0.127). The agreement between serology and Mantoux test was not significant.

Conclusion

The commercial serological test evaluated showed poor sensitivity and specificity and suggests no utility for detection of pulmonary tuberculosis.