Transcriptome analysis of mycobacteria in sputum samples of pulmonary tuberculosis patients

Abstract

Pulmonary tuberculosis, the disease caused by Mycobacterium tuberculosis, still retains a top rank among the deadliest communicable diseases. Sputum expectorated during the disease continues to be a primary diagnostic specimen and also serves as a reservoir of bacteria. The expression pattern of mycobacteria in sputum will lead to an insight into bacterial adaptation at the most highly transmissible stage of infection and can also help in identifying newer diagnostic as well as drug targets. Thus, in the present study, a whole genome microarray of Mycobacterium tuberculosis was used to elucidate the transcriptional profile of mycobacteria in the sputum samples of smear positive pulmonary tuberculosis patients. Overall, the mycobacteria in sputum appeared to be in a low energy and low replicative state as compared to in vitro grown log phase M. tb with downregulation of genes involved in ATP synthesis, aerobic respiration and translational machinery. Simultaneously, downregulation was also seen in the genes involved in secretion machinery of mycobacteria along with the downregulation of genes involved in the synthesis of phthiocerol dimycocerosate and phenol glycolipids. In contrast, the majority of the genes which showed an upregulation in sputum mycobacteria were of unknown function. Further identification of these genes may provide new insights into the mycobacterial behavior during this phase of infection and may help in deciphering candidates for development of better diagnostic and drug candidates.

The transcriptome of Mycobacterium tuberculosis in a lipid-rich dormancy model through RNAseq analysis

“…Sputum becomes in turn the microenvironment where the transmissible stage of Mtb resides, whose microscopic phenotype and transcriptional profile suggested there is a shift in the lipid metabolism. Mtb recovered from sputum showed cytoplasmic lipid bodies revealed with auramine-nile red staining, significant overexpression of the large Kst-R regulon of cholesterol catabolism, overpression of tgs1 gene required for triacylglycerol synthesis and down-regulation of genes involved in phthiocerol dimycocerosate and phenol glycolipid synthesis indicating cell wall remodeling 23 , 24 . There is also evidence that hypercholesterolemia increases susceptibility to TB in mice 25 and a survey in Singapore found positive correlation between a cholesterol diet and an increased risk to active TB in Chinese population 26 .…”

Transcriptional Profile of Mycobacterium tuberculosis in an in vitro Model of Intraocular Tuberculosis

“…Each functional group contained a different number of upregulated and downregulated transcripts (Figure 7D ). The hypergeometric test ( p < 0.05) was applied to determine the functional categories that were enriched in differentially expressed transcripts (Figure 7D ) (Leong and Kipling, 2009 ; Qureshi and Sacan, 2013 ; Sharma et al, 2017 ). The annotated list of identified M. tuberculosis transcripts with a potential role in invasion, transcriptional mechanism, environmental adaptation, and survival inside RPE cells at 3-dpi has been provided in Table 2 according to their functional categories as sorted by the TubercuList database ( http://tuberculist.epfl.ch ).…”

“…Among these genes, four genes ( Rv1971, Rv0169, Rv0592 , and Rv1967 ) belong to mammalian cell entry (mce) family protein that plays a crucial role in invasion and virulence of M. tuberculosis (Zhang and Xie, 2011 ). Recently, Rv1971 from this family was also reported to be upregulated in sputum samples of pulmonary TB patients (Sharma et al, 2017 ) and bioinformatically observed to be a highly antigenic protein involved in immune responses (Zhang et al, 2009 ). Interestingly, this gene was also detected in human IOTB samples (Table 3 ).…”

“…For each sample, from FE software, QC metric sets, thresholds, and charting tools were determined, and the resulting text files (.txt) of the control and experimental samples were analyzed for data normalization, statistical significance, and gene expression analysis using Agilent Technologies GeneSpring® GX software (version 14.9). Significantly modulated genes were selected based on Benjamini–Hochberg procedure (Benjamini and Hochberg, 1995 , 2000 ; Keselman et al, 2002 ; Pawitan et al, 2005 ) with a fold change >2 and p < 0.05, based on previous microarray studies with M. tuberculosis (Sharma et al, 2017 ).…”

<b>Sequenciamento, anotação e análise do genoma completo de <i>Mycobacterium bovis</i> cepa SP38</b>

“…Acredita-se que essas famílias de parálogos sejam importantes fatores de virulência, atuando na evasão do sistema imune por meio da variabilidade genética (FISHBEIN et al, 2015;PHELAN et al, 2016). Em seguida destacamse os genes envolvidos em funções de housekeeping, como produção e conservação de energia (5,23%), citados como fundamentais para o momento da infecção no hospedeiro em análises de transcriptoma (SHARMA et al, 2017), transporte e metabolismo de lipídeos (4,62%), que contém genes essenciais para a sobrevivência nos macrófagos (FISHER;PLIKAYTIS;SHINNICK, 2002;RUSSELL et al, 2010), e transcrição (4,62%), que são primordiais para manutenção bacteriana no hospedeiro (RAMAN et al, 2004).…”