Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Fat, lazy and indifferent - profiling persistent drug-tolerant tubercule bacilli from patient sputa during treatment predicts drug efficacy

ABSTRACT: Amplified M.tb RNA derived from 15 subjects at multiple time intervals before and during chemotherapy (totalling 52 samples) was profiled alongside M.tb H37Rv RNA extracted from in vitro log phase bacilli (2 biological replicates hybridised in duplicate) as a standardised comparator. Amplified mycobacterial RNA (2 µg) were directly labelled with Cy3 fluorophore using the Universal Linkage System (ULS, Kreatech Diagnostics). Microarray hybridisations were conducted as previously described using an M.tb complex pan-genome microarray generated by the Bacterial Microarray Group at St. George’s (ArrayExpress accession number A-BUGS-41).

INSTRUMENT(S): NA

ORGANISM(S): Mycobacterium tuberculosis H37Rv

SUBMITTER: Simon Waddell

PROVIDER: E-MTAB-3872 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Profiling persistent tubercule bacilli from patient sputa during therapy predicts early drug efficacy.

Honeyborne Isobella I McHugh Timothy D TD Kuittinen Iitu I Cichonska Anna A Evangelopoulos Dimitrios D Ronacher Katharina K van Helden Paul D PD Gillespie Stephen H SH Fernandez-Reyes Delmiro D Walzl Gerhard G Rousu Juho J Butcher Philip D PD Waddell Simon J SJ

BMC medicine 20160407

<h4>Background</h4>New treatment options are needed to maintain and improve therapy for tuberculosis, which caused the death of 1.5 million people in 2013 despite potential for an 86 % treatment success rate. A greater understanding of Mycobacterium tuberculosis (M.tb) bacilli that persist through drug therapy will aid drug development programs. Predictive biomarkers for treatment efficacy are also a research priority.<h4>Methods and results</h4>Genome-wide transcriptional profiling was used to ...[more]

PMID: 27055815

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Project description:Understanding mechanisms of resistance to M. tuberculosis (M.tb) infection in humans could identify novel therapeutic strategies as it has for other infectious diseases. We hypothesized that monocytes from household contacts of pulmonary tuberculosis patients that failed to convert their tuberculin skin tests (TSTNEG) would respond differently to M.tb infection when compared to matched tuberculin skin test positive controls (TSTPOS). We obtained genome-wide transcriptional profiles of M.tb-infected peripheral blood monocytes from 10 TSTNEG and 18 TSTPOS Ugandans. We used Gene Set Enrichment Analysis and interaction networks to identify cellular processes associated with TST status. We discovered gene sets associated with histone deacetylases that were differentially expressed when comparing TSTPOS and TSTNEG subjects. We used small molecule inhibitors to demonstrate that histone deacetylase function is important for the pro-inflammatory response to M.tb infection in monocytes. Monocytes from individuals who appear to resist M.tb infection differentially activate pathways controlled by histone deacetylase in response to M.tb infection when compared to those who are susceptible to infection. These data identify a potential cellular mechanism underlying the clinical phenomenon of persistently negative tuberculin skin tests despite known exposure to an infectious contact. CD14+ monocytes from 18 TSTPOS and 10 TSTNEG subjects were isolated by positive selection. Depending on the yield, between 5E5 and 1E6 monocytes were plated in duplicate in a 24-well tissue culture plate and rested overnight at 37oC, 5% CO2. The next day, cells were mock-infected or infected with virulent M. tuberculosis strain H37Rv at a multiplicity of infection (MOI) 10:1. After a six hour incubation at 37C, 5% CO2, the mycobacteria were killed and total RNA harvested using TRIzol-LS (Invitrogen). TRIzol samples were stored at -80oC until all infections were completed. To minimize batch effects, equal numbers of TSTPOS and TSTNEG subjects were processed on each of six days and frozen aliquots of M.tb were used for infections. RNA was extracted using the RNeasy Mini Kit with on-column DNAse digestion. RNA was transcribed into complementary DNA, labeled, and hybridized onto Illumina HT-12 microarrays according to the manufacturerâs instructions. Probe intensities were quantified using Illumina iScan platform and processed using BeadArray without performing normalization or background correction.

Dataset Information

Fat, lazy and indifferent - profiling persistent drug-tolerant tubercule bacilli from patient sputa during treatment predicts drug efficacy

Publications

Profiling persistent tubercule bacilli from patient sputa during therapy predicts early drug efficacy.

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