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Visceral leishmaniasis (VL) caused by Leishmania donovani remains of public health concern in rural India. Those at risk of VL are also at risk of other neglected tropical diseases (NTDs) including soil transmitted helminths. Intestinal helminths are potent regulators of host immune responses sometimes mediated through cross-talk with gut microbiota. We evaluate a meta-taxonomic approach to determine the composition of prokaryotic and eukaryotic gut microflora using amplicon-based sequencing of 16S ribosomal RNA (16S rRNA) and 18S rRNA gene regions. The most abundant bacterial taxa identified in faecal samples from Bihar State India were Prevotella (37.1%), Faecalibacterium (11.3%), Escherichia-Shigella (9.1%), Alloprevotella (4.5%), Bacteroides (4.1%), Ruminococcaceae UCG-002 (1.6%), and Bifidobacterium (1.5%). Eukaryotic taxa identified (excluding plant genera) included Blastocystis (57.9%; Order: Stramenopiles), Dientamoeba (12.1%; Family: Tritrichomonadea), Pentatrichomonas (10.1%; Family: Trichomonodea), Entamoeba (3.5%; Family: Entamoebida), Ascaridida (0.8%; Family: Chromodorea; concordant with Ascaris by microscopy), Rhabditida (0.8%; Family: Chromodorea; concordant with Strongyloides), and Cyclophyllidea (0.2%; Order: Eucestoda; concordant with Hymenolepis). Overall alpha (Shannon’s, Faith’s and Pielou’s indices) and beta (Bray-Curtis dissimilarity statistic; weighted UniFrac distances) diversity of taxa did not differ significantly by age, sex, geographic subdistrict, or VL case (N = 23) versus endemic control (EC; N = 23) status. However, taxon-specific associations occurred: (i) Ruminococcaceae UCG- 014 and Gastranaerophilales_uncultured bacterium were enriched in EC compared to VL cases; (ii) Pentatrichomonas was more abundant in VL cases than in EC, whereas the reverse occurred for Entamoeba. Across the cohort, high Escherichia-Shigella was associated with reduced bacterial diversity, while high Blastocystis was associated with high bacterial diversity and low Escherichia-Shigella. Individuals with high Blastocystis had low Bacteroidaceae and high Clostridiales vadin BB60 whereas the reverse held true for low Blastocystis. This scoping study provides useful baseline data upon which to develop a broader analysis of pathogenic enteric microflora and their influence on gut microbial health and NTDs generally. [ABSTRACT FROM AUTHOR]

Tuberculin skin test (TST) is used most widely for the detection of latent tuberculosis infection (LTBI), even though evidences suggest that it could be underreporting the prevalence of LTBI particularly in high disease-burden settings. We have explored whether in vivo (TST) and in vitro (cell-proliferative) T cell responses to PPD can serve as complementary measures. In addition, we also probed whether in vitro T cell response to cell-membrane antigens (Mem) of Mycobacterium tuberculosis (MTB) can serve as a biomarker for LTBI. Study subjects comprised 43 healthcare workers (HCWs), and 9 smear-positive TB patients served as ‘disease control’. To measure proliferative T cell responses, 0.1 ml blood (diluted 1:10) was incubated (5 days) with test or control antigen. Cells were stained with fluorescent antibodies to T cell (CD3+/CD4+/CD8+) surface markers and, after fixation and permeabilization, to nuclear proliferation marker Ki67. Data was acquired on a flow cytometer. HCWs who had an intimate exposure to MTB showed significantly higher TST positivity (85%) than the rest (43%), notwithstanding their BCG vaccination status. The proliferative responses of CD4+ and CD8+ subsets of T cells were comparable. Sixty seven and 100% TST-negative HCWs, respectively, were positive for proliferative T cell response to PPD and MTBMem. Cumulative positivity (TST or in vitro) was 86% with PPD and 100% with MTBMem indicating complementarity of the two responses. As standalone in vitro assay, MTBMem provided a significantly higher positivity (95%) than PPD (67%). T cell responses of TB patients were ‘generally’ depressed, having implications for the development of immunological assays for ‘progressive’ LTBI. Altogether, these results demonstrate that in vivo and in vitro T cell responses to PPD are complementary and in vitro response to MTBMem can be developed as a highly sensitive biomarker for LTBI. [ABSTRACT FROM AUTHOR]