Research Articles (Genetics)

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Browsing Research Articles (Genetics) by Subject "Amplicon-based RNA sequencing"

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    Distinct host-immune response toward species related intracellular mycobacterial killing : a transcriptomic study
    (Taylor & Francis, 2020) Madhvi, Abhilasha; Mishra, Hridesh; Chegoua, Novel N.; Tromp, Gerard; Van Heerden, Carel J.; Pietersen, R. D.; Leisching, Gina; Baker, Bienyameen
    The comparison of the host immune response when challenged with pathogenic and nonpatho- genic species of mycobacteria can provide answers to the unresolved question of how pathogens subvert or inhibit an effective response. We infected human monocyte derived macrophages (hMDMs) with different species of mycobacteria, in increasing order of pathogenicity, i.e. M. smegmatis, M. bovis BCG, and M. tuberculosis R179 that had been cultured in the absence of detergents. RNA was isolated post-infection and transcriptomic analysis using amplicons (Ampliseq) revealed 274 differentially expressed genes (DEGs) across three species, out of which we selected 19 DEGs for further validation. We used qRT-PCR to confirm the differential expression of 19 DEGs. We studied biological network through Ingenuity Pathway Analysis® (IPA) which revealed up-regulated pathways of the interferon and interleukin family related to the killing of M. smegmatis. Apart from interferon and interleukin family, we found one up-regulated (EIF2AK2) and two down-regulated (MT1A and TRIB3) genes as unique potential targets found by Ampliseq and qRT-PCR which may be involved in the intracellular mycobacterial killing. The roles of these genes have not previously been described in tuberculosis. Multiplex ELISA of culture supernatants showed increased host immune response toward M. smegmatis as compared to M. bovis BCG and M.tb R179. These results enhance our understanding of host immune response against M.tb infection.