NATCON 2022 Panel discussion: "TB vaccination - Going Forward".

Vaccination is important tuberculosis (TB) preventive strategy that is essential to achieve the goals of the End TB strategy. The BCG vaccination at birth offers protection against TB in young children but not in adolescents and adults. New TB vaccines are the need of the hour. The TB vaccine development pipeline in the past years is encouraging with newer TB vaccines in clinical trials in humans. The focus of the newer TB vaccine is the prevention of infection, disease, and recurrence of TB disease. Therapeutic vaccines focus on better treatment outcomes and prevention of TB recurrence. BCG revaccination is of current interest. Novel, safe, and efficient TB vaccines that prevent TB infection and disease if introduced in 2025 could drastically reduce the rate of TB incidence. However, the development of an effective vaccine for TB is challenging. Engagement of stakeholders, mobilizing funding, and advocacy could accelerate the newer TB vaccine development process.

Inactivated COVID-19 vaccines: durability of Covaxin/BBV152 induced immunity against variants of concern.

BACKGROUND: Covaxin/BBV152 is one of the most widely used vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and one of the few vaccines used extensively in low- and middle-income countries (LMIC). METHODS: We investigated the effect of Covaxin on the SARS-CoV-2 specific IgG and IgA and neutralizing antibody (NAb) levels at baseline (M0) and at Months 1 (M1), 2 (M2), 3 (M3), 4 (M4), 6 (M6) and 12 (M12) following vaccination in healthcare workers. In addition, we also examined the NAb levels against variant lineages of B.1.617.2 (Delta, India), B.1.617.2.1 (Delta Plus, India), B.1.351 (Beta, SA), B.1.1.7 (Alpha, UK) and B.1.1.529 (Omicron). RESULTS: Covaxin induces enhanced SARS-CoV-2 binding antibodies of IgG and IgA responses against both spike (S) and nucleocapsid (N) antigens at M1, M2, M3, M4, M6 and M12 in comparison with M0. Our data also reveal that NAb levels against the ancestral strain (Wuhan, wild type) are elevated and sustained at M1, M2, M3, M4, M6 and M12 in comparison with M0 and against variant lineages of B.1.617.2 (Delta, India), B.1.617.2.1 (Delta Plus, India), B.1.351 (Beta, SA) and B.1.1.7 (Alpha, UK) are elevated at M3, M6 and M12 in comparison with M0. However, NAb levels against B.1.1.529 (Omicron) was consistently below the limit of detection except at M12. CONCLUSION: Thus, Covaxin induces an enhanced humoral immune response, with persistence till at least 12 months post-vaccination against most SARS-CoV-2 variants.

Prime-Boost Vaccination With Covaxin/BBV152 Induces Heightened Systemic Cytokine and Chemokine Responses.

Covaxin/BBV152 is a whole virion inactivated SARS-CoV-2 vaccine. The effect of prime-boost vaccination with Covaxin on systemic immune responses is not known. We investigated the effect of Covaxin on the plasma levels of a wide panel of cytokines and chemokines at baseline (M0) and at months 1 (M1), 2 (M2) and 3 (M3) following prime-boost vaccination in healthy volunteers. Our results demonstrate that Covaxin induces enhanced plasma levels of Type 1 cytokines (IFNγ, IL-2, TNFα), Type 2/regulatory cytokines (IL-4, IL-5, IL-10 and IL-13), Type 17 cytokine (IL-17A), other pro-inflammatory cytokines (IL-6, IL-12, IL-1α, IL-1ß) and other cytokines (IL-3 and IL-7) but diminished plasma levels of IL-25, IL-33, GM-CSF and Type 1 IFNs. Covaxin also induced enhanced plasma levels of CC chemokine (CCL4) and CXC chemokines (CXCL1, CXCL2 and CX3CL1) but diminished levels of CXCL10. Covaxin vaccination induces enhanced cytokine and chemokine responses as early as month 1, following prime-boost vaccination, indicating robust activation of innate and adaptive immune responses in vaccine recipients.

Antibody responses to the BBV152 vaccine in individuals previously infected with SARS-CoV-2: A pilot study.

Background & objectives: Vaccination against SARS-CoV-2 is a recommendation from the World Health Organization as the foremost preference in the current situation to control the COVID-19 pandemic. BBV152 is one of the approved vaccines against SARS-CoV-2 in India. In this study, we determined SARS-CoV-2-specific antibody levels at day 0 (baseline, before vaccination), day 28 ± 2 post-first dose (month 1) and day 56 ± 2 post-first dose (month 2) of BBV152 whole-virion-inactivated SARS-CoV-2 recipients, and compared the antibody responses of individuals with confirmed pre-vaccination SARS-CoV-2 infection to those individuals without prior evidence of infection. Methods: Blood samples were collected from 114 healthcare professionals and frontline workers who received BBV152 vaccine from February to May & June 2021. Prior infection with SARS-CoV-2 was determined at baseline. Serum samples were used to estimate SARS-CoV-2 nucleoprotein-specific IgG [IgG (N)], spike protein-specific IgG [IgG (S)] and neutralizing antibodies (NAb). Results: Participants with previous SARS-CoV-2 infection after a single vaccine dose elicited IgG (N) and IgG (S) antibody levels along with NAb binding inhibition responses levels were similar to infection-naïve vaccinated participants who had taken two doses of vaccine. Interpretation & conclusions: Our preliminary data suggested that a single dose of BBV152-induced humoral immunity in previously infected individuals was equivalent to two doses of the vaccine in infection-naïve individuals. However, these findings need to be confirmed with large sized cohort studies.

Profile of patients with pulmonary non-tuberculous mycobacterial disease mimicking pulmonary tuberculosis.

INTRODUCTION: With the introduction of newer molecular diagnostic tools, an increasing number of Non-tuberculous Mycobacteria (NTM) affecting the respiratory system and mimicking symptoms of pulmonary tuberculosis (PTB) are being identified. They may be misdiagnosed and treated as PTB, often categorized as treatment failures if they do not respond to treatment. This manuscript aims to characterize patients with pulmonary NTM disease. METHODS: Patient characteristics of bacteriologically confirmed pulmonary NTM disease, attending the ICMR-National Institute for Research in Tuberculosis, Chennai were prospectively compiled over a two-year period (2017-2018). RESULTS: A total of 122 patients with recurrent chest symptoms and not responding to anti-tuberculosis treatment were screened for NTM. Thirty-nine cases (26 males and 13 females) of symptomatic pulmonary NTM were diagnosed. The mean (SD) patient age and body mass index were 48.6 ± 11 years and 16.3 ± 3. All male participants were smokers, had at least one episode of previous ATT. Mycobacterium kansasii (48.7%) was the most frequently isolated species followed by Mycobacterium intracellulare (20.5%), Mycobacterium abscessus (7.6%) followed by Mycobacterium avium, Mycobacterium fortuitum, Mycobacterium kyorinense, and Mycobacterium simiae. Infection with multiple NTMs was seen in four patients. Isoniazid resistance was identified in 20 patients. Based on species identified, treatment was initiated as per American Thoracic Society guidelines and continued up to 12 months of culture negativity. CONCLUSIONS: M. kansasii is the commonest pulmonary NTM isolated in Tamilnadu with a higher prevalence in males and elderly. Sensitization of both patients and providers is essential to avoid misdiagnosis and delay in diagnosis of pulmonary NTM disease as pulmonary TB.

Low Serum Concentrations of Rifampicin and Pyrazinamide Associated with Poor Treatment Outcomes in Children with Tuberculosis Related to HIV Status.

OBJECTIVES: To compare the pharmacokinetics of rifampicin (RMP), isoniazid (INH) and pyrazinamide (PZA) between HIV-infected and HIV-uninfected children with tuberculosis (TB) and correlate it with TB treatment outcome. METHODS: HIV-uninfected (n = 84) and HIV-infected (n = 77) children with TB receiving standard thrice weekly treatment were recruited from 6 hospitals in India. Semi-intensive pharmacokinetic sampling was performed during intensive phase of TB treatment after directly observed administration of drugs. Drug concentrations were measured by high performance liquid chromatography. INH acetylator status was determined, and nutritional assessment was done. Children were followed-up and treatment outcomes noted. RESULTS: Children with HIV and TB had significantly lower RMP peak concentration (Cmax) (2.6 vs. 5.1 µg/mL; P < 0.001) and exposure [area under the time-concentration curve (AUC0-8); 10.4 vs. 23.4 µg/mL h; P < 0.001] than those with TB. Among HIV-infected children, a significantly higher proportion had stunting (77% vs. 29%; P < 0.001) and underweight (73% vs. 38%; P < 0.001) compared with children with TB. Combining both groups, RMP Cmax (P = 0.001; adjusted odds ratio = 1.437; 95% confidence interval: 1.157-1.784) and PZA Cmax (P = 0.027; adjusted odds ratio = 1.041; 95% confidence interval: 1.005-1.079) significantly influenced treatment outcome. CONCLUSIONS: HIV infection was associated with lower Cmax of RMP and INH and AUC0-8 of RMP. Over 90% of children in both groups had subtherapeutic RMP Cmax. Cmax of RMP and PZA significantly influenced TB treatment outcome in children with TB. The findings have important clinical implications and suggest the need to increase anti-TB drug doses in children with HIV and TB.

Type 2 diabetes mellitus coincident with pulmonary or latent tuberculosis results in modulation of adipocytokines.

Type 2 diabetes mellitus (T2DM) is recognized as major risk factor for the progress of active pulmonary tuberculosis (PTB), although the mechanistic link between diabetes and tuberculosis remains poorly characterized. Moreover, the influence of poorly controlled diabetes on the baseline levels of adipocytokines in the context of tuberculosis has not been explored in detail. To characterize the influence of coexistent DM on adipocytokine levels in pulmonary or latent TB (LTB), we examined circulating levels of adipocytokines in the plasma of individuals with PTB-DM or LTB-DM and compared them with those without DM (PTB or LTB). PTB-DM or LTB-DM is characterized by diminished circulating levels of adiponectin and adipsin and/or heightened circulating levels of leptin, visfatin and PAI-1. In addition, adiponectin and adipsin exhibit a significant negative correlation, whereas leptin, visfatin and PAI-1 display a significant positive correlation with HbA1C levels and random blood glucose levels. Therefore, our data reveal that PTB-DM or LTB-DM is characterized by alterations in the systemic levels of adipocytokines, indicating that altered adipose tissue inflammation underlying Type 2 diabetes potentially contributes to pathogenesis of TB disease.

Pharmacokinetics of first-line antituberculosis drugs in HIV-infected children with tuberculosis treated with intermittent regimens in India.

The objective of this report was to study the pharmacokinetics of rifampin (RMP), isoniazid (INH), and pyrazinamide (PZA) in HIV-infected children with tuberculosis (TB) treated with a thrice-weekly anti-TB regimen in the government program in India. Seventy-seven HIV-infected children with TB aged 1 to 15 years from six hospitals in India were recruited. During the intensive phase of TB treatment with directly observed administration of the drugs, a complete pharmacokinetic study was performed. Drug concentrations were measured by high-performance liquid chromatography. A multivariable regression analysis was done to explore the factors impacting drug levels and treatment outcomes. The proportions of children with subnormal peak concentrations (Cmax) of RMP, INH, and PZA were 97%, 28%, and 33%, respectively. Children less than 5 years old had a lower median Cmax and lower exposure (area under the time-concentration curve from 0 to 8 h [AUC0-8]) of INH (Cmax, 2.5 versus 5.1 µg/ml, respectively [P=0.016]; AUC0-8, 11.1 versus 22.0 µg/ml·h, respectively [P=0.047[) and PZA (Cmax, 34.1 versus 42.3 µg/ml, respectively [P=0.055]; AUC0-8, 177.9 versus 221.7 µg/ml·h, respectively [P=0.05]) than those more than 5 years old. In children with unfavorable versus favorable outcomes, the median Cmax of RMP (1.0 versus 2.8 µg/ml, respectively; P=0.002) and PZA (31.9 versus 44.4 µg/ml, respectively; P=0.045) were significantly lower. Among all factors studied, the PZA Cmax influenced TB treatment outcome (P=0.011; adjusted odds ratio, 1.094; 95% confidence interval, 1.021 to 1.173). A high proportion of children with HIV and TB had a subnormal RMP Cmax. The PZA Cmax significantly influenced treatment outcome. These findings have important clinical implications and emphasize that drug doses in HIV-infected children with TB have to be optimized.

Effect of 1,25-dihydroxy vitamin D3 on cathelicidin expression in patients with and without cavitary tuberculosis.

1,25-dihydroxy vitamin D3 (1,25(OH)2D3) is a potent immuno-modulator which induces LL-37, the active peptide of cathelicidin, and restricts the growth of Mycobacterium tuberculosis (Mtb) in human macrophages. In the present study, we investigated the effect of 1,25(OH)2D3 on cathelicidin antimicrobial peptide (CAMP) expression in healthy controls (HCs) and pulmonary tuberculosis (PTB) patients. Peripheral blood mononuclear cells (PBMCs) from 50 HCs and 35 PTB patients were cultured for 72 h either with Mtb alone or Mtb with 1,25(OH)2D3 at 10(-7) M concentration. 1,25(OH)2D3 significantly up regulated the macrophage phagocytosis, CD14, CAMP gene expression and hCAP18 protein in HCs and PTB patients (p < 0.05). A significant positive correlation was observed between macrophage phagocytosis and CAMP gene expression in both the study groups (p < 0.05). Moreover, 1,25(OH)2D3 up regulated CAMP gene expression was more prominent in PTB patients without lung cavity (less severe form of disease) as compared to patients with cavitary TB (severe form of disease) (p < 0.05). The present study suggests that vitamin D may be used as an adjunct to anti-TB treatment and may be useful for a quicker recovery from less severe forms of TB disease.

1,25-Dihydroxy vitamin D3 downregulates pro-inflammatory cytokine response in pulmonary tuberculosis.

1,25-Dihydroxy vitamin D3 [1,25(OH)2D3] is a potent immunomodulator and regulates various immune responses to Mycobacterium tuberculosis (Mtb). The present study aimed to understand the effect of 1,25(OH)2D3 on pro-inflammatory cytokine response to Mtb antigen. Peripheral blood mononuclear cells from 42 healthy controls (HCs) and 42 pulmonary tuberculosis (PTB) patients were cultured with culture filtrate antigen (CFA) of Mtb with and without 1,25(OH)2D3 at 10(-7)M concentration for 72 h. The levels of IL-1α, IL-1ß, TNF-α, TNF-ß, IL-17 and IL-23 were estimated in the culture supernatants by ELISA. 1,25(OH)2D3 significantly suppressed all the CFA induced pro-inflammatory cytokines (p<0.05) studied except IL-1ß in both HCs and PTB patients. Among the PTB patients, the observed suppression was visible both in patients with and without cavitary tuberculosis. The present study results suggest that 1,25(OH)2D3 downregulates the production of pro-inflammatory cytokines and may control the exacerbated inflammatory response that may protect the host from excessive tissue damage at the site of infection.

Are we justified in treating for multidrug-resistant tuberculosis based on positive follow-up smear results?

SETTING: National Institute for Research in Tuberculosis, India. OBJECTIVE: To assess, among new culture-confirmed smear-positive pulmonary tuberculosis (TB) patients, the proportion of follow-up smear-positives that were culture-negative (S+C-) by month of follow-up examination, human immunodeficiency virus (HIV) status, pre-treatment drug susceptibility status and smear grading. DESIGN: We extracted follow-up smear (fluorescence microscopy) and culture (Löwenstein-Jensen) results of patients enrolled in clinical trials from January 2000 to August 2012 and treated with the WHO Category I regimen (2EHRZ3/4HR3). RESULTS: Of 520 patients, including 176 who were HIV-infected, respectively 199, 81, 47 and 43 were smear-positive at months 2, 4, 5 and 6; of these, respectively 138 (69%), 62 (75%), 32 (68%) and 27 (63%) were culture-negative. The S+C- phenomenon was more pronounced among '1+ positive' patients than in 2+ or 3+ positive patients and in 'pan-susceptible' patients than in those with any resistance, and did not vary by HIV status. CONCLUSION: Nearly two thirds of patients with follow-up smears positive at months 5 and 6 were culture-negative. Starting multidrug-resistant TB (MDR-TB) treatment empirically based on smear results, even in resource-limited settings, is incorrect and can have hazardous consequences. There is an urgent need to revisit the WHO recommendation concerning empirical MDR-TB treatment.

Number of sputum specimens during treatment follow-up of tuberculosis patients: two or one?

SETTING: National Institute for Research in Tuberculosis clinics in Chennai and Madurai, India. OBJECTIVE: To examine the pattern of serial smears (negative-negative [NN], negative-positive [NP], positive-negative [PN], positive-positive [PP]) during treatment follow-up of culture-confirmed new smear-positive tuberculosis (TB) patients, and the proportion of culture-negatives in each category. DESIGN: We reviewed the records and extracted follow-up smear (fluorescent microscopy) and culture (Löwenstein-Jensen) results of patients enrolled in clinical trials from January 2000 to August 2012 and treated with the Category I regimen (2EHRZ3/4HR3). Data entry and analysis were performed using EpiData. RESULTS: Among 520 patients (176 infected with the human immunodeficiency virus), the proportions of culture-negative patients with NN, discordant (PN or NP) and PP patterns were approximately 98%, 80% and 40%, respectively. The smear-positive culture-negative phenomenon was more frequent in follow-up smear results graded 1+, followed by 2+ and 3+. CONCLUSION: There is justification for discontinuing the examination of second specimens during treatment follow-up among TB patients. However, a positive result on the first smear needs to be confirmed by a second positive result before making clinical management decisions. The World Health Organization may need to reconsider its recommendation on this issue.

Effect of vitamin D3 on chemokine expression in pulmonary tuberculosis.

1,25 Dihydroxy vitamin D(3) (vitamin D(3)) is an immunomodulator and its deficiency has been associated with susceptibility to tuberculosis. We have studied the immunoregulatory role of vitamin D(3) on various chemokine expression in pulmonary tuberculosis. Peripheral blood mononuclear cells obtained from 21 pulmonary tuberculosis (PTB) patients and 24 healthy controls (HCs) were cultured for 48 h with culture filtrate antigen (CFA) of Mycobacterium tuberculosis with or without vitamin D(3) at a concentration 1 × 10(-7)M. The relative mRNA expression of monocyte chemoattractant protein-1 (MCP-1, CCL2), macrophage inflammatory protein-1α (MIP-1α, CCL3), macrophage inflammatory protein-1ß (MIP-1ß, CCL4), and regulated upon-activation, normal T cell-expressed and secreted (RANTES, CCL5) and IFN-γ inducible protein-10 (IP-10, CXCL10) chemokines were estimated from 48 h old macrophages using real-time polymerase chain reaction (RT-PCR). The culture supernatants were used to estimate the various chemokines including monokine induced by IFN-γ (MIG, CXCL9) levels using cytometric bead array. In HCs, vitamin D(3) significantly suppressed the MCP-1 mRNA expression of CFA stimulated cells (p=0.0027), while no such effect was observed in PTB patients. Vitamin D(3) showed no significant effect on MIP-1α, MIP-1ß and RANTES in both the study groups. The CFA induced IP-10 mRNA and protein expression was significantly suppressed by vitamin D(3) in both the study groups (p<0.05). A similar suppressive effect of vitamin D(3) was observed with MIG protein in healthy controls (p=0.0029) and a trend towards a suppression was observed in PTB patients. The suppressive effect of vitamin D(3) is more prominent in CXC chemokines rather than CC chemokines. This suggests that vitamin D(3) may down regulate the recruitment and activation of T-cells through CXC chemokines at the site of infection and may act as a potential anti-inflammatory agent.

Toll-like receptor and TIRAP gene polymorphisms in pulmonary tuberculosis patients of South India.

Toll-like receptors (TLRs) are pattern recognition receptors and play an important role in innate immunity. Changes in TLRs and signaling molecules that result from polymorphisms are often associated with susceptibility to various infectious diseases. In the present study, we investigated whether variants in the TLR-1 1805T/G (Ile602Ser), TLR-2 2258G/A (Arg753Gln), TLR-4 896A/G (Asp299Gly), TLR-4 1196C/T (Thr399Ile), TLR-6 745C/T (Ser249Pro), TIRAP 975C/T (Ser180Leu) genes and TLR-9 promoter region polymorphisms at positions -1237C/T and -1486C/T are associated with susceptibility or resistance to pulmonary tuberculosis (PTB). Genotyping of TLR and TIRAP gene polymorphisms was performed by polymerase chain reaction followed by restriction fragment length polymorphism method in 212 healthy control subjects (HCs) and 206 PTB patients. The allele and genotype frequencies of various TLR genes were not different between the HCs and PTB patients. However, the study is underpowered to detect minor associations. The frequency of T allele of TIRAP 975C/T (Ser180Leu) polymorphism was significantly increased among PTB patients as compared to HCs [p = 0.026; Odds ratio (OR) 1.49, 95% Confidence interval (CI) 1.049-2.22]. A trend towards an increased frequency of TT genotype of TIRAP 975C/T was also observed in PTB patients [p = 0.078, OR 3.10 95% CI (0.96-10.05)]. The present study suggests that T allele of TIRAP 975C/T polymorphism may be associated with susceptibility to pulmonary TB in south Indian population. Further study on the regulatory role of this polymorphism may be helpful to understand the innate immunity in TB.

Effect of vitamin D3 on phagocytic potential of macrophages with live Mycobacterium tuberculosis and lymphoproliferative response in pulmonary tuberculosis.

Immune responses are elicited through antigen presentation and recognition by macrophages and T-lymphocytes, respectively. The immunomodulatory effect of vitamin D(3) on macrophage phagocytic potential with live Mycobacterium tuberculosis, spontaneous and M. tuberculosis culture filtrate antigen induced lymphocyte responses were studied in pulmonary tuberculosis patients (PTBPs) ( n = 31) and normal healthy subjects (NHSs) ( n = 43). Vitamin D(3) at a concentration of 10(-7) M significantly enhanced the macrophage phagocytosis of live M. tuberculosis in normal subjects with low phagocytic potential (less than 10%) ( p = 0.015). No such increase was observed in PTBPs. Vitamin D(3) significantly decreased the spontaneous lymphoproliferative response ( p = 0.022) and increased the apoptosis of peripheral blood mononuclear cells in PTBPs ( p = 0.024). In normals, vitamin D(3) increased the spontaneous lymphoproliferative response. An inverse correlation between macrophage phagocytosis and spontaneous response was observed in NHSs, whereas a direct correlation was seen between vitamin D(3)-treated cells in normal subjects under in vitro condition. Vitamin D(3) decreased the M. tuberculosis culture filtrate antigen induced lymphocyte response significantly in normal subjects ( p = 0.0003), while it had no influence on the lymphocyte response in PTBPs. The present study suggests that exposure to vitamin D(3) increases the phagocytic potential and spontaneous lymphoproliferative response but brings down the antigen-induced response in normals. In tuberculosis, addition of vitamin D(3) has no significant effect on antigen-induced lymphoproliferative response. This may be due to the unresponsive nature of the cells to the action of vitamin D(3) by virtue of the disease, which renders them inactive.