Longitudinal Dynamics of a Blood Transcriptomic Signature of Tuberculosis.

Rationale: Performance of blood transcriptomic tuberculosis (TB) signatures in longitudinal studies and effects of TB-preventive therapy and coinfection with HIV or respiratory organisms on transcriptomic signatures has not been systematically studied. Objectives: We evaluated longitudinal kinetics of an 11-gene blood transcriptomic TB signature, RISK11, and effects of TB-preventive therapy (TPT) and respiratory organisms on RISK11 signature score, in HIV-uninfected and HIV-infected individuals. Methods: RISK11 was measured in a longitudinal study of RISK11-guided TPT in HIV-uninfected adults, a cross-sectional respiratory organisms cohort, or a longitudinal study in people living with HIV (PLHIV). HIV-uninfected RISK11+ participants were randomized to TPT or no TPT; RISK11- participants received no TPT. PLHIV received standard-of-care antiretroviral therapy and TPT. In the cross-sectional respiratory organisms cohort, viruses and bacteria in nasopharyngeal and oropharyngeal swabs were quantified by real-time quantitative PCR. Measurements and Main Results: RISK11+ status was transient in most of the 128 HIV-negative participants with longitudinal samples; more than 70% of RISK11+ participants reverted to RISK11- by 3 months, irrespective of TPT. By comparison, reversion from a RISK11+ state was less common in 645 PLHIV (42.1%). Non-HIV viral and nontuberculous bacterial organisms were detected in 7.2% and 38.9% of the 1,000 respiratory organisms cohort participants, respectively, and among those investigated for TB, 3.8% had prevalent disease. Median RISK11 scores (%) were higher in participants with viral organisms alone (46.7%), viral and bacterial organisms (42.8%), or prevalent TB (85.7%) than those with bacterial organisms other than TB (13.4%) or no organisms (14.2%). RISK11 could not discriminate between prevalent TB and viral organisms. Conclusions: Positive RISK11 signature status is often transient, possibly due to intercurrent viral infection, highlighting potentially important challenges for implementation of these biomarkers as new tools for TB control.

Prevalence and antimicrobial resistance profiles of respiratory microbial flora in African children with HIV-associated chronic lung disease.

BACKGROUND: HIV-associated chronic lung disease (CLD) is common among children living with HIV (CLWH) in sub-Saharan Africa, including those on antiretroviral therapy (ART). However, the pathogenesis of CLD and its possible association with microbial determinants remain poorly understood. We investigated the prevalence, and antibiotic susceptibility of Streptococcus pneumoniae (SP), Staphylococcus aureus (SA), Haemophilus influenzae (HI), and Moraxella catarrhalis (MC) among CLWH (established on ART) who had CLD (CLD+), or not (CLD-) in Zimbabwe and Malawi. METHODS: Nasopharyngeal swabs (NP) and sputa were collected from CLD+ CLWH (defined as forced-expiratory volume per second z-score < - 1 without reversibility post-bronchodilation with salbutamol), at enrolment as part of a randomised, placebo-controlled trial of azithromycin (BREATHE trial - NCT02426112 ), and from age- and sex-matched CLD- CLWH. Samples were cultured, and antibiotic susceptibility testing was conducted using disk diffusion. Risk factors for bacterial carriage were identified using questionnaires and analysed using multivariate logistic regression. RESULTS: A total of 410 participants (336 CLD+, 74 CLD-) were enrolled (median age, 15 years [IQR = 13-18]). SP and MC carriage in NP were higher in CLD+ than in CLD- children: 46% (154/336) vs. 26% (19/74), p = 0.008; and 14% (49/336) vs. 3% (2/74), p = 0.012, respectively. SP isolates from the NP of CLD+ children were more likely to be non-susceptible to penicillin than those from CLD- children (36% [53/144] vs 11% [2/18], p = 0.036). Methicillin-resistant SA was uncommon [4% (7/195)]. In multivariate analysis, key factors associated with NP bacterial carriage included having CLD (SP: adjusted odds ratio (aOR) 2 [95% CI 1.1-3.9]), younger age (SP: aOR 3.2 [1.8-5.8]), viral load suppression (SP: aOR 0.6 [0.4-1.0], SA: 0.5 [0.3-0.9]), stunting (SP: aOR 1.6 [1.1-2.6]) and male sex (SA: aOR 1.7 [1.0-2.9]). Sputum bacterial carriage was similar in both groups (50%) and was associated with Zimbabwean site (SP: aOR 3.1 [1.4-7.3], SA: 2.1 [1.1-4.2]), being on ART for a longer period (SP: aOR 0.3 [0.1-0.8]), and hot compared to rainy season (SP: aOR 2.3 [1.2-4.4]). CONCLUSIONS: CLD+ CLWH were more likely to be colonised by MC and SP, including penicillin-non-susceptible SP strains, than CLD- CLWH. The role of these bacteria in CLD pathogenesis, including the risk of acute exacerbations, should be further studied.

Tuberculosis Diagnosis in Children Using Xpert Ultra on Different Respiratory Specimens.

Rationale: Microbiological confirmation of pulmonary tuberculosis in children is desirable.Objectives: To investigate the diagnostic accuracy and incremental yield of Xpert MTB/RIF Ultra (Ultra; Cepheid), a new rapid test, on repeated induced sputum, nasopharyngeal aspirates, and combinations of specimens.Methods: Consecutive South African children hospitalized with suspected pulmonary tuberculosis were enrolled.Measurements and Main Results: Induced sputum (IS) and nasopharyngeal aspirates (NPAs) were obtained. NPAs were frozen; IS underwent liquid culture, and an aliquot was frozen. Ultra was performed on thawed NPAs and IS specimens individually. Children were categorized as confirmed, unconfirmed, or unlikely tuberculosis according to NIH consensus case definitions. The diagnostic accuracy of Ultra was compared with liquid culture on IS. In total, 195 children (median age: 23.3 mo; 32 [16.4%] HIV-infected) had one IS and NPA, and 130 had two NPAs. There were 40 (20.5%) culture-confirmed cases. Ultra was positive on NPAs in 26 (13.3%) and on IS in 31 (15.9%). Sensitivity and specificity of Ultra on one NPA were 46% and 98%, respectively, and similar by HIV status. Sensitivity and specificity of Ultra on one IS were 74.3% and 96.9% respectively. Combining one NPA and one IS increased sensitivity to 80%. Sensitivity using Ultra on two NPAs was 54.2%, increasing to 87.5% with an IS Ultra.Conclusions: IS provides a better specimen than repeated NPA for rapid diagnosis using Ultra. However, Ultra testing of combinations of specimens provides a novel strategy that can be adapted to identify most children with confirmed pulmonary tuberculosis.

Microbiological diagnosis of pulmonary tuberculosis in children by oral swab polymerase chain reaction.

Microbiological diagnosis of pediatric pulmonary tuberculosis (TB) is challenging due to the difficulty of collecting and testing sputum from children. We investigated whether easily-obtained oral swab samples are useful alternatives or supplements to sputum. Oral swabs and induced sputum (IS) were collected from 201 South African children with suspected pulmonary TB. IS samples were tested by mycobacterial culture and Xpert MTB/RIF. Oral swabs were tested by PCR targeting IS6110. Children were categorized as Confirmed TB (microbiologic confirmation on IS), Unconfirmed TB (clinical diagnosis only), or Unlikely TB (recovery without TB treatment). Relative to Confirmed TB, PCR on two oral swabs per child was 43% sensitive and 93% specific. This sensitivity fell below that of sputum Xpert (64%). Among children with either Confirmed or Unconfirmed TB, PCR on two oral swabs per child was 31% sensitive and 93% specific, which was more sensitive than sputum testing among this group (21%). Although oral swab analysis had low sensitivity in sputum-positive children, it detected TB in a significant proportion of sputum-negative children who were clinically diagnosed with TB. Specificity at 93% was suboptimal but may improve with the use of automated methods. With further development, oral swabs may become useful supplements to sputum as samples for diagnosis of pulmonary TB in children.

Impact of Xpert MTB/RIF for TB diagnosis in a primary care clinic with high TB and HIV prevalence in South Africa: a pragmatic randomised trial.

BACKGROUND: Xpert MTB/RIF is approved for use in tuberculosis (TB) and rifampicin-resistance diagnosis. However, data are limited on the impact of Xpert under routine conditions in settings with high TB burden. METHODS AND FINDINGS: A pragmatic prospective cluster-randomised trial of Xpert for all individuals with presumptive (symptomatic) TB compared to the routine diagnostic algorithm of sputum microscopy and limited use of culture was conducted in a large TB/HIV primary care clinic. The primary outcome was the proportion of bacteriologically confirmed TB cases not initiating TB treatment by 3 mo after presentation. Secondary outcomes included time to TB treatment and mortality. Unblinded randomisation occurred on a weekly basis. Xpert and smear microscopy were performed on site. Analysis was both by intention to treat (ITT) and per protocol. Between 7 September 2010 and 28 October 2011, 1,985 participants were assigned to the Xpert (n = 982) and routine (n = 1,003) diagnostic algorithms (ITT analysis); 882 received Xpert and 1,063 routine (per protocol analysis). 13% (32/257) of individuals with bacteriologically confirmed TB (smear, culture, or Xpert) did not initiate treatment by 3 mo after presentation in the Xpert arm, compared to 25% (41/167) in the routine arm (ITT analysis, risk ratio 0.51, 95% CI 0.33-0.77, p = 0.0052). The yield of bacteriologically confirmed TB cases among patients with presumptive TB was 17% (167/1,003) with routine diagnosis and 26% (257/982) with Xpert diagnosis (ITT analysis, risk ratio 1.57, 95% CI 1.32-1.87, p<0.001). This difference in diagnosis rates resulted in a higher rate of treatment initiation in the Xpert arm: 23% (229/1,003) and 28% (277/982) in the routine and Xpert arms, respectively (ITT analysis, risk ratio 1.24, 95% CI 1.06-1.44, p = 0.013). Time to treatment initiation was improved overall (ITT analysis, hazard ratio 0.76, 95% CI 0.63-0.92, p = 0.005) and among HIV-infected participants (ITT analysis, hazard ratio 0.67, 95% CI 0.53-0.85, p = 0.001). There was no difference in 6-mo mortality with Xpert versus routine diagnosis. Study limitations included incorrect intervention allocation for a high proportion of participants and that the study was conducted in a single clinic. CONCLUSIONS: These data suggest that in this routine primary care setting, use of Xpert to diagnose TB increased the number of individuals with bacteriologically confirmed TB who were treated by 3 mo and reduced time to treatment initiation, particularly among HIV-infected participants. TRIAL REGISTRATION: Pan African Clinical Trials Registry PACTR201010000255244. Please see later in the article for the Editors' Summary.