Browsing by Author "Chonticha Klungthong"

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    Application of one-step reverse transcription droplet digital pcr for dengue virus detection and quantification in clinical specimens
    (2021-04-01) Dumrong Mairiang; Adisak Songjaeng; Prachya Hansuealueang; Yuwares Malila; Paphavee Lertsethtakarn; Sasikorn Silapong; Yongyuth Poolpanichupatam; Chonticha Klungthong; Kwanrutai Chin-Inmanu; Somchai Thiemmeca; Nattaya Tangthawornchaikul; Kanokwan Sriraksa; Wannee Limpitikul; Sirijitt Vasanawathana; Damon W. Ellison; Prida Malasit; Prapat Suriyaphol; Panisadee Avirutnan; Siriraj Hospital; Songkhla Hospital; Armed Forces Research Institute of Medical Sciences, Thailand; Khon Kaen Regional Hospital; Thailand National Center for Genetic Engineering and Biotechnology
    Detection and quantification of viruses in laboratory and clinical samples are standard assays in dengue virus (DENV) studies. The quantitative reverse transcription polymerase chain reaction (qRT-PCR) is considered to be the standard for DENV detection and quantification due to its high sensitivity. However, qRT-PCR offers only quantification relative to a standard curve and consists of several “in-house” components resulting in interlaboratory variations. We developed and optimized a protocol for applying one-step RT-droplet digital PCR (RT-ddPCR) for DENV detection and quantification. The lower limit of detection (LLOD95) and the lower limit of quantification (LLOQ) for RT-ddPCR were estimated to be 1.851 log10-copies/reaction and 2.337 log10-copies/reaction, respectively. The sensitivity of RT-ddPCR was found to be superior to qRT-PCR (94.87% vs. 90.38%, p = 0.039) while no false positives were detected. Quantification of DENV in clinical samples was independently performed in three laboratories showing interlaboratory variations with biases <0.5 log10-copies/mL. The RT-ddPCR protocol presented here could help harmonize DENV quantifi cation results and improve findings in the field such as identifying a DENV titer threshold correlating with disease severity.
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    Clinical isolates of dengue virus with distinctive susceptibility to nitric oxide radical induce differential gene responses in THP-1 cells
    (2008-07-05) Sukathida Ubol; Takol Chareonsirisuthigul; Jitra Kasisith; Chonticha Klungthong; Mahidol University; Armed Forces Research Institute of Medical Sciences, Thailand
    In the present study, 10 clinical isolates of dengue virus were selected according to their susceptibility to the inhibitory effect of nitric oxide radical, NO. Five of them are nitric oxide-susceptible viruses while the other five are nitric oxide-resistant viruses. These isolates were investigated to identify genetic factors that are responsible for the different phenotypes. Due to the evidence showing that NO suppresses DENV RNA polymerase activity, we, therefore, hypothesized that the RdRp domain of NS5 may responsible for NO inhibition. To answer this question, sequences of NS5 gene of NO-susceptible viruses and NO-resistant viruses were compared. We found that these two groups of viruses contain different amino acid sequence at position 621 to 646 in the active site of NS5. These data suggested that response to the inhibitory effect of nitric oxide radical may, at least in part, be regulated by NS5. The effect of these two different phenotypes of viruses on host cells was studied using cDNA array screening. The cDNA array analysis demonstrated that the nitric oxide-resistant group had a stronger influence on host cells since it induced changes in the expression of a greater number of genes than did the nitric oxide-susceptible group, 97 genes versus 71 genes, respectively. The NO-resistant virus also stimulated cytokines known to be virulent factors, such as IL 6, IL 8, RANTES, and the inflammatory factors. In conclusion, our data demonstrated that dengue viruses isolated from patients show genotypic and phenotypic differences which may correlate with virulence. © 2008 Elsevier Inc. All rights reserved.
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    Comparison of real-time SYBR green dengue assay with real-time taqman RT-PCR dengue assay and the conventional nested PCR for diagnosis of primary and secondary dengue infection
    (2011-10-01) Damodar Paudel; Richard Jarman; Kriengsak Limkittikul; Chonticha Klungthong; Supat Chamnanchanunt; Ananda Nisalak; Robert Gibbons; Watcharee Chokejindachai; Nepal Orthopaedic Hospital; Mahidol University; Armed Forces Research Institute of Medical Sciences, Thailand
    Background: Dengue fever and dengue hemorrhagic fever are caused by dengue virus. Dengue infection remains a burning problem of many countries. To diagnose acute dengue in the early phase we improve the low cost, rapid SYBR green real time assay and compared the sensitivity and specificity with real time Taqman ® assay and conventional nested PCR assay. Aims: To develop low cost, rapid and reliable real time SYBR green diagnostic dengue assay and compare with Taqman real-time assay and conventional nested PCR (modified Lanciotti). Materials and Methods: Eight cultured virus strains were diluted in tenth dilution down to undetectable level by the PCR to optimize the primer, temperature (annealing, and extension and to detect the limit of detection of the assay. Hundred and ninety three ELISA and PCR proved dengue clinical samples were tested with real time SYBR ® Green assay, real time Taqman ® assay to compare the sensitivity and specificity. Results: Sensitivity and specificity of real time SYBR® green dengue assay (84% and 66%, respectively) was almost comparable to those (81% and 74%) of Taqman real time PCR dengue assay. Real time SYBR ® green RT-PCR was equally sensitive in primary and secondary infection while real time Taqman was less sensitive in the secondary infection. Sensitivity of real time Taqman on DENV3 (87%) was equal to SYBR green real time PCR dengue assay. Conclusion: We developed low cost rapid diagnostic SYBR green dengue assay. Further study is needed to make duplex primer assay for the serotyping of dengue virus.
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    Construction of specific DNA probe for the detection of Salmonella in food
    (1997-03-01) Apiradee Pilantanapak; Panida Jayanetra; Watanalai Panbangred; Chonticha Klungthong; Aroon Bangtrakulnonth; Mahidol University; Thailand Ministry of Public Health; Burapha University
    The Salmonella specific DNA fragment from genomic DNA of S. typhimurium ATCC 23566 was cloned in E. coli and successfully used as a digoxigenin labeled probe for detecting the presence of Salmonella serotypes in both artificially contaminated food and natural contaminated food samples.
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    Detection of Escherichia coli in water and food by using DNA probe and polymerase chain reaction
    (Mahidol University. Mahidol University Library and Knowledge Center, 2024) Chonticha Klungthong; Watanalai Panbangred; Panida Jayanetra; Chuenchit Boonchird
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    Epidemiological and Molecular Characterization of Dengue Virus Circulating in Bhutan, 2013-2014
    (2015-08-21) Sangay Zangmo; Chonticha Klungthong; Piyawan Chinnawirotpisan; Srisurang Tantimavanich; Nathamon Kosoltanapiwat; Butsaya Thaisomboonsuk; Kelzang Phuntsho; Sonam Wangchuk; In-Kyu Yoon; Stefan Fernandez; Mahidol University. Faculty of Medical Technology. Department of Clinical Microbiology and Applied Technology
    Dengue is one of the most significant public health problems in tropical and subtropical countries, and is increasingly being detected in traditionally non-endemic areas. In Bhutan, dengue virus (DENV) has only recently been detected and limited information is available. In this study, we analyzed the epidemiological and molecular characteristics of DENV in two southern districts in Bhutan from 2013–2014. During this period, 379 patients were clinically diagnosed with suspected dengue, of whom 119 (31.4%) were positive for DENV infection by NS1 ELISA and/or nested RT-PCR. DENV serotypes 1, 2 and 3 were detected with DENV-1 being predominant. Phylogenetic analysis of DENV-1 using envelope gene demonstrated genotype V, closely related to strains from northern India.
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    Epidemiological and molecular characterization of dengue virus circulating in Bhutan, 2013-2014
    (2015-01-01) Sangay Zangmo; Chonticha Klungthong; Piyawan Chinnawirotpisan; Srisurang Tantimavanich; Nathamon Kosoltanapiwat; Butsaya Thaisomboonsuk; Kelzang Phuntsho; Sonam Wangchuk; In Kyu Yoon; Stefan Fernandez; Mahidol University; Ministry of Health; Armed Forces Research Institute of Medical Sciences, Thailand
    © 2015 Zangmo et al. Dengue is one of the most significant public health problems in tropical and subtropical countries, and is increasingly being detected in traditionally non-endemic areas. In Bhutan, dengue virus (DENV) has only recently been detected and limited information is available. In this study, we analyzed the epidemiological and molecular characteristics of DENV in two southern districts in Bhutan from 2013–2014. During this period, 379 patients were clinically diagnosed with suspected dengue, of whom 119 (31.4%) were positive for DENV infection by NS1 ELISA and/or nested RT-PCR. DENV serotypes 1, 2 and 3 were detected with DENV-1 being predominant. Phylogenetic analysis of DENV-1 using envelope gene demonstrated genotype V, closely related to strains from northern India.
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    Genomic surveillance of SARS-CoV-2 in Thailand reveals mixed imported populations, a local lineage expansion and a virus with truncated ORF7a
    (2021-01-15) Khajohn Joonlasak; Elizabeth M. Batty; Theerarat Kochakarn; Bhakbhoom Panthan; Krittikorn Kümpornsin; Poramate Jiaranai; Arporn Wangwiwatsin; Angkana Huang; Namfon Kotanan; Peera Jaru-Ampornpan; Wudtichai Manasatienkij; Treewat Watthanachockchai; Kingkan Rakmanee; Anthony R. Jones; Stefan Fernandez; Insee Sensorn; Somnuek Sungkanuparph; Ekawat Pasomsub; Chonticha Klungthong; Thanat Chookajorn; Wasun Chantratita; Khon Kaen University; Armed Forces Research Institute of Medical Sciences, Thailand; Faculty of Medicine Ramathibodi Hospital, Mahidol University; Mahidol University; Thailand National Center for Genetic Engineering and Biotechnology; Nuffield Department of Medicine; Wellcome Sanger Institute; COVID-19 Network Investigations Alliance
    Coronavirus Disease 2019 (COVID-19) is a global public health threat. Genomic surveillance of SARS-CoV-2 was implemented in March of 2020 at a major diagnostic hub in Bangkok, Thailand. Several virus lineages supposedly originated in many countries were found, and a Thai-specific lineage, designated A/Thai-1, has expanded to be predominant in Thailand. A virus sample in the SARS-CoV-2 A/Thai-1 lineage contains a frame-shift deletion at ORF7a, encoding a putative host antagonizing factor of the virus.
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    High prevalence of hepatitis G viremia among kidney transplant patients in Thailand
    (1997-10-01) Boonyos Raengsakulrach; Leena Ong-aj-yooth; Thanarak Thaiprasert; Sanga Nilwarangkur; Sompong Ong-aj-yooth; Sumitda Narupiti; Vipa Thirawuth; Chonticha Klungthong; Rapin Snitbhan; David W. Vaughn; Armed Forces Research Institute of Medical Sciences, Thailand; Mahidol University
    Patients receiving kidney transplants (KT) are at high risk for blood borne viral infections. To determine the prevalence of a recently discovered hepatitis G virus (HGV) in this patient group, reverse transcription-polymerase chain reaction (RT-PCR) employing primers derived from the NS5 region of the viral genome was utilized. HGV RNA was detected in 40 of 94 KT patients (43%), as compared to 3 of 69 healthy subjects (4.3%). Cocirculation of HGV and hepatitis C virus (HCV) RNA was detected in 12 patients (13%). Comparison of patients with and without HGV revealed that the former had received hemodialysis before transplantation for a significantly longer duration than the latter (28 vs. 17 months, respectively; P < 0.05). The amount of blood transfused and mean levels of liver enzymes, including alkaline phosphatase, alanine transaminase, and aspartate transaminase, were the same in both groups. Sequence analysis of 275-base pair DNA clones obtained from 2 patients revealed approximately 92% sequence homology to the published HGV and GB virus C sequences. These results suggested that HGV infection among Thai KT patients was high and the role of HGV in causing liver disease remains to be determined.
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    The molecular epidemiology of dengue virus serotype 4 in Bangkok, Thailand
    (2004-11-10) Chonticha Klungthong; Chunlin Zhang; Mammen P. Mammen; Sukathida Ubol; Edward C. Holmes; Armed Forces Research Institute of Medical Sciences, Thailand; Mahidol University; University of Oxford
    Dengue represents a major public health problem in Thailand, with all four viral serotypes co-circulating. Dengue virus serotype 4 (DENV-4) is the least frequently sampled serotype, although one that is often associated with hemorrhagic fever during secondary infection. To determine the evolutionary forces shaping the genetic diversity of DENV-4, and particularly whether its changing prevalence could be attributed to instances of adaptive evolution in the viral genome, we undertook a large-scale molecular epidemiological analysis of DENV-4 in Bangkok, Thailand, using both E gene and complete coding region sequences. This analysis revealed extensive genetic diversity within a single locality at a single time, including the discovery of a new and divergent genotype of DENV-4, as well as a pattern of continual lineage turnover. We also recorded the highest average rate of evolutionary change for this serotype, at 1.072 × 10-3nucleotide substitutions per site, per year. However, despite this abundant genetic variation, there was no evidence for adaptive evolution in any gene, codon, or lineage of DENV-4, with the highest rate of nonsynonymous substitution observed in NS2A. Consequently, the rapid turnover of DENV-4 lineages through time is most likely the consequence of a high rate of deleterious mutation in the viral genome coupled to seasonal fluctuations in the size of the vector population. © 2004 Elsevier Inc. All rights reserved.
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    Molecular evolution and epidemiology of dengue virus type 4 in Thailand
    (Mahidol University. Mahidol University Library and Knowledge Center, 2023) Chonticha Klungthong; Sukathida Ubol