Displaying publications 1 - 20 of 38 in total

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  1. Jumat MI, Sarmiento ME, Acosta A, Chin KL
    J Appl Microbiol, 2023 Jun 01;134(6).
    PMID: 37197901 DOI: 10.1093/jambio/lxad104
    Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), remains the leading cause of mortality due to infectious diseases, only surpassed in 2020 by COVID-19. Despite the development in diagnostics, therapeutics, and evaluation of new vaccines for TB, this infectious disease remains uncontrollable due to the emergence of multidrug-resistant (MDR) and extremely drug-resistant (XDR) TB, among other factors. The development in transcriptomics (RNomics) has enabled the study of gene expression in TB. It is considered that non-coding RNAs (ncRNAs) from host [microRNAs (miRNAs)] and Mtb [small RNAs (sRNAs)] are important elements in TB pathogenesis, immune resistance, and susceptibility. Many studies have shown the importance of host miRNAs in regulating immune response against Mtb via in vitro and in vivo mice models. The bacterial sRNAs play a major role in survival, adaptation, and virulence. Here, we review the characterization and function of host and bacteria ncRNAs in TB and their potential use in clinical applications as diagnostic, prognostic, and therapeutic biomarkers.
  2. 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.
  3. Chin KL, Anibarro L, Sarmiento ME, Acosta A
    Trop Med Infect Dis, 2023 Jan 28;8(2).
    PMID: 36828505 DOI: 10.3390/tropicalmed8020089
    Globally, it is estimated that one-quarter of the world's population is latently infected with Mycobacterium tuberculosis (Mtb), also known as latent tuberculosis infection (LTBI). Recently, this condition has been referred to as tuberculosis infection (TBI), considering the dynamic spectrum of the infection, as 5-10% of the latently infected population will develop active TB (ATB). The chances of TBI development increase due to close contact with index TB patients. The emergence of multidrug-resistant TB (MDR-TB) and the risk of development of latent MDR-TB has further complicated the situation. Detection of TBI is challenging as the infected individual does not present symptoms. Currently, there is no gold standard for TBI diagnosis, and the only screening tests are tuberculin skin test (TST) and interferon gamma release assays (IGRAs). However, these tests have several limitations, including the inability to differentiate between ATB and TBI, false-positive results in BCG-vaccinated individuals (only for TST), false-negative results in children, elderly, and immunocompromised patients, and the inability to predict the progression to ATB, among others. Thus, new host markers and Mtb-specific antigens are being tested to develop new diagnostic methods. Besides screening, TBI therapy is a key intervention for TB control. However, the long-course treatment and associated side effects result in non-adherence to the treatment. Additionally, the latent MDR strains are not susceptible to the current TBI treatments, which add an additional challenge. This review discusses the current situation of TBI, as well as the challenges and efforts involved in its control.
  4. 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.
  5. 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.

  6. 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 
  7. 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.
  8. 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.
  9. Chin KL, Sarmiento ME, Mustapha ZA, Jani J, Jamal NB, Stanis CS, et al.
    Tuberculosis (Edinb), 2020 12;125:102003.
    PMID: 33099253 DOI: 10.1016/j.tube.2020.102003
    Tuberculosis (TB) is the deadliest of infectious diseases. TB diagnosis, based on sputum microscopy, culture, and nucleic acid amplification tests (NAATs) to identify its main causative agent, Mycobacterium tuberculosis (MTB), remains challenging. The current available NAATs, endorsed by World Health Organization (WHO), can differentiate MTB from some MTB complex (MTBC) members. Using bioinformatics, we identified a single nucleotide polymorphism (SNP) in lprM (Rv1970) gene that differentiate MTB from other MTBC members. A forward mismatch amplification mutation assay (MAMA) primer was designed for the targeted mutation and was used in a semi-nested melt-MAMA qPCR (lprM-MAMA). Using the optimized protocol, lprM-MAMA was positive with all MTB reference and clinical strains, and negative with other MTBC members, non-tuberculous mycobacteria (NTM) and other non-mycobacterial (NM) reference strains. The limit of detection (LOD) of lprM-MAMA was 76.29 fg. Xpert® MTB/RIF (Xpert)-positive sputum samples were also positive by lprM-MAMA, except for samples classified as having "very low" bacterial load by Xpert. Xpert-negative sputum samples were also negative by lprM-MAMA. In conclusion, lprM-MAMA demonstrated to be a useful tool for specific MTB diagnosis. Further evaluation with higher number of reference strains, including NTM and NM; and sputum samples are required to determine its potential for clinical application.
  10. 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.
  11. 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.
  12. Mat Luwi NE, Kadir R, Mohamud R, A Garcia-Santana ML, Acevedo R, Sarmiento ME, et al.
    Int J Mycobacteriol, 2020 8 31;9(3):261-267.
    PMID: 32862158 DOI: 10.4103/ijmy.ijmy_82_20
    Background: Tuberculosis (TB) is the leading cause of mortality due to infectious diseases. The development of new generation vaccines against TB is of paramount importance for the control of the disease. In previous studies, liposomes obtained from lipids of Mycobacterium smegmatis (LMs) demonstrated their immunogenicity and protective capacity against Mycobacterium tuberculosis in mice. To characterize the immunomodulatory capacity of this experimental vaccine candidate, in the current study, the stimulatory capacity of LMs was determined on bone marrow-derived dendritic cells (BMDCs) from mice.

    Methods: LMs were obtained and incubated with mature BMDCs. The internalization of LMs by BMDCs was studied by confocal microscopy, and the LMs immune-stimulatory capacity was determined by the expression of surface molecules (CD86 and MHCII) and the cytokine production (interleukin [IL]-12, interferon-Υ, tumor necrosis factor-α, and IL-10) 24 h after exposure to LMs.

    Results: The interaction of LMs with BMDCs and its internalization was demonstrated as well as the immune activation of BMDCs, characterized by the increased expression of CD86 and the production of IL-12. The LMs internalization and immune activation of BMDCs were blocked in the presence of cytochalasin, filipin III and chlorpromazine, which demonstrated that internalization of LMs by BMDCs is a key process for the LMs induced immune activation of BMDCs.

    Conclusions: The results obtained support the further evaluation of LMs as a mycobacterial vaccine, adjuvant, and in immunotherapy.

  13. 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.
  14. Chin KL, Sarmiento ME, Alvarez-Cabrera N, Norazmi MN, Acosta A
    Eur J Clin Microbiol Infect Dis, 2020 May;39(5):799-826.
    PMID: 31853742 DOI: 10.1007/s10096-019-03771-0
    Currently, there is a trend of increasing incidence in pulmonary non-tuberculous mycobacterial infections (PNTM) together with a decrease in tuberculosis (TB) incidence, particularly in developed countries. The prevalence of PNTM in underdeveloped and developing countries remains unclear as there is still a lack of detection methods that could clearly diagnose PNTM applicable in these low-resource settings. Since non-tuberculous mycobacteria (NTM) are environmental pathogens, the vicinity favouring host-pathogen interactions is known as important predisposing factor for PNTM. The ongoing changes in world population, as well as socio-political and economic factors, are linked to the rise in the incidence of PNTM. Development is an important factor for the improvement of population well-being, but it has also been linked, in general, to detrimental environmental consequences, including the rise of emergent (usually neglected) infectious diseases, such as PNTM. The rise of neglected PNTM infections requires the expansion of the current efforts on the development of diagnostics, therapies and vaccines for mycobacterial diseases, which at present, are mainly focused on TB. This review discuss the current situation of PNTM and its predisposing factors, as well as the efforts and challenges for their control.
  15. 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
  16. Camacho F, Moreno E, Garcia-Alles LF, Chinea Santiago G, Gilleron M, Vasquez A, et al.
    Front Immunol, 2020;11:566710.
    PMID: 33162982 DOI: 10.3389/fimmu.2020.566710
    Lipids, glycolipids and lipopeptides derived from Mycobacterium tuberculosis (Mtb) are presented to T cells by monomorphic molecules known as CD1. This is the case of the Mtb-specific sulfoglycolipid Ac2SGL, which is presented by CD1b molecules and is recognized by T cells found in tuberculosis (TB) patients and in individuals with latent infections. Our group, using filamentous phage display technology, obtained two specific ligands against the CD1b-Ac2SGL complex: (i) a single chain T cell receptor (scTCR) from a human T cell clone recognizing the CD1b-AcSGL complex; and (ii) a light chain domain antibody (dAbκ11). Both ligands showed lower reactivity to a synthetic analog of Ac2SGL (SGL12), having a shorter acyl chain as compared to the natural antigen. Here we put forward the hypothesis that the CD1b endogenous spacer lipid (EnSpacer) plays an important role in the recognition of the CD1b-Ac2SGL complex by specific T cells. To support this hypothesis we combined: (a) molecular binding assays for both the scTCR and the dAbκ11 antibody domain against a small panel of synthetic Ac2SGL analogs having different acyl chains, (b) molecular modeling of the CD1b-Ac2SGL/EnSpacer complex, and (c) modeling of the interactions of this complex with the scTCR. Our results contribute to understand the mechanisms of lipid presentation by CD1b molecules and their interactions with T-cell receptors and other specific ligands, which may help to develop specific tools targeting Mtb infected cells for therapeutic and diagnostic applications.
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