Parasites, Hosts and Diseases

"Penchom Janwan"

Brief Communications

Evaluation of IgG4 Subclass Antibody Detection by Peptide-Based ELISA for the Diagnosis of Human Paragonimiasis Heterotrema: Pewpan M. Intapan, Oranuch Sanpool, Penchom Janwan, Porntip Laummaunwai, Nimit Morakote, Yoon Kong, Wanchai Maleewong; Korean J Parasitol 2013;51(6):763-766.
Published online December 31, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.6.763

Abstract
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A synthetic peptide was prepared based on the antigenic region of Paragonimus westermani pre-procathepsin L, and its applicability for immunodiagnosis for human paragonimiasis (due to Paragonimus heterotremus) was tested using an ELISA to detect IgG4 antibodies in the sera of patients. Sera from other helminthiases, tuberculosis, and healthy volunteers were used as the references. This peptide-based assay system gave sensitivity, specificity, accuracy, and positive and negative predictive values of 100%, 94.6%, 96.2%, 100%, and 88.9%, respectively. Cross reactivity was frequently seen against the sera of fascioliasis (75%) and hookworm infections (50%). Since differential diagnosis between paragonimiasis and fascioliasis can be easily done by clinical presentation and fascioliasis serology, this cross reaction is not a serious problem. Sera from patients with other parasitoses (0-25%) rarely responded to this synthetic antigen. This synthetic peptide antigen seems to be useful for development of a standardized diagnostic system for paragonimiasis.

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An immunochromatographic test using whole blood for rapid diagnosis of human paragonimiasis and its diagnostic usefulness
Patcharaporn Boonroumkaew, Lakkhana Sadaow, Penchom Janwan, Rutchanee Rodpai, Oranuch Sanpool, Tongjit Thanchomnang, Hiroshi Yamasaki, Pewpan M. Intapan, Wanchai Maleewong
Food and Waterborne Parasitology.2024; 37: e00246. CrossRef
Development of point-of-care testing tool using immunochromatography for rapid diagnosis of human paragonimiasis
Lakkhana Sadaow, Oranuch Sanpool, Hiroshi Yamasaki, Wanchai Maleewong, Pewpan M. Intapan
Acta Tropica.2020; 203: 105325. CrossRef
Paragonimus and paragonimiasis in Asia: An update
Ayako Yoshida, Pham Ngoc Doanh, Haruhiko Maruyama
Acta Tropica.2019; 199: 105074. CrossRef
A Case of Pulmonary Paragonimiasis with Chronic Abdominal Pain and Erythematous Rash in a 6-year-old Girl
Ju Young Kim, Min Kyu Park, Yong Ju Lee, Sun Huh, Ky Young Cho
Pediatric Infection and Vaccine.2018; 25(1): 54. CrossRef
A tool for mass-screening of paragonimiasis: an enzyme-linked immunosorbent assay with urine samples
Xu Guang Qiu, Fukumi Nakamura-Uchiyama, Yukifumi Nawa, Makoto Itoh
Tropical Medicine and Health.2016;[Epub] CrossRef
Expression characteristics and specific antibody reactivity of diverse cathepsin F members of Paragonimus westermani
Chun-Seob Ahn, Byoung-Kuk Na, Dong-ll Chung, Jeong-Geun Kim, Jin-Taek Kim, Yoon Kong
Parasitology International.2015; 64(1): 37. CrossRef
North American paragonimiasis: epidemiology and diagnostic strategies
Peter U Fischer, Gary J Weil
Expert Review of Anti-infective Therapy.2015; 13(6): 779. CrossRef

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Detection of Gnathostoma spinigerum Antibodies in Sera of Non-Traumatic Subarachnoid Hemorrhage Patients in Thailand: Amnat Kitkhuandee, Waranon Munkong, Kittisak Sawanyawisuth, Penchom Janwan, Wanchai Maleewong, Pewpan M. Intapan; Korean J Parasitol 2013;51(6):755-757.
Published online December 31, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.6.755

Abstract
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Gnathostoma spinigerum can cause subarachnoid hemorrhage (SAH). The detection of specific antibodies in serum against G. spinigerum antigen is helpful for diagnosis of neurognathostomiasis. There is limited data on the frequency of G. spinigerum infection in non-traumatic SAH. A series of patients diagnosed as non-traumatic SAH at the Srinagarind Hospital, Khon Kaen University, Thailand between January 2011 and January 2013 were studied. CT or MR imaging of the brain was used for diagnosis of SAH. Patients were categorized as aneurysmal subarachnoid hemorrhage (A-SAH) or non-aneurysmal subarachnoid hemorrhage (NA-SAH) according to the results of cerebral angiograms. The presence of specific antibodies in serum against 21- or 24-kDa G. spinigerum antigen was determined using the immunoblot technique. The detection rate of antibodies was compared between the 2 groups. Of the 118 non-traumatic SAH patients for whom cerebral angiogram and immunoblot data were available, 80 (67.8%) patients had A-SAH, whereas 38 (32.2%) had NA-SAH. Overall, 23.7% were positive for specific antibodies against 21- and/or 24-kDa G. spinigerum antigen. No significant differences were found in the positive rate of specific antibodies against G. spinigerum in both groups (P-value=0.350).

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A Recombinant Matrix Metalloproteinase Protein from Gnathostoma spinigerum for Serodiagnosis of Neurognathostomiasis: Penchom Janwan, Pewpan M. Intapan, Hiroshi Yamasaki, Porntip Laummaunwai, Kittisak Sawanyawisuth, Chaisiri Wongkham, Chatchai Tayapiwatana, Amnat Kitkhuandee, Viraphong Lulitanond, Yukifumi Nawa, Wanchai Maleewong; Korean J Parasitol 2013;51(6):751-754.
Published online December 31, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.6.751

Abstract
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Neurognathostomiasis is a severe form of human gnathostomiasis which can lead to disease and death. Diagnosis of neurognathostomiasis is made presumptively by using clinical manifestations. Immunoblotting, which recognizes antigenic components of molecular mass 21 kDa and 24 kDa in larval extracts of Gnathostoma spinigerum (Gs 21/24), has high sensitivity and specificity for diagnosis of neurognathostomiasis. However, only very small amounts of the Gs 21/24 antigens can be prepared from parasites harvested from natural or experimental animals. To overcome this problem, we recently produced a recombinant matrix metalloproteinase (rMMP) protein from G. spinigerum. In this study, we evaluated this rMMP alongside the Gs 21/24 antigens for serodiagnosis of human neurognathostomiasis. We studied sera from 40 patients from Srinagarind Hospital, Khon Kaen University, Thailand, with clinical criteria consistent with those of neurognathostomiasis, and sera from 30 healthy control adults from Thailand. All sera were tested for specific IgG antibodies against both G. spinigerum crude larval extract and rMMP protein using immunoblot analysis. The sensitivity and specificity for both antigenic preparations were all 100%. These results show that G. spinigerum rMMP protein can be used as an alternative diagnostic antigen, in place of larval extract, for serodiagnosis of neurognathostomiasis.

Citations

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Protein and antigen profiles of third-stage larvae of Gnathostoma spinigerum assessed with next-generation sequencing transcriptomic information
Kathyleen Nogrado, Tipparat Thiangtrongjit, Poom Adisakwattana, Paron Dekumyoy, Sant Muangnoicharoen, Charin Thawornkuno, Onrapak Reamtong
Scientific Reports.2022;[Epub] CrossRef
Development of Immunochromatographic Test Kit for Rapid Detection of Specific IgG4 Antibody in Whole-Blood Samples for Diagnosis of Human Gnathostomiasis
Penchom Janwan, Pewpan M. Intapan, Lakkhana Sadaow, Rutchanee Rodpai, Hiroshi Yamasaki, Patcharaporn Boonroumkaew, Oranuch Sanpool, Tongjit Thanchomnang, Phuangphaka Sadee, Wanchai Maleewong
Diagnostics.2021; 11(5): 862. CrossRef
Proteomics of Gnathostomiasis: A Way Forward for Diagnosis and Treatment Development
Tipparat Thiangtrongjit, Kathyleen Nogrado, Thawatchai Ketboonlue, Preeyarat Malaitong, Poom Adisakwattana, Onrapak Reamtong
Pathogens.2021; 10(9): 1080. CrossRef
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Sara Althubaiti, Şenay Kafkas, Marwa Abdelhakim, Robert Hoehndorf
Journal of Biomedical Semantics.2020;[Epub] CrossRef
Human gnathostomiasis: a neglected food-borne zoonosis
Guo-Hua Liu, Miao-Miao Sun, Hany M. Elsheikha, Yi-Tian Fu, Hiromu Sugiyama, Katsuhiko Ando, Woon-Mok Sohn, Xing-Quan Zhu, Chaoqun Yao
Parasites & Vectors.2020;[Epub] CrossRef
Surveillance and diagnosis of zoonotic foodborne parasites
Reza Zolfaghari Emameh, Sami Purmonen, Antti Sukura, Seppo Parkkila
Food Science & Nutrition.2018; 6(1): 3. CrossRef

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Molecular Detection of Ancylostoma duodenale, Ancylostoma ceylanicum, and Necator americanus in Humans in Northeastern and Southern Thailand: Issarapong Phosuk, Pewpan M. Intapan, Tongjit Thanchomnang, Oranuch Sanpool, Penchom Janwan, Porntip Laummaunwai, Witthaya Aamnart, Nimit Morakote, Wanchai Maleewong; Korean J Parasitol 2013;51(6):747-749.
Published online December 31, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.6.747

Abstract
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The 2 principal species of hookworms infecting humans are Necator americanus and Ancylostoma duodenale. Case studies on zoonotic hookworm infections with Ancylostoma ceylanicum and/or Ancylostoma caninum are known mainly from Asian countries. Of these 2 zoonotic species, only A. ceylanicum can develop to adulthood in humans. In the present study, we report a molecular-based survey of human hookworm infections present in southern and northeastern Thailand. Thirty larval hookworm samples were obtained from fecal agar plate cultures of 10 patients in northeastren Thailand and 20 in southern Thailand. Partial ITS1, 5.8S, and ITS2 regions of the ribosomal DNA genes were amplified using PCR. The amplicons were sequenced, aligned, and compared with other hookworm sequences in GenBank database. The results showed that, in Thailand, N. americanus is more prevalent than Ancylostoma spp. and is found in both study areas. Sporadic cases of A. ceylanicum and A. duodenale infection were seen in northeastern Thailand.

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Molecular identification and genetic diversity of zoonotic hookworm infections in domestic dogs from northeastern, Thailand
Rutchanee Rodpai, Oranuch Sanpool, Lakkhana Sadaow, Patcharaporn Boonroumkaew, Pewpan M. Intapan, Wanchai Maleewong, Manachai Yingklang, Penchom Janwan, Kotchaphon Vaisusuk, Wasupon Chatan, Supawadee Piratae, Tongjit Thanchomnang
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Wararat Sangwalee, Jun Norkaew, Sengchoy Inthachak, Penchom Janwan, Rutchanee Rodpai, Oranuch Sanpool, Lakkhana Sadaow, Patcharaporn Boonroumkaew, Pewpan M. Intapan, Wanchai Maleewong, Tongjit Thanchomnang, Raffi V. Aroian
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Original Articles

Molecular Differentiation of Opisthorchis viverrini and Clonorchis sinensis Eggs by Multiplex Real-Time PCR with High Resolution Melting Analysis: Worasak Kaewkong, Pewpan M. Intapan, Oranuch Sanpool, Penchom Janwan, Tongjit Thanchomnang, Porntip Laummaunwai, Viraphong Lulitanond, Pham Ngoc Doanh, Wanchai Maleewong; Korean J Parasitol 2013;51(6):689-694.
Published online December 31, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.6.689

Abstract
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Opisthorchis viverrini and Clonorchis sinensis are parasites known to be carcinogenic and causative agents of cholangiocarcinoma in Asia. The standard method for diagnosis for those parasite infections is stool examination to detect parasite eggs. However, the method has low sensitivity, and eggs of O. viverrini and C. sinensis are difficult to distinguish from each other and from those of some other trematodes. Here, we report a multiplex real-time PCR coupled with high resolution melting (HRM) analysis for the differentiation of O. viverrini and C. sinensis eggs in fecal samples. Using 2 pairs of species-specific primers, DNA sequences from a portion of the mitochondrial NADH dehydrogenase subunit 2 (nad 2) gene, were amplified to generate 209 and 165 bp products for O. viverrini and C. sinensis, respectively. The distinct characteristics of HRM patterns were analyzed, and the melting temperatures peaked at 82.4±0.09℃ and 85.9±0.08℃ for O. viverrini and C. sinensis, respectively. This technique was able to detect as few as 1 egg of O. viverrini and 2 eggs of C. sinensis in a 150 mg fecal sample, which is equivalent to 7 and 14 eggs per gram of feces, respectively. The method is species-specific, rapid, simple, and does not require fluorescent probes or post-PCR processing for discrimination of eggs of the 2 species. It offers a new tool for differentiation and detection of Asian liver fluke infections in stool specimens.

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Molecular Variation in the Paragonimus heterotremus Complex in Thailand and Myanmar: Oranuch Sanpool, Pewpan M. Intapan, Tongjit Thanchomnang, Penchom Janwan, Yukifumi Nawa, David Blair, Wanchai Maleewong; Korean J Parasitol 2013;51(6):677-681.
Published online December 31, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.6.677

Abstract
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Paragonimiasis is an important food-borne parasitic zoonosis caused by infection with lung flukes of the genus Paragonimus. Of the 7 members of the genus known in Thailand until recently, only P. heterotremus has been confirmed as causing human disease. An 8th species, P. pseudoheterotremus, has recently been proposed from Thailand, and has been found in humans. Molecular data place this species as a sister species to P. heterotremus, and it is likely that P. pseudoheterotremus is not specifically distinct from P. heterotremus. In this study, we collected metacercariae of both nominal species (identification based on metacercarial morphology) from freshwater crabs from Phetchabun Province in northern Thailand, Saraburi Province in central Thailand, and Surat Thani Province in southern Thailand. In addition, we purchased freshwater crabs imported from Myanmar at Myawaddy Province, western Thailand, close to the Myanmar-Thailand border. The DNAs extracted from excysted metacercariae were PCR-amplified and sequenced for ITS2 and cox1 genes. The ITS2 sequences were nearly identical among all samples (99-100%). Phylogenies inferred from all available partial cox1 sequences contained several clusters. Sequences from Indian P. heterotremus formed a sister group to sequences from P. pseudoheterotremus-type metacercariae. Sequences of P. heterotremus from Thailand, Vietnam, and China formed a separate distinct clade. One metacercaria from Phitsanulok Province was distinct from all others. There is clearly considerable genetic variation in the P. heterotremus complex in Thailand and the form referred to as P. pseudoheterotremus is widely distributed in Thailand and the Thai-Myanmar border region.

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Molecular Differentiation of Schistosoma japonicum and Schistosoma mekongi by Real-Time PCR with High Resolution Melting Analysis: Amornmas Kongklieng, Worasak Kaewkong, Pewpan M. Intapan, Oranuch Sanpool, Penchom Janwan, Tongjit Thanchomnang, Viraphong Lulitanond, Pusadee Sri-Aroon, Yanin Limpanont, Wanchai Maleewong; Korean J Parasitol 2013;51(6):651-656.
Published online December 31, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.6.651

Abstract
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Human schistosomiasis caused by Schistosoma japonicum and Schistosoma mekongi is a chronic and debilitating helminthic disease still prevalent in several countries of Asia. Due to morphological similarities of cercariae and eggs of these 2 species, microscopic differentiation is difficult. High resolution melting (HRM) real-time PCR is developed as an alternative tool for the detection and differentiation of these 2 species. A primer pair was designed for targeting the 18S ribosomal RNA gene to generate PCR products of 156 base pairs for both species. The melting points of S. japonicum and S. mekongi PCR products were 84.5±0.07℃ and 85.7±0.07℃, respectively. The method permits amplification from a single cercaria or an egg. The HRM real-time PCR is a rapid and simple tool for differentiation of S. japonicum and S. mekongi in the intermediate and final hosts.

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Rapid Detection and Identification of Wuchereria bancrofti, Brugia malayi, B. pahangi, and Dirofilaria immitis in Mosquito Vectors and Blood Samples by High Resolution Melting Real-Time P: Tongjit Thanchomnang, Pewpan M. Intapan, Chairat Tantrawatpan, Viraphong Lulitanond, Sudchit Chungpivat, Piyanan Taweethavonsawat, Worasak Kaewkong, Oranuch Sanpool, Penchom Janwan, Wej Choochote, Wanchai Maleewong; Korean J Parasitol 2013;51(6):645-650.
Published online December 31, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.6.645

Abstract
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A simple, rapid, and high-throughput method for detection and identification of Wuchereria bancrofti, Brugia malayi, Brugia pahangi, and Dirofilaria immitis in mosquito vectors and blood samples was developed using a real-time PCR combined with high-resolution melting (HRM) analysis. Amplicons of the 4 filarial species were generated from 5S rRNA and spliced leader sequences by the real-time PCR and their melting temperatures were determined by the HRM method. Melting of amplicons from W. bancrofti, B. malayi, D. immitis, and B. pahangi peaked at 81.5±0.2℃, 79.0±0.3℃, 76.8±0.1℃, and 79.9±0.1℃, respectively. This assay is relatively cheap since it does not require synthesis of hybridization probes. Its sensitivity and specificity were 100%. It is a rapid and technically simple approach, and an important tool for population surveys as well as molecular xenomonitoring of parasites in vectors.

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Modulation of Antibody Responses against Gnathostoma spinigerum in Mice Immunized with Crude Antigen Formulated in CpG Oligonucleotide and Montanide ISA720: Pewpan M. Intapan, Chakrit Hirunpetcharat, Churairat Kularbkaew, Wiboonchai Yutanawiboonchai, Penchom Janwan, Wanchai Maleewong; Korean J Parasitol 2013;51(6):637-644.
Published online December 31, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.6.637

Abstract
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This study aimed to investigate the antibody responses in mice immunized with Gnathostoma spinigerum crude antigen (GsAg) incorporated with the combined adjuvant, a synthetic oligonucleotide containing unmethylated CpG motif (CpG ODN 1826) and a stable water in oil emulsion (Montanide ISA720). Mice immunized with GsAg and combined adjuvant produced all antibody classes and subclasses to GsAg except IgA. IgG2a/2b/3 but not IgG1 subclasses were enhanced by immunization with CpG ODN 1826 when compared with the control groups immunized with non-CpG ODN and Montanide ISA or only with Montanide ISA, suggesting a biased induction of a Th1-type response by CpG ODN. After challenge infection with live G. spinigerum larvae, the levels of IgG2a/2b/3 antibody subclasses decreased immediately and continuously, while the IgG1 subclass remained at high levels. This also corresponded to a continuous decrease of the IgG2a/IgG1 ratio after infection. Only IgM and IgG1 antibodies, but not IgG2a/2b/3, were significantly produced in adjuvant control groups after infection. These findings suggest that G. spinigerum infection potently induces a Th2-type biased response.

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