Displaying publications 301 - 320 of 1901 in total

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  1. Inhibition of choline kinase as an antiamoebic approach in Entamoeba histolytica infection
    Teh ZH, Lim BH, See Too WC, Few LL
    Trop Biomed, 2023 Dec 01;40(4):430-438.
    PMID: 38308830 DOI: 10.47665/tb.40.4.008
    Entamoeba histolytica is the parasite responsible for amoebiasis, which can result in amoebic colitis or amoebic liver abscess. Metronidazole has been the conventional treatment for intestinal amoebiasis, but concerns regarding resistance have emerged due to the identification of resistance pathways in E. histolytica. This study investigates a novel anti-amoebic approach targeting the CDP-choline pathway. Inhibition studies were conducted using potential choline kinase (CK) inhibitors to inhibit the EhCK enzyme, and RNA interference was employed to knock down the EhCK gene. Km and Vmax of purified EhCK and hCKa2 proteins were determined by pyruvate kinase-lactate dehydrogenase (PK-LDH) coupled assay. The IC50 values for EhCK and hCKa2 were determined with several commercial CK inhibitors. Selected inhibitors were incubated with E. histolytica trophozoites for 48 hours to determine the EC50 for each inhibitor. Silencing of gene encoding EhCK was carried out using duplex siRNA and the gene expression level was measured by real-time qPCR. Based on the IC50 values, three of the inhibitors, namely CK37, flavopiridol and H-89 were more potent against EhCK than hCKa2. Trophozoites growth inhibition showed that only HDTAB, H-89 and control drug metronidazole could penetrate and induce cell death after 48-hour incubation. siRNA concentration of 10 µg/mL was used for the transfection of positive control GAPDH, EhCK, and non-targeting GFP siRNAs. RNAi experiment concluded with positive control GAPDH downregulated by 99% while the level of EhCK mRNA was downregulated by 47%. In this study, potential inhibitors of EhCK and siRNA have been identified, paving the way for further refinement and testing to enhance their potency against EhCK while sparing hCK. The utilization of these specific inhibitors and siRNA targeting EhCK represents a novel approach to impede the growth of E. histolytica by disrupting its phospholipid synthesis pathway.
    Matched MeSH terms: RNA, Small Interfering/metabolism
  2. Fulltext CD4/CD8 Ratio Recovery Among People Living With HIV Starting With First-Line Integrase Strand Transfer Inhibitors: A Prospective Regional Cohort Analysis
    Han WM, Avihingsanon A, Rajasuriar R, Tanuma J, Mundhe S, Lee MP, et al.
    J Acquir Immune Defic Syndr, 2023 Feb 01;92(2):180-188.
    PMID: 36625858 DOI: 10.1097/QAI.0000000000003121
    BACKGROUND: We evaluated trends in CD4/CD8 ratio among people living with HIV (PLWH) starting antiretroviral therapy (ART) with first-line integrase strand transfer inhibitors (INSTI) compared with non-INSTI-based ART, and the incidence of CD4/CD8 ratio normalization.

    METHODS: All PLWH enrolled in adult HIV cohorts of IeDEA Asia-Pacific who started with triple-ART with at least 1 CD4, CD8 (3-month window), and HIV-1 RNA measurement post-ART were included. CD4/CD8 ratio normalization was defined as a ratio ≥1. Longitudinal changes in CD4/CD8 ratio were analyzed by linear mixed model, the incidence of the normalization by Cox regression, and the differences in ratio recovery by group-based trajectory modeling.

    RESULTS: A total of 5529 PLWH were included; 80% male, median age 35 years (interquartile range [IQR], 29-43). First-line regimens were comprised of 65% NNRTI, 19% PI, and 16% INSTI. The baseline CD4/CD8 ratio was 0.19 (IQR, 0.09-0.33). PLWH starting with NNRTI- (P = 0.005) or PI-based ART (P = 0.030) had lower CD4/CD8 recovery over 5 years compared with INSTI. During 24,304 person-years of follow-up, 32% had CD4/CD8 ratio normalization. After adjusting for age, sex, baseline CD4, HIV-1 RNA, HCV, and year of ART initiation, PLWH started with INSTI had higher odds of achieving CD4/CD8 ratio normalization than NNRTI- (P < 0.001) or PI-based ART (P = 0.015). In group-based trajectory modeling analysis, INSTI was associated with greater odds of being in the higher ratio trajectory.

    CONCLUSIONS: INSTI use was associated with higher rates of CD4/CD8 ratio recovery and normalization in our cohort. These results emphasize the relative benefits of INSTI-based ART for immune restoration.

    Matched MeSH terms: RNA/therapeutic use
  3. Surveillance, isolation and genomic characterization of Pteropine orthoreovirus of probable bat origin among patients with acute respiratory infection in Malaysia
    Tee KK, Chan PQ, Loh AM, Singh S, Teo CH, Iyadorai T, et al.
    J Med Virol, 2023 Feb;95(2):e28520.
    PMID: 36691929 DOI: 10.1002/jmv.28520
    Pteropine orthoreovirus (PRV), an emerging bat-borne virus, has been linked to cases of acute respiratory infections (ARI) in humans. The prevalence, epidemiology and genomic diversity of PRV among ARI of unknown origin were studied. Among 632 urban outpatients tested negative for all known respiratory viruses, 2.2% were PRV-positive. Patients mainly presented with moderate to severe forms of cough, sore throat and muscle ache, but rarely with fever. Phylogenetic analysis revealed that over 90% of patients infected with the Melaka virus (MelV)-like PRV, while one patient infected with the Pulau virus previously found only in fruit bats. Human oral keratinocytes and nasopharyngeal epithelial cells were susceptible to clinical isolates of PRV, including the newly isolated MelV-like 12MYKLU1034. Whole genome sequence of 12MYKLU1034 using Nanopore technique revealed a novel reassortant strain. Evolutionary analysis of the global PRV strains suggests the continuous evolution of PRV through genetic reassortment among PRV strains circulating in human, bats and non-human primate hosts, creating a spectrum of reassortant lineages with complex evolutionary characteristics. In summary, the role of PRV as a common etiologic agent of ARI is evident. Continuous monitoring of PRV prevalence, pathogenicity and diversity among human and animal hosts is important to trace the emergence of novel reassortants.
    Matched MeSH terms: RNA, Viral/genetics
  4. Differentially Expressed Genes and Their Clinical Significance in Ischaemic Stroke: An In-Silico Study
    Appunni S, Rubens M, Ramamoorthy V, Sharma H, Singh AK, Swarup V, et al.
    Malays J Med Sci, 2020 Dec;27(6):53-67.
    PMID: 33447134 DOI: 10.21315/mjms2020.27.6.6
    Background: Ischaemic stroke (IS), a multifactorial neurological disorder, is mediated by interplay between genes and the environment and, thus, blood-based IS biomarkers are of significant clinical value. Therefore, this study aimed to find global differentially expressed genes (DEGs) in-silico, to identify key enriched genes via gene set enrichment analysis (GSEA) and to determine the clinical significance of these genes in IS.

    Methods: Microarray expression dataset GSE22255 was retrieved from the Gene Expression Omnibus (GEO) database. It includes messenger ribonucleic acid (mRNA) expression data for the peripheral blood mononuclear cells of 20 controls and 20 IS patients. The bioconductor-package 'affy' was used to calculate expression and a pairwise t-test was applied to screen DEGs (P < 0.01). Further, GSEA was used to determine the enrichment of DEGs specific to gene ontology (GO) annotations.

    Results: GSEA analysis revealed 21 genes to be significantly plausible gene markers, enriched in multiple pathways among all the DEGs (n = 881). Ten gene sets were found to be core enriched in specific GO annotations. JunD, NCX3 and fibroblast growth factor receptor 4 (FGFR4) were under-represented and glycoprotein M6-B (GPM6B) was persistently over-represented.

    Conclusion: The identified genes are either associated with the pathophysiology of IS or they affect post-IS neuronal regeneration, thereby influencing clinical outcome. These genes should, therefore, be evaluated for their utility as suitable markers for predicting IS in clinical scenarios.

    Matched MeSH terms: RNA; RNA, Messenger
  5. Fulltext Rapid-response RNA-fluorescence in situ hybridization (FISH) assay platform for coronavirus antiviral high-throughput screening
    Chan R, Mugisha CS, Chuenchob V, Moquin SA, Manjunatha UH, Jarrousse N, et al.
    SLAS Discov, 2024 Dec;29(8):100189.
    PMID: 39499968 DOI: 10.1016/j.slasd.2024.100189
    Over the past 25 years, the global community has faced challenges posed by three distinct outbreaks of coronaviruses including severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The identification of a novel alphacoronavirus canine CoV (CCoV-HuPn2018) in human patients in Malaysia underscores the potential for crossover infections to humans. The threat of the ever-evolving nature of viral infections as well as the lingering health and socioeconomic effects of the recent SARS-CoV-2 pandemic emphasize the urgent need for advanced antiviral drug screening tools that can be quickly implemented to strengthen preparedness and preventive measures against future outbreaks. Here, we present the development and validation of a novel RNA-fluorescence in situ hybridization (FISH) imaging assay as a 384-well, high-throughput rapid response platform for antiviral drug discovery. RNA-FISH is a powerful tool to visualize specific mRNA in cultured cells using a high-content imaging platform. The flexibility of RNA-FISH probe sets allows for the rapid design of viral genome-specific probes, enabling in vitro assay development to test for inhibition of viral replication by either biologic or small molecule inhibitors. Screening of 170 antiviral compounds in concentration-response demonstrates a strong correlation between the RNA-FISH assay and an immunofluorescence assay (IFA) for both human coronaviruses HCoV-OC43 and HCoV-229E. Additionally, we successfully applied this methodology in the context of CCoV strain 1-71, proving rapid development and deployment, opening new avenues for the evaluation of antiviral drugs to potential future emerging threats.
    Matched MeSH terms: RNA, Viral/genetics
  6. Fulltext Unraveling potential gene biomarkers for dengue infection through RNA sequencing
    Suppiah J, Md Sani SS, Hassan SS, Nadzar NIF, Ibrahim N', Thayan R, et al.
    Virus Genes, 2025 Feb;61(1):26-37.
    PMID: 39397194 DOI: 10.1007/s11262-024-02114-2
    Dengue virus hijacks host cell mechanisms and immune responses in order to replicate efficiently. The interaction between the host and the virus affects the host's gene expression, which remains largely unexplored. This pilot study aimed to profile the host transcriptome as a potential strategy for identifying specific biomarkers for dengue prediction and detection. High-throughput RNA sequencing (RNA-seq) was employed to generate host transcriptome profiles in 16 dengue patients and 10 healthy controls. Differentially expressed genes (DEGs) were identified in patients with severe dengue and those with dengue with warning signs compared to healthy individuals. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to elucidate the functions of upregulated and downregulated genes. Compared to healthy controls, 6466 genes were significantly differentially expressed (p RNA-sequencing holds promising implications and potential to augment existing dengue detection algorithms, contributing significantly to improved diagnostic and prognostic capabilities.
    Matched MeSH terms: Sequence Analysis, RNA*
  7. Association of Fusobacterium nucleatum infection with the clinicopathological characteristics in colorectal cancer patients
    Pang SW, Armon S, Chook JB, Chew J, Peh KB, Lim WW, et al.
    Mol Biol Rep, 2024 Jan 16;51(1):124.
    PMID: 38227097 DOI: 10.1007/s11033-023-09150-5
    BACKGROUND: Colorectal cancer (CRC) is a global health problem. The gut microbiome is now recognized as an important underlying factor to the initiation and progression of CRC. Fusobacterium nucleatum (FN) is one of the most studied bacteria in the aetiology of CRC. This study provided cohort evidence on the association of FN infection with clinicopathologic features in CRC patients.

    METHODS: We analysed the cancerous and adjacent non-cancerous formalin-fixed paraffin embedded (FFPE) tissue of 83 CRC patients from a single medical centre in Malaysia. TaqMan probe-based qPCR targeting the 16S rRNA gene was used to detect the presence of FN in the extracted FFPE DNA. The differences in FN expression between cancer and non-cancer tissues were evaluated. Association studies between FN infection in the tumour and relative FN abundance with available clinical data were conducted.

    RESULTS: FN was more abundant in the cancerous tissue compared to non-cancerous tissue (p = 0.0025). FN infection in the tumour was significantly associated with lymph node metastasis (p = 0.047) and cancer staging (p = 0.032), but not with other clinicopathologic variables. In double-positive patients where FN was detected in both cancerous and non-cancerous tissue, the expression fold-change of FN, calculated using 2-ΔΔCT formula, was significantly higher in patients with tumour size equal to or greater than 5 cm (p = 0.033) and in KRAS-mutated patients (p = 0.046).

    CONCLUSIONS: FN is enriched in CRC tumour tissue and is associated with tumour size, lymph node metastasis, cancer staging, and KRAS mutation in this single-centre small cohort study.

    Matched MeSH terms: RNA, Ribosomal, 16S/genetics
  8. The systematic status of Gonocephalus robinsonii Boulenger, 1908 (Squamata: Agamidae: Draconinae)
    Denzer W, Manthey U, Mahlow K, Böhme W
    Zootaxa, 2015;4039(1):129-44.
    PMID: 26624470 DOI: 10.11646/zootaxa.4039.1.5
    The generic assignment of the draconine lizard Gonocephalus robinsonii from the highlands of West-Malaysia has been uncertain since the original description. Here we present a study based on morphology, previously published karyotype data and molecular phylogenetics using 16S rRNA sequences to evaluate the systematic status of G. robinsonii. As a result we describe Malayodracon gen. nov. to accommodate the species.
    Matched MeSH terms: RNA, Ribosomal, 16S
  9. Polymerase chain reaction-based discrimination of viable from non-viable Mycoplasma gallisepticum
    Tan CG, Ideris A, Omar AR, Yii CP, Kleven SH
    Onderstepoort J Vet Res, 2014 09 02;81(1):e1-e7.
    PMID: 25686255 DOI: 10.4102/ojvr.v81i1.708
    The present study was based on the reverse transcription polymerase chain reaction (RT-PCR) of the 16S ribosomal nucleic acid (rRNA) of Mycoplasma for detection of viable Mycoplasma gallisepticum. To determine the stability of M. gallisepticum 16S rRNA in vitro, three inactivation methods were used and the suspensions were stored at different temperatures. The 16S rRNA of M. gallisepticum was detected up to approximately 20-25 h at 37 °C, 22-25 h at 16 °C, and 23-27 h at 4 °C. The test, therefore, could detect viable or recently dead M. gallisepticum (< 20 h). The RT-PCR method was applied during an in vivo study of drug efficacy under experimental conditions, where commercial broiler-breeder eggs were inoculated with M. gallisepticum into the yolk. Hatched chicks that had been inoculated in ovo were treated with Macrolide 1. The method was then applied in a flock of day 0 chicks with naturally acquired vertical transmission of M. gallisepticum, treated with Macrolide 2. Swabs of the respiratory tract were obtained for PCR and RT-PCR evaluations to determine the viability of M. gallisepticum. This study proved that the combination of both PCR and RT-PCR enables detection and differentiation of viable from non-viable M. gallisepticum.
    Matched MeSH terms: RNA, Bacterial/genetics; RNA, Ribosomal, 16S/genetics
  10. The complete mitogenome of the endangered freshwater crayfish Cherax tenuimanus (Smith 1912) (Crustacea: Decapoda: Parastacidae)
    Austin CM, Tan MH, Gan HY, Gan HM
    Mitochondrial DNA A DNA Mapp Seq Anal, 2016 11;27(6):4176-4177.
    PMID: 25630729
    Next-Gen sequencing was used to recover the complete mitochondrial genome of Cherax tenuimanus. The mitogenome consists of 15,797 base pairs (68.14% A + T content) containing 13 protein-coding genes, two ribosomal subunit genes, 22 transfer RNAs, and a 779 bp non-coding AT-rich region. Mitogenomes have now been recovered for all six species of Cherax native to Western Australia.
    Matched MeSH terms: RNA, Ribosomal/genetics; RNA, Transfer/genetics
  11. Fulltext Molecular phylogeny of Orthetrum dragonflies reveals cryptic species of Orthetrum pruinosum
    Yong HS, Lim PE, Tan J, Ng YF, Eamsobhana P, Suana IW
    Sci Rep, 2014 Jul 03;4:5553.
    PMID: 24989852 DOI: 10.1038/srep05553
    Dragonflies of the genus Orthetrum are members of the suborder Anisoptera, family Libellulidae. There are species pairs whose members are not easily separated from each other by morphological characters. In the present study, the DNA nucleotide sequences of mitochondrial and nuclear genes were employed to elucidate the phylogeny and systematics of Orthetrum dragonflies. Phylogenetic analyses could not resolve the various subfamilies of the family Libellulidae unequivocally. The nuclear 28S rRNA gene is highly conserved and could not resolve congeneric species of Orthetrum. Individual mitochondrial genes (COI, COII, and 16S rRNA) and combination of these genes as well as the nuclear ITS1&2 genes clearly differentiate morphologically similar species, such as the reddish species pairs O. chrysis and O. testaceum, and the bluish-coloured species O. glaucum and O. luzonicum. This study also reveals distinct genetic lineages between O. pruinosum schneideri (occurring in Malaysia) and O. pruinosum neglectum (occurring north of Peninsular Malaysia from India to Japan), indicating these taxa are cryptic species.
    Matched MeSH terms: RNA, Ribosomal, 16S/genetics; RNA, Ribosomal, 28S/genetics
  12. Fulltext Comparative transcriptome analysis of leaves and roots in response to sudden increase in salinity in Brassica napus by RNA-seq
    Yong HY, Zou Z, Kok EP, Kwan BH, Chow K, Nasu S, et al.
    Biomed Res Int, 2014;2014:467395.
    PMID: 25177691 DOI: 10.1155/2014/467395
    Amphidiploid species in the Brassicaceae family, such as Brassica napus, are more tolerant to environmental stress than their diploid ancestors.A relatively salt tolerant B. napus line, N119, identified in our previous study, was used. N119 maintained lower Na(+) content, and Na(+)/K(+) and Na(+)/Ca(2+) ratios in the leaves than a susceptible line. The transcriptome profiles of both the leaves and the roots 1 h and 12 h after stress were investigated. De novo assembly of individual transcriptome followed by sequence clustering yielded 161,537 nonredundant sequences. A total of 14,719 transcripts were differentially expressed in either organs at either time points. GO and KO enrichment analyses indicated that the same 49 GO terms and seven KO terms were, respectively, overrepresented in upregulated transcripts in both organs at 1 h after stress. Certain overrepresented GO term of genes upregulated at 1 h after stress in the leaves became overrepresented in genes downregulated at 12 h. A total of 582 transcription factors and 438 transporter genes were differentially regulated in both organs in response to salt shock. The transcriptome depicting gene network in the leaves and the roots regulated by salt shock provides valuable information on salt resistance genes for future application to crop improvement.
    Matched MeSH terms: Sequence Analysis, RNA/methods; RNA, Plant/genetics*
  13. Fulltext Pachyseris inattesa sp. n. (Cnidaria, Anthozoa, Scleractinia): a new reef coral species from the Red Sea and its phylogenetic relationships
    Terraneo TI, Berumen ML, Arrigoni R, Waheed Z, Bouwmeester J, Caragnano A, et al.
    Zookeys, 2014.
    PMID: 25152672 DOI: 10.3897/zookeys.433.8036
    A new scleractinian coral species, Pachyseris inattesa sp. n., is described from the Red Sea. Despite a superficial resemblance with some species in the agariciid genus Leptoseris with which it has been previously confused, P. inattesa sp. n. has micro-morphological characters typical of the genus Pachyseris. This genus, once part of the Agariciidae, is comprised of five extant species and is widely distributed throughout the tropical Indo-Pacific. It is currently incertae sedis as a result of recent molecular analysis and appears to be closely related to the Euphylliidae. A molecular phylogenetic reconstruction including P. inattesa sp. n., the genus type species P. rugosa, and P. speciosa, all present in the Red Sea, was performed using the mitochondrial intergenic spacer between COI and 16S-rRNA. The results confirm that P. inattesa sp. n. is a monophyletic lineage closely related to the other Pachyseris species examined.
    Matched MeSH terms: RNA, Ribosomal, 16S
  14. Fulltext Prevalence of unclassified bacteria in tropical coastal waters of malaysia revealed by metagenomic approach
    Chan KG, Chong TM
    Genome Announc, 2014;2(3).
    PMID: 24812226 DOI: 10.1128/genomeA.00419-14
    The metagenomes of marine prokaryotes from coastal seawaters in Malaysia are reported in this study. The investigation of the microbial communities using 16S rRNA gene amplicon metagenomic sequencing revealed that majority of the bacteria in the seawater samples remain unclassified.
    Matched MeSH terms: RNA, Ribosomal, 16S
  15. MORPHOLOGY AND MOLECULAR CHARACTERIZATION OF PSEUDO-NITZSCHIA (BACILLARIOPHYCEAE) FROM MALAYSIAN BORNEO, INCLUDING THE NEW SPECIES PSEUDO-NITZSCHIA CIRCUMPORA SP. NOV
    Lim HC, Leaw CP, Su SN, Teng ST, Usup G, Mohammad-Noor N, et al.
    J Phycol, 2012 Oct;48(5):1232-47.
    PMID: 27011282 DOI: 10.1111/j.1529-8817.2012.01213.x
    Field sampling was undertaken to investigate the occurrence of Pseudo-nitzschia Peragallo species in eight locations along the coast of Malaysian Borneo. A total of 108 strains of Pseudo-nitzschia species were isolated, and their morphology examined with SEM and TEM. Additionally, molecular data from nuclear-encoded partial LSU rDNA, and ITS regions, were characterized. A total of five species were confidently identified based on a combination of distinct morphological characteristics and supporting molecular evidence: P. brasiliana Lundholm, Hasle & Fryxell, P. cuspidata (Hasle) Hasle, P. dolorosa Lundholm & Moestrup, P. micropora Priisholm, Moestrup & Lundholm, and P. pungens (Grunow) Hasle var. pungens. However, one morphotype from Sarawak, while somewhat similar to P. caciantha, showed significant morphological distinction from this and any other of the currently described species. Most notably this morphotype possessed a characteristic pore arrangement in the poroids, with the fine pores in each perforation sector arranged in circles. Pair-wise sequence comparison of the LSU rDNA between this unidentified morphotype and P. caciantha Lundholm, Moestrup & Hasle, revealed 2.7% genetic divergence. Phylogenetic analyses strongly supported the monophyly of the morphotype. Based upon these supporting data it is here described as a new species, Pseudo-nitzschia circumpora sp. nov. A key to the six species of Pseudo-nitzschia from Malaysian Borneo is presented. Molecular signatures for all species were established based on structural comparisons of ITS2 rRNA transcripts.
    Matched MeSH terms: RNA, Ribosomal
  16. Molecular characterization of Giardia duodenalis isolated from Semai Pahang Orang Asli (Peninsular Malaysia aborigines)
    Mahdy AK, Surin J, Mohd-Adnan A, Wan KL, Lim YA
    Parasitology, 2009 Sep;136(11):1237-41.
    PMID: 19660153 DOI: 10.1017/S0031182009990527
    This study was conducted to determine the genotypes of Giardia duodenalis isolated from human faecal samples at Pos Betau, Pahang, Malaysia. Faecal specimens were collected and examined for G. duodenalis cysts using Trichrome staining techniques. Molecular identification was carried out by the amplification of a region of the small subunit of the nuclear ribosomal RNA (SSU rRNA) gene using nested PCR and subsequent sequencing. The sequences from 15 isolates from G. duodenalis were subjected to phylogenetic analysis (including appropriate outgroups) using the neighbor-joining and maximum parsimony methods. The trees identified G. duodenalis assemblages A and B, with a predominance of assemblage B. The predominance of anthroponotic genotypes indicates the possibility of anthroponotic transmission of these protozoa in this Semai Pahang Orang Asli community.
    Matched MeSH terms: RNA, Ribosomal/genetics; RNA, Protozoan/genetics
  17. Fulltext Getting the most out of RNA-seq data analysis
    Khang TF, Lau CY
    PeerJ, 2015;3:e1360.
    PMID: 26539333 DOI: 10.7717/peerj.1360
    Background. A common research goal in transcriptome projects is to find genes that are differentially expressed in different phenotype classes. Biologists might wish to validate such gene candidates experimentally, or use them for downstream systems biology analysis. Producing a coherent differential gene expression analysis from RNA-seq count data requires an understanding of how numerous sources of variation such as the replicate size, the hypothesized biological effect size, and the specific method for making differential expression calls interact. We believe an explicit demonstration of such interactions in real RNA-seq data sets is of practical interest to biologists. Results. Using two large public RNA-seq data sets-one representing strong, and another mild, biological effect size-we simulated different replicate size scenarios, and tested the performance of several commonly-used methods for calling differentially expressed genes in each of them. We found that, when biological effect size was mild, RNA-seq experiments should focus on experimental validation of differentially expressed gene candidates. Importantly, at least triplicates must be used, and the differentially expressed genes should be called using methods with high positive predictive value (PPV), such as NOISeq or GFOLD. In contrast, when biological effect size was strong, differentially expressed genes mined from unreplicated experiments using NOISeq, ASC and GFOLD had between 30 to 50% mean PPV, an increase of more than 30-fold compared to the cases of mild biological effect size. Among methods with good PPV performance, having triplicates or more substantially improved mean PPV to over 90% for GFOLD, 60% for DESeq2, 50% for NOISeq, and 30% for edgeR. At a replicate size of six, we found DESeq2 and edgeR to be reasonable methods for calling differentially expressed genes at systems level analysis, as their PPV and sensitivity trade-off were superior to the other methods'. Conclusion. When biological effect size is weak, systems level investigation is not possible using RNAseq data, and no meaningful result can be obtained in unreplicated experiments. Nonetheless, NOISeq or GFOLD may yield limited numbers of gene candidates with good validation potential, when triplicates or more are available. When biological effect size is strong, NOISeq and GFOLD are effective tools for detecting differentially expressed genes in unreplicated RNA-seq experiments for qPCR validation. When triplicates or more are available, GFOLD is a sharp tool for identifying high confidence differentially expressed genes for targeted qPCR validation; for downstream systems level analysis, combined results from DESeq2 and edgeR are useful.
    Matched MeSH terms: Sequence Analysis, RNA
  18. In vitro isolation and molecular identification of reptarenavirus in Malaysia
    Abba Y, Hassim H, Hamzah H, Ibrahim OE, Ilyasu Y, Bande F, et al.
    Virus Genes, 2016 Oct;52(5):640-50.
    PMID: 27142080 DOI: 10.1007/s11262-016-1345-7
    Boid inclusion body disease (BIBD) is a viral disease of boids caused by reptarenavirus. In this study, tissue from naturally infected boid snakes were homogenized and propagated in African Monkey kidney (Vero) and rat embryonic fibroblast (REF) cells. Virus replication was determined by the presence of cytopathic effect, while viral morphology was observed using transmission electron microscopy. Viral RNA was amplified using RT-PCR with primers specific for the L-segment of reptarenavirus; similarly, quantification of viral replication was done using qPCR at 24-144 h postinfection. Viral cytopathology was characterized by cell rounding and detachment in both Vero and REF cells. The viral morphology showed round-to-pleomorphic particles ranging from 105 to 150 nm which had sand-like granules. Sanger sequencing identified four closely associated reptarenavirus species from 15 (37.5 %) of the total samples tested, and these were named as follows: reptarenavirus UPM-MY 01, 02, 03, and 04. These isolates were phylogenetically closely related to the University Helsinki virus (UHV), Boa Arenavirus NL (ROUTV; BAV), and unidentified reptarenavirus L20 (URAV-L20). Comparison of deduced amino acid sequences further confirmed identities to L-protein of UHV, L-polymerase of BAV and RNA-dependent RNA polymerase of URAV-L20. Viral replication in Vero cells increased steadily from 24 to 72 h and peaked at 144 h. This is the first study in South East Asia to isolate and characterize reptarenavirus in boid snakes with BIBD.
    Matched MeSH terms: RNA Replicase/genetics; RNA, Viral/genetics
  19. Fulltext Molecular Characterization of Leptospira spp. in Environmental Samples from North-Eastern Malaysia Revealed a Pathogenic Strain, Leptospira alstonii
    Azali MA, Yean Yean C, Harun A, Aminuddin Baki NN, Ismail N
    J Trop Med, 2016;2016:2060241.
    PMID: 27127522 DOI: 10.1155/2016/2060241
    The presence of pathogenic Leptospira spp. in the environment poses threats to human health. The aim of this study was to detect and characterize Leptospira spp. from environmental samples. A total of 144 samples comprised of 72 soil and 72 water samples were collected from markets and recreational areas in a north-eastern state in Malaysia. Samples were cultured on Ellinghausen and McCullough modified by Johnson and Harris media. Leptospires were positive in 22.9% (n = 33) of the isolates. Based on partial sequences of 16S rRNA, a pathogenic leptospire, Leptospira alstonii (n = 1/33), was identified in 3% of the isolates followed by intermediate leptospire (L. wolffii, n = 1/33, and L. licerasiae, n = 7/33) and nonpathogenic leptospire, L. meyeri (n = 22/33) in 24.2% and 66.7%, respectively. This study demonstrates the presence of a clinically significant pathogenic L. alstonii in the environments which could pose health risks to the occupants and visitors.
    Matched MeSH terms: RNA, Ribosomal, 16S
  20. Fulltext Integrated pathway analysis of nasopharyngeal carcinoma implicates the axonemal dynein complex in the Malaysian cohort
    Chin YM, Tan LP, Abdul Aziz N, Mushiroda T, Kubo M, Mohd Kornain NK, et al.
    Int J Cancer, 2016 10 15;139(8):1731-9.
    PMID: 27236004 DOI: 10.1002/ijc.30207
    Nasopharyngeal carcinoma (NPC) is an epithelial squamous cell carcinoma on the mucosal lining of the nasopharynx. The etiology of NPC remains elusive despite many reported studies. Most studies employ a single platform approach, neglecting the cumulative influence of both the genome and transcriptome toward NPC development. We aim to employ an integrated pathway approach to identify dysregulated pathways linked to NPC. Our approach combines imputation NPC GWAS data from a Malaysian cohort as well as published expression data GSE12452 from both NPC and non-NPC nasopharynx tissues. Pathway association for GWAS data was performed using MAGENTA while for expression data, GSA-SNP was used with gene p values derived from differential expression values from GEO2R. Our study identified NPC association in the gene ontology (GO) axonemal dynein complex pathway (pGWAS-GSEA  = 1.98 × 10(-2) ; pExpr-GSEA  = 1.27 × 10(-24) ; pBonf-Combined  = 4.15 × 10(-21) ). This association was replicated in a separate cohort using gene expression data from NPC and non-NPC nasopharynx tissues (pAmpliSeq-GSEA  = 6.56 × 10(-4) ). Loss of function in the axonemal dynein complex causes impaired cilia function, leading to poor mucociliary clearance and subsequently upper or lower respiratory tract infection, the former of which includes the nasopharynx. Our approach illustrates the potential use of integrated pathway analysis in detecting gene sets involved in the development of NPC in the Malaysian cohort.
    Matched MeSH terms: RNA, Neoplasm/genetics; RNA, Neoplasm/isolation & purification; RNA, Neoplasm/metabolism
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