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Development of an immunochromatographic lateral flow dipstick for the detection of Mycobacterium tuberculosis 16 kDa antigen (Mtb-strip)

Mohd Amiruddin MN, Ang GY, Yu CY, Falero-Diaz G, Otero O, Reyes F, et al.

J Microbiol Methods, 2020 09;176:106003.
PMID: 32702386 DOI: 10.1016/j.mimet.2020.106003

Mycobacterium tuberculosis (Mtb) is a pathogenic bacterium that causes tuberculosis (TB). This contagious disease remains a severe health problem in the world. The disease is transmitted via inhalation of airborne droplets carrying Mtb from TB patients. Early detection of the disease is vital to prevent transmission of the infection to people in close contact with the patients. To date, there is a need of a simple, rapid, sensitive and specific diagnostic test for TB. Previous studies showed the potential of Mtb 16 kDa antigen (Ag16) in TB diagnosis. In this study, lateral flow immunoassay, also called simple strip immunoassay or immunochromatographic test (ICT) for detection of Ag16 was developed (Mtb-strip) and assessed as a potential rapid TB diagnosis method. A monoclonal antibody against Ag16 was optimized as the capturing and detection antibody on the Mtb-strip. Parameters affecting the performance of the Mtb-strip were also optimized before a complete prototype was developed. Analytical sensitivity showed that Mtb-strip was capable to detect as low as 125 ng of purified Ag16. The analytical sensitivity of Mtb-strip suggests its potential usefulness in different clinical applications.
Fulltext Mitochondrial DNA sequence of the horseshoe crab Tachypleus gigas

Sarmiento ME, Chin KL, Lau NS, Aziah I, Norazmi MN, Acosta A, et al.

Mitochondrial DNA B Resour, 2021 May 23;6(6):1710-1714.
PMID: 34104748 DOI: 10.1080/23802359.2021.1930213

This paper reports on the complete mitochondrial (mt) genome of a horseshoe crab, Tachypleus gigas (T. gigas), in Kuala Kemaman, Terengganu, Malaysia. Whole-genome sequencing of hemocyte DNA was performed with Illumina HiSeq system and the generated reads were de novo assembled with ABySS 2.1.5 and reassembled using mitoZ against Carcinoscorpius rotundicauda and Limulus polyphemus, resulting in a contig of 15 Kb. Phylogenetic analysis of the assembled mt genome suggests that the Tachypleus gigas is closely related to Tachypleus tridentatus than to Carcinoscorpius rotundicauda.
Comparative transcriptome profiling of horseshoe crab Tachypleus gigas hemocytes in response to lipopolysaccharides

Sarmiento ME, Chin KL, Lau NS, Aziah I, Ismail N, Norazmi MN, et al.

Fish Shellfish Immunol, 2021 Oct;117:148-156.
PMID: 34358702 DOI: 10.1016/j.fsi.2021.08.001

Horseshoe crabs (HSCs) are living fossil species of marine arthropods with a long evolutionary history spanning approximately 500 million years. Their survival is helped by their innate immune system that comprises cellular and humoral immune components to protect them against invading pathogens. To help understand the genetic mechanisms involved, the present study utilised the Illumina HiSeq platform to perform transcriptomic analysis of hemocytes from the HSC, Tachypleus gigas, that were challenged with lipopolysaccharides (LPS). The high-throughput sequencing resulted in 352,077,208 and 386,749,136 raw reads corresponding to 282,490,910 and 305,709,830 high-quality mappable reads for the control and LPS-treated hemocyte samples, respectively. Based on the log-fold change of > 0.3 or
Fulltext Transcriptomic Signature of Horseshoe Crab Carcinoscorpius rotundicauda Hemocytes' Response to Lipopolysaccharides

Sarmiento ME, Chin KL, Lau NS, Ismail N, Norazmi MN, Acosta A, et al.

Curr Issues Mol Biol, 2022 Nov 25;44(12):5866-5878.
PMID: 36547060 DOI: 10.3390/cimb44120399

Carcinoscorpius rotundicauda (C. rotundicauda) is one of the four species of horseshoe crabs (HSCs). The HSC hemocytes store defense molecules that are released upon encountering invading pathogens. The HSCs rely on this innate immunity to continue its existence as a living fossil for more than 480 million years. To gain insight into the innate mechanisms involved, transcriptomic analysis was performed on isolated C. rotundicauda hemocytes challenged with lipopolysaccharides (LPS), the main components of the outer cell membrane of gram-negative bacteria. RNA-sequencing with Illumina HiSeq platform resulted in 232,628,086 and 245,448,176 raw reads corresponding to 190,326,253 and 201,180,020 high-quality mappable reads from control and LPS-stimulated hemocytes, respectively. Following LPS-stimulation, 79 genes were significantly upregulated and 265 genes were downregulated. The differentially expressed genes (DEGs) were related to multiple immune functional categories and pathways such as those of the cytoskeleton, Toll and Imd, apoptosis, MAP kinase (MAPK), inositol phosphate metabolism, phagosome, leucocyte endothelial migration, and gram-negative bacterial infection, among others. This study provides important information about the mechanisms of response to LPS, which is relevant for the understanding the HSCs' immune response.
TCR-like domain antibody against Mycobacterium tuberculosis (Mtb) heat shock protein antigen presented by HLA-A*11 and HLA-A*24

Dass SA, Norazmi MN, Acosta A, Sarmiento ME, Tye GJ

Int J Biol Macromol, 2020 Jul 15;155:305-314.
PMID: 32240734 DOI: 10.1016/j.ijbiomac.2020.03.229

T cell receptor (TCR)-like antibodies, obtained with the use of phage display technology, sandwich the best of the both arms of the adaptive immune system. In this study, in vitro selections against the latency associated Mycobacterium tuberculosis (Mtb) heat shock protein 16 kDa antigen (16 kDa) presented by HLA-A*011 and HLA-A*24 were carried out with the use of a human domain phage antibody library. TCR-like domain antibodies (A11Ab and A24Ab) were successfully generated recognizing 16 kDa epitopes presented by HLA-A*011 and HLA-A*24 molecules respectively. Both antibodies were found to be functional in soluble form and exhibited strong binding capacity against its targets. The results obtained support the future evaluation of these ligands for the development of diagnostic and therapeutic tools for tuberculosis infection.
Engineered Mycobacterium tuberculosis antigen assembly into core-shell nanobeads for diagnosis of tuberculosis

Sheffee NS, Rubio-Reyes P, Mirabal M, Calero R, Carrillo-Calvet H, Chen S, et al.

Nanomedicine, 2021 06;34:102374.
PMID: 33675981 DOI: 10.1016/j.nano.2021.102374

Despite recent advances in diagnosis, tuberculosis (TB) remains one of the ten leading causes of death worldwide. Here, we engineered Mycobacterium tuberculosis (Mtb) proteins (ESAT6, CFP10, and MTB7.7) to self-assemble into core-shell nanobeads for enhanced TB diagnosis. Respective purified Mtb antigen-coated polyester beads were characterized and their functionality in TB diagnosis was tested in whole blood cytokine release assays. Sensitivity and specificity were studied in 11 pulmonary TB patients (PTB) and 26 healthy individuals composed of 14 Tuberculin Skin Test negative (TSTn) and 12 TST positive (TSTp). The production of 6 cytokines was determined (IFNγ, IP10, IL2, TNFα, CCL3, and CCL11). To differentiate PTB from healthy individuals (TSTp + TSTn), the best individual cytokines were IL2 and CCL11 (>80% sensitivity and specificity) and the best combination was IP10 + IL2 (>90% sensitivity and specificity). We describe an innovative approach using full-length antigens attached to biopolyester nanobeads enabling sensitive and specific detection of human TB.
Dry-reagent gold nanoparticle-based lateral flow biosensor for the simultaneous detection of Vibrio cholerae serogroups O1 and O139

Yu CY, Ang GY, Chua AL, Tan EH, Lee SY, Falero-Diaz G, et al.

J Microbiol Methods, 2011 Sep;86(3):277-82.
PMID: 21571011 DOI: 10.1016/j.mimet.2011.04.020

Cholera is a communicable disease caused by consumption of contaminated food and water. This potentially fatal intestinal infection is characterised by profuse secretion of rice watery stool that can rapidly lead to severe dehydration and shock, thus requiring treatment to be given immediately. Epidemic and pandemic cholera are exclusively associated with Vibrio cholerae serogroups O1 and O139. In light of the need for rapid diagnosis of cholera and to prevent spread of outbreaks, we have developed and evaluated a direct one-step lateral flow biosensor for the simultaneous detection of both V. cholerae O1 and O139 serogroups using alkaline peptone water culture. Serogroup specific monoclonal antibodies raised against lipopolysaccharides (LPS) were used to functionalize the colloidal gold nanoparticles for dual detection in the biosensor. The assay is based on immunochromatographic principle where antigen-antibody reaction would result in the accumulation of gold nanoparticles and thus, the appearance of a red line on the strip. The dry-reagent dipstick format of the biosensor ensure user-friendly application, rapid result that can be read with the naked eyes and cold-chain free storage that is well-suited to be performed at resource-limited settings.
Fulltext Sequence comparison of six human microRNAs genes between tuberculosis patients and healthy individuals

Amila A, Acosta A, Sarmiento ME, Suraiya S, Zafarina Z, Panneerchelvam S, et al.

Int J Mycobacteriol, 2015 Dec;4(4):341-6.
PMID: 26964819 DOI: 10.1016/j.ijmyco.2015.06.009

MicroRNAs (miRNAs) play an important role in diseases development. Therefore, human miRNAs may be able to inhibit the survival of Mycobacterium tuberculosis (Mtb) in the human host by targeting critical genes of the pathogen. Mutations within miRNAs can alter their target selection, thereby preventing them from inhibiting Mtb genes, thus increasing host susceptibility to the disease.
Fulltext Cellular and humoral immunogenicity of recombinant Mycobacterium smegmatis expressing Ag85B epitopes in mice

Kadir NA, Sarmiento ME, Acosta A, Norazmi MN

Int J Mycobacteriol, 2016 Mar;5(1):7-13.
PMID: 26927984 DOI: 10.1016/j.ijmyco.2015.09.006

The search for new vaccines more efficacious than bacille Calmette-Guérin for tuberculosis prevention is of paramount importance for the control of the disease. The expression of Mycobacterium tuberculosis antigens in Mycobacterium smegmatis is one of the current strategies for the development of new-generation vaccines against tuberculosis. The objective of this study was to evaluate the immunogenicity in mice of M. smegmatis expressing epitopes from Ag85B antigen.
Fulltext Vaccines for TB: Lessons from the Past Translating into Future Potentials

Tye GJ, Lew MH, Choong YS, Lim TS, Sarmiento ME, Acosta A, et al.

J Immunol Res, 2015;2015:916780.
PMID: 26146643 DOI: 10.1155/2015/916780

Development of vaccines for infectious diseases has come a long way with recent advancements in adjuvant developments and discovery of new antigens that are capable of eliciting strong immunological responses for sterile eradication of disease. Tuberculosis (TB) that kills nearly 2 million of the population every year is also one of the highlights of the recent developments. The availability or not of diagnostic methods for infection has implications for the control of the disease by the health systems but is not related to the immune surveillance, a phenomenon derived from the interaction between the bacteria and their host. Here, we will review the immunology of TB and current vaccine candidates for TB. Current strategies of developing new vaccines against TB will also be reviewed in order to further discuss new insights into immunotherapeutic approaches involving adjuvant and antigens combinations that might be of potential for the control of TB.
Fulltext Immunogenicity of recombinant Mycobacterium bovis bacille Calmette-Guèrin clones expressing T and B cell epitopes of Mycobacterium tuberculosis antigens

Mohamud R, Azlan M, Yero D, Alvarez N, Sarmiento ME, Acosta A, et al.

BMC Immunol, 2013;14 Suppl 1:S5.
PMID: 23458635 DOI: 10.1186/1471-2172-14-S1-S5

Recombinant Mycobacterium bovis bacille Calmette-Guèrin (rBCG) expressing three T cell epitopes of Mycobacterium tuberculosis (MTB) Ag85B antigen (P1, P2, P3) fused to the Mtb8.4 protein (rBCG018) or a combination of these antigens fused to B cell epitopes from ESAT-6, CFP-10 and MTP40 proteins (rBCG032) were used to immunize Balb/c mice. Total IgG responses were determined against Mtb8.4 antigen and ESAT-6 and CFP-10 B cell epitopes after immunization with rBCG032. Mice immunized with rBCG032 showed a significant increase in IgG1 and IgG2a antibodies against ESAT-6 and MTP40 (P1) B cell epitopes and IgG3 against both P1 and P2 B cell epitopes of MPT40. Splenocytes from mice immunized with rBCG018 proliferated against Ag85B P2 and P3 T cell epitopes and Mtb8.4 protein whereas those from mice-immunized with rBCG032 responded against all Ag85B epitopes and the ESAT-6 B cell epitope. CD4⁺ and CD8⁺ lymphocytes from mice immunized with rBCG018 produced primarily Th1 type cytokines in response to the T cell epitopes. Similar pattern of recognition against the T cell epitopes were obtained with rBCG032 with the additional recognition of ESAT-6, CFP-10 and one of the MTP40 B cell epitopes with the same pattern of cytokines. This study demonstrates that rBCG constructs expressing either T or T and B cell epitopes of MTB induced appropriate immunogenicity against MTB.
Fulltext COVID-19 Lethality in Sub-Saharan Africa and Helminth Immune Modulation

Fonte L, Acosta A, Sarmiento ME, Ginori M, García G, Norazmi MN

Front Immunol, 2020;11:574910.
PMID: 33117371 DOI: 10.3389/fimmu.2020.574910
Fulltext Does our Mycobacteriome Influence COVID-19 Morbidity and Lethality?

Acosta A, Fonte L, Sarmiento ME, Norazmi MN

Front Microbiol, 2021;12:589165.
PMID: 33613475 DOI: 10.3389/fmicb.2021.589165
Microbial biodiversity in the throats of pulmonary tuberculosis patients and tuberculin skin test (TST) positive and negative healthy individuals in Malaysia

Semail N, Suraiya S, Calero R, Mirabal M, Carrillo H, Ezzeddin Kamil MH, et al.

Tuberculosis (Edinb), 2020 09;124:101965.
PMID: 32692651 DOI: 10.1016/j.tube.2020.101965

The purpose of this study was to investigate the composition of throat microbiota in pulmonary tuberculosis patients (PTB) in comparison to healthy tuberculin skin test positive (TSTp) and negative (TSTn) individuals. Throat swabs samples were collected, and the microbiota was characterized. Richer operational taxonomic units (OTUs) were present in PTB group, compared to TSTp and TSTn. Regarding alpha diversity analysis there was a higher community diversity in TSTn compared to TSTp. Beta diversity analysis showed different species composition in TSTp compared to TSTn and PTB. There was higher presence of Firmicutes in PTB and TSTn compared to TSTp group at phylum level. At the genus level, Leuconostoc and Enterococcus were higher in TSTn compared to TSTp and Pediococcus, Chryseobacterium, Bifidobacterium, Butyrivibrio, and Bulleidia were higher in PTB compared to TSTn. Streptococcus was higher in TSTn compared to PTB and Lactobacillus in PTB compared to TSTp. At species level, Streptococcus sobrinus and Bulleidia moorei were higher in PTB compared to TSTn individuals, while Lactobacillus salivarius was higher in PTB compared to TSTp. The differences in the microbiome composition could influence the resistance/susceptibility to Mtb infection.
Fulltext Invasion and Metastasis Suppression by Anti-Neonatal Nav1.5 Antibodies in Breast Cancer

Sharudin NA, Murtadha Noor Din AH, Azahar II, Mohd Azlan M, Yaacob NS, Sarmiento ME, et al.

Asian Pac J Cancer Prev, 2022 Sep 01;23(9):2953-2964.
PMID: 36172657 DOI: 10.31557/APJCP.2022.23.9.2953

BACKGROUND: Detectable neonatal Nav1.5 (nNav1.5) expression in tumour breast tissue positive for lymph node metastasis and triple-negative subtype serves as a valid tumour-associated antigen to target and prevent breast cancer invasion and metastasis. Therapeutic antibodies against tumour antigens have become the predominant class of new drugs in cancer therapy because of their fewer adverse effects and high specificity.
OBJECTIVE: This study was designed to investigate the therapeutic and anti-metastatic potential of the two newly obtained anti-nNav1.5 antibodies, polyclonal anti-nNav1.5 (pAb-nNav1.5) and monoclonal anti-nNav1.5 (mAb-nNav1.5), on breast cancer invasion and metastasis.
METHODS: MDA-MB-231 and 4T1 cells were used as in vitro models to study the effect of pAb-nNav1.5 (59.2 µg/ml) and mAb-nNav1.5 (10 µg/ml) (24 hours treatment) on cell invasion. 4T1-induced mammary tumours in BALB/c female mice were used as an in vivo model to study the effect of a single dose of intravenous pAb-nNav1.5 (1 mg/ml) and mAb-nNav1.5 (1 mg/ml) on the occurrence of metastasis. Real-time PCR and immunofluorescence staining were conducted to assess the effect of antibody treatment on nNav1.5 mRNA and protein expression, respectively. The animals' body weight, organs, lesions, and tumour mass were also measured and compared.
RESULTS: pAb-nNav1.5 and mAb-nNav1.5 treatments effectively suppressed the invasion of MDA-MB-231 and 4T1 cells in the 3D spheroid invasion assay. Both antibodies significantly reduced nNav1.5 gene and protein expression in these cell lines. Treatment with pAb-nNav1.5 and mAb-nNav1.5 successfully reduced mammary tumour tissue size and mass and prevented lesions in vital organs of the mammary tumour animal model whilst maintaining the animal's healthy weight. mRNA expression of nNav1.5 in mammary tumour tissues was only reduced by mAb-nNav1.5.
CONCLUSION: Overall, this work verifies the uniqueness of targeting nNav1.5 in breast cancer invasion and metastasis prevention, but more importantly, humanised versions of mAb-nNav1.5 may be valuable passive immunotherapeutic agents to target nNav1.5 in breast cancer.
Fulltext Comparative study of IgA VH 3 gene usage in healthy TST(-) and TST(+) population exposed to tuberculosis: deep sequencing analysis

Chin ST, Ignatius J, Suraiya S, Tye GJ, Sarmiento ME, Acosta A, et al.

Immunology, 2015 Feb;144(2):302-11.
PMID: 25158076 DOI: 10.1111/imm.12372

The acquired immune response against tuberculosis is commonly associated with T-cell responses with little known about the role of B cells or antibodies. There have been suggestions that B cells and humoral immunity can modulate the immune response to Mycobacterium tuberculosis. However, the mechanisms involving B-cell responses in M. tuberculosis are not fully understood, in particular the antibody gene preferences. We hypothesized that a preferential use of V genes can be seen associated with resistance to infection mainly in the IgA isotype, which is of prominent importance for infection by pathogens via the mucosal route. We studied healthy individuals with long-term exposure to tuberculosis, infected (TST(+) ) and uninfected TST(-) ) with M. tuberculosis. From a total of 22 V genes analysed, the TST(-) population preferred the VH 3-23 and Vκ1 genes. The VH 3-23 genes were subsequently subjected to 454 amplicon sequencing. The TST(-) population showed a higher frequency of the D3-10 segment compared with the D3-22 segment for the TST(+) population. The J segment usage pattern was similar for both populations with J4 segment being used the most. A preferential pairing of J4 segments to D3-3 was seen for the TST(-) population. The antibodyome difference between both populations suggests a preference for antibodies with VH 3-23, D3-3, JH 4 gene usage by the TST(-) population that could be associated with resistance to infection with M. tuberculosis.
Immune TB Antibody Phage Display Library as a Tool To Study B Cell Immunity in TB Infections

Hamidon NH, Suraiya S, Sarmiento ME, Acosta A, Norazmi MN, Lim TS

Appl Biochem Biotechnol, 2018 Mar;184(3):852-868.
PMID: 28884285 DOI: 10.1007/s12010-017-2582-5

B cells and in particular antibodies has always played second fiddle to cellular immunity in regard to tuberculosis (TB). However, recent studies has helped position humoral immunity especially antibodies back into the foray in relation to TB immunity. Therefore, the ability to correlate the natural antibody responses of infected individuals toward TB antigens would help strengthen this concept. Phage display is an intriguing approach that can be utilized to study antibody-mediated responses against a particular infection via harvesting the B cell repertoire from infected individuals. The development of disease-specific antibody libraries or immune libraries is useful to better understand antibody-mediated immune responses against specific disease antigens. This study describes the generation of an immune single-chain variable fragment (scFv) library derived from TB-infected individuals. The immune library with an estimated diversity of 109 independent clones was then applied for the identification of monoclonal antibodies against Mycobacterium tuberculosis α-crystalline as a model antigen. Biopanning of the library isolated three monoclonal antibodies with unique gene usage. This strengthens the role of antibodies in TB immunity in addition to the role played by cellular immunity. The developed library can be applied against other TB antigens and aid antibody-derived TB immunity studies in the future.
Parasitic infections in Malaysian aborigines with pulmonary tuberculosis: a comparative cross-sectional study

Wong WK, Mohd-Nor N, Noordin R, Foo PC, Mohamed Z, Haq JA, et al.

Parasitol Res, 2019 Sep;118(9):2635-2642.
PMID: 31363922 DOI: 10.1007/s00436-019-06406-7

The geographical distribution of tuberculosis (TB) overlaps with various parasitic infections. Uncovering the characteristics of coinfecting parasites that potentially affect the host susceptibility to TB is pertinent as it may provide input to current TB therapeutic and prophylactic measures. The present study was aimed at examining the types of parasitic infections in TB patients and healthy TB contacts (HC) in Orang Asli, Malaysian aborigines, who dwelled in the co-endemic areas. Stool and serum samples were collected from Orang Asli who fulfilled the selection criteria and provided written informed consents. Selected parasitic infections in the two study groups were determined by stool examination and commercial serum antibody immunoassays. The prevalence of parasitic infections in TB and HC participants were 100% (n = 82) and 94.6% (n = 55) respectively. The parasitic infections comprised toxocariasis, trichuriasis, amoebiasis, toxoplasmosis, hookworm infection, ascariasis, strongyloidiasis, and brugian filariasis, in decreasing order of prevalence. Overall, helminth or protozoa infection did not show any significant association with the study groups. However, when the species of the parasite was considered, individuals exposed to trichuriasis and toxoplasmosis showed significant odds reduction (odds ratio (OR) 0.338; 95% confidence interval (CI) 0.166, 0.688) and odds increment (OR 2.193; 95% CI 1.051, 4.576) to have active pulmonary TB, respectively. In conclusion, trichuriasis and toxoplasmosis may have distinct negative and positive associations respectively with the increase of host susceptibility to TB.
Interactions of domain antibody (dAbκ11) with Mycobacterium tuberculosis Ac2SGL in complex with CD1b

Law CT, Camacho F, Garcia-Alles LF, Gilleron M, Sarmiento ME, Norazmi MN, et al.

Tuberculosis (Edinb), 2019 01;114:9-16.
PMID: 30711162 DOI: 10.1016/j.tube.2018.11.002

Tuberculosis (TB) is the main cause of mortality among all infectious diseases. The presentation of lipids by CD1b molecules and the interactions of the CD1b-lipid complexes with the immune receptors are important for the understanding of the immune response to Mycobacterium tuberculosis (Mtb), and to develop TB control methods. A specific domain antibody (dAbk11) recognizing the complex of CD1b with Mtb sulphoglycolipid (Ac2SGL) had been previously developed. In order to study the interactions of dAbk11 with Ac2SGL:CD1b, the conformation of Ac2SGL within CD1b was first modelled. The orientation of dAbκ11 with Ac2SGL:CD1b was then predicted by a docking experiment and the complex was sampled using molecular dynamics simulation. Data showed that dAbκ11 Tyr32 OH plays a decisive role in interacting with Ac2SGL alkyl tail HO17. The binding free energy calculation showed that Ac2SGL establish strong hydrophobic interactions with dAbκ11. The model also predicted a higher affinity for the natural sulfoglycolipid (Ac2SGL) than the synthetic analogue (SGL12), which was supported by the ELISA data. These results shed light on the likely mechanism of interactions between Ac2SGL:CD1b and dAbκ11, thus making possible to envision the strategies for dAbκ11 optimization for possible future applications.
Small RNA datasets of drug-susceptible Mycobacterium tuberculosis strains from Sabah, Malaysia

Jumat MI, Jani J, Mustapha ZA, Rodrigues KF, Azizan N, Acosta A, et al.

Data Brief, 2023 Feb;46:108795.
PMID: 36483477 DOI: 10.1016/j.dib.2022.108795

These datasets present a list of small RNAs from three drug-susceptible Mycobacterium tuberculosis strains isolated from Sabah, Malaysia. Sputum samples were obtained from three tuberculosis patients belonging to different districts. The bacteria were detected using GeneXpert MTB/RIF, isolated and cultured in BACTECTM MGITTM 320, and tested for their drug susceptibility. Total RNAs were extracted, sequenced, and analyzed using bioinformatic tools to filter out small RNA present in the Mycobacterium tuberculosis strains. Small RNA sequencing generated total raw reads of 63,252,209, 63,636,812, and 61,148,224 and total trimmed reads (15-30 nucleotides) of 51,533,188, 53,520,197, and 51,363,772 for Mycobacterium tuberculosis strain SBH49, SBH149, and SBH372, respectively. The raw data were submitted to the Sequence Read Archive (SRA) database of the National Center for Biotechnology Information (NCBI) under the accession numbers of SRX16744291 (SBH49), SRX16744292 (SBH149), and SRX16744293 (SBH372). Small RNAs play important roles in cellular processes such as cell differentiation, cell signaling, development of resistance to antibiotics and immune response, and metabolism regulation. The small RNAs determined here could provide further insights into various cellular processes crucial for Mycobacterium tuberculosis survivability and a better understanding of their gene regulation which ultimately opens a new pathway for combating tuberculosis infection.

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