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Mammomonogamus roundworm (Nematoda: Syngamidae) recovered from the duodenum of a Thai patient: a first and unusual case originating in Thailand

Eamsobhana P, Mongkolporn T, Punthuprapasa P, Yoolek A

Trans R Soc Trop Med Hyg, 2006 Apr;100(4):387-91.
PMID: 16257022

A pair of Mammomonogamus laryngeus roundworms in copula was recovered from the duodenum of a 72-year-old male Thai patient from Kanchanaburi Province. Eggs were also found in the stool of the patient. This is the first case of Mammomonogamus infection originating in Thailand, as the previous two reports from Thailand attributed the infection as originating in Malaysia. The occurrence of adult worms in the duodenum is unusual and differs from previous findings in the larynx, posterior pharynx, tracheal wall and bronchi. This adds a new dimension to the manifestation of the infection.
Fulltext Prognostic Factors in Recurrent Congenital Muscular Torticollis

Chotigavanichaya C, Phongprapapan P, Wongcharoenwatana J, Eamsobhana P, Ariyawatkul T, Kaewpornsawan K

Malays Orthop J, 2021 Mar;15(1):43-47.
PMID: 33880147 DOI: 10.5704/MOJ.2103.007

Introduction: Congenital muscular torticollis (CMT), primarily resulting from unilateral shortening and fibrosis of the sternocleidomastoid muscle. One of the common surgical complications is recurrent deformity. However, the associations between unipolar or bipolar release, age of the patient, and the recurrence of the disease are unclear. Therefore, the purpose of this study was to evaluate the factors associated with recurrence after surgery.
Materials and Methods: A retrospective review was performed in 47 patients who were diagnosed with CMT and had been treated surgically with unipolar or bipolar release between January 2007 and December 2015. Demographic data (sex, sides, surgical technique, age at time of surgery, period of follow-up, complications and recurrence) were recorded.
Results: Forty-seven patients with an average age of 8.7 years old at time of surgery. Twenty-six patients had right-sided muscular torticollis, while 21 had left-sided. The average follow-up time was 2 years (range, 2-4 years). The average age of unipolar release was 8.8 years old (range, 218 years old), while the average age of bipolar release was 8.7 years old (range, 2-13 years old). Recurrence occurred in 11 patients (9 in unipolar and 2 in bipolar release). Sex, side of deformity, type of surgery and age at time of surgery showed no statistically significant as a factor for recurrence rate, however recurrence of unipolar more than bipolar surgery was nearly two times revealing clinical significance.
Conclusions: Sex, side of deformity, type of surgery and age at time of surgery were not associated with the recurrence deformity.
Fulltext Evaluation of dot immunogold filtration assay (DIGFA) for rapid serodiagnosis of eosinophilic meningitis due to Angio-strongylus cantonensis (Nematoda: Metastrongyloidea)

Eamsobhana P, Prasartvit A, Gan XX, Yong HS

Trop Biomed, 2015 Mar;32(1):121-5.
PMID: 25801261

Angiostrongylus cantonensis is the most frequent cause of eosinophilic meningitis in humans in Thailand and worldwide. Because of difficulty of recovering the Angiostrongylus larvae from infected patients, detection of parasite-specific antibodies is used to support clinical diagnosis. This study tested serum samples from eosinophilic meningitis patients and individuals at risk of infection with A. cantonensis to evaluate a recently developed simple and rapid dot-immunogold filtration assay (DIGFA) for detection of specific antibodies against A. cantonensis. Purified 31-kDa glycoprotein of A. cantonensis and protein A colloidal gold conjugate were employed to detect the 31-kDa anti-A. cantonensis antibody in patients sera from the parasite endemic areas of northeast Thailand. The results were compared with those obtained by dot-blot enzyme-linked immunosorbent assay (ELISA) with 31-kDa A. cantonensis antigen. The overall positivity rate of DIGFA and dot-blot ELISA for A. cantonensis infection in 98 clinically diagnosed cases from three highly endemic districts in Khon Kaen province were 39.79% and 37.75%, respectively. Among 86 sera of subjects at risk of infection with A. cantonensis, 24.41% were positive by DIGFA and 23.25% by dot-blot ELISA. There were good correlation between the visual grading of DIGFA and dot-blot ELISA in both groups of defined sera. DIGFA is as sensitive and specific as dot-blot ELISA for confirming eosinophilic meningitis due to A. cantonensis infection, with advantages of simplicity, rapidity and without the use of specific and expensive equipment, and can be used in field settings.
Genetic variation of NADH dehydrogenase subunit 1 (nad1) mitochondrial gene sequence in adult Necator americanus hookworms recovered from a female patient in Thailand

Eamsobhana P, Yong HS, Roongruangchai K, Tungtrongchitr A, Wanachiwanawin D

Trop Biomed, 2020 Jun 01;37(2):536-541.
PMID: 33612820

Two female and one male adult hookworms were recovered from a female patient in Thailand. Based on gross and microscopic morphology, the three hookworms are members of Necator americanus. Phylogenetic reconstruction based on partial NADH dehydrogenase subunit 1 (nad1) mitochondrial gene sequences shows that these hookworms belong to the same genetic lineage as N. americanus adult worm from Zhejiang, China. The male and female hookworms were genetically distinct, belonging to two different nad1-haplotypes. This is the first report targeting the nad1 gene on the identification and genetic characterization of the human hookworms originated from infected patient. The nad1 gene marker is useful for species and higher taxa differentiation of hookworms.
Phylogenetics and systematics of Angiostrongylus lungworms and related taxa (Nematoda: Metastrongyloidea) inferred from the nuclear small subunit (SSU) ribosomal DNA sequences

Eamsobhana P, Lim PE, Yong HS

J Helminthol, 2015 May;89(3):317-25.
PMID: 24622302 DOI: 10.1017/S0022149X14000108

The Angiostrongylus lungworms are of public health and veterinary concern in many countries. At the family level, the Angiostrongylus lungworms have been included in the family Angiostrongylidae or the family Metastrongylidae. The present study was undertaken to determine the usefulness and suitability of the nuclear 18S (small subunit, SSU) rDNA sequences for differentiating various taxa of the genus Angiostrongylus, as well as to determine the systematics and phylogenetic relationship of Angiostrongylus species and other metastrongyloid taxa. This study revealed six 18S (SSU) haplotypes in A. cantonensis, indicating considerable genetic diversity. The uncorrected pairwise 'p' distances among A. cantonensis ranged from 0 to 0.86%. The 18S (SSU) rDNA sequences unequivocally distinguished the five Angiostrongylus species, confirmed the close relationship of A. cantonensis and A. malaysiensis and that of A. costaricensis and A. dujardini, and were consistent with the family status of Angiostrongylidae and Metastrongylidae. In all cases, the congeneric metastrongyloid species clustered together. There was no supporting evidence to include the genus Skrjabingylus as a member of Metastrongylidae. The genera Aelurostrongylus and Didelphostrongylus were not recovered with Angiostrongylus, indicating polyphyly of the Angiostrongylidae. Of the currently recognized families of Metastrongyloidea, only Crenosomatidae appeared to be monophyletic. In view of the unsettled questions regarding the phylogenetic relationships of various taxa of the metastrongyloid worms, further analyses using more markers and more taxa are warranted.
Angiostrongylus mackerrasae and A. cantonensis (Nematoda: Metastrongyloidea) belong to same genetic lineage: evidence from mitochondrial protein-coding genes

Song SL, Yong HS, Eamsobhana P

J Helminthol, 2018 Jul;92(4):524-529.
PMID: 28693647 DOI: 10.1017/S0022149X1700061X

Angiostrongylus mackerrasae is a parasitic nematode of rats found in Australia. When first reported, it was referred to as A. cantonensis. Recent molecular studies, including the mitochondrial genome, indicate that it is highly similar to A. cantonensis. These studies did not include A. malaysiensis, another member of the A. cantonensis species complex, for comparison. The present study examined the genetic distance and phylogenetic relationship between the component taxa (A. cantonensis, A. mackerrasae and A. malaysiensis) of the A. cantonensis species complex, based on the 12 protein-coding genes (PCGs) of their mitochondrial genome. Both the nucleotide and amino acid sequences were analysed. Angiostrongylus mackerrasae and A. cantonensis are members of the same genetic lineage and both are genetically distinct from A. malaysiensis. The genetic distance based on concatenated nucleotide sequences of 12 mt-PCGs between A. mackerrasae and A. cantonensis from Thailand is p = 1.73%, while that between the Thai and Chinese taxa of A. cantonensis is p = 3.52%; the genetic distance between A. mackerrasae and A. cantonensis from China is p = 3.70%. The results indicate that A. mackerrasae and A. cantonensis belong to the same genetic lineage, and that A. mackerrasae may be conspecific with A. cantonensis. It remains to be resolved whether A. mackerrasae is conspecific with A. cantonensis or undergoing incipient speciation.
Detection of circulating antigens of Parastrongylus cantonensis in human sera by sandwich ELISA with specific monoclonal antibody

Eamsobhana P, Mak JW, Yong HS

Southeast Asian J Trop Med Public Health, 1995 Dec;26(4):712-5.
PMID: 9139382

A specific monoclonal antibody (AW-3C2) as revealed by ELISA was produced against the adult worm antigens of Parastrongylus cantonensis and used in a sandwich ELISA for the detection of circulating antigens in the sera of parastrongyliasis patients and those with other parasitic diseases. A total of 60 sera was used in this study. Of these, 10 each were from patients with parastrongyliasis, cysticercosis, filariasis, gnathostomiasis, malaria and toxocariasis. The control group consisted of 53 serum samples from normal healthy Thais and Malaysians. The mean +/- optical density (OD) values for the normal Thai and Malaysian groups were 0.126 +/- 0.028 and 0.124 +/- 0.029, respectively. The mean OD values of the parastrongyliasis patient group differed significantly from that of the normal groups as well as those of other parasitic infections. Using a cut-off point of OD +/- 3SD of the control groups as indicating a positive reading, the specificity of the assay with this monoclonal antibody was 100% while the sensitivity was 50%.
Fulltext Detection of circulating antigens of Parastrongylus cantonensis in human sera by dot-blot ELISA and sandwich ELISA using monoclonal antibody

Eamsobhana P, Yong HS, Mak JW, Wattanakulpanich D

Southeast Asian J Trop Med Public Health, 1997 Sep;28(3):624-8.
PMID: 9561620

A dot-blot ELISA was compared with a previously performed sandwich ELISA for the detection of Parastrongylus cantonensis antigens in sera from patients. Using the same monoclonal antibody and the same sera, 6 of 10 sera (60%) from parastronglyiasis patients were positive in dot-blot ELISA, whereas with sandwich ELISA, 5 of the same patient sera (50%) were positive. The specificity in both assays was 100% using 50 sera from patients with other parasitic diseases; of these, 10 each were from patients with cysticercosis, filariasis, gnathostomiasis, malaria and toxocariasis. The control group consisted of 53 sera from normal health Thais and Malaysians. The sensitivity of the assays was, however, slightly better with dot-blot ELISA and because it is simple, quick and cost-effective, it may be a test of choice for specific diagnosis of human parastrongyliasis.
Complete mitochondrial genome of Angiostrongylus malaysiensis lungworm and molecular phylogeny of Metastrongyloid nematodes

Yong HS, Song SL, Eamsobhana P, Lim PE

Acta Trop, 2016 May 17;161:33-40.
PMID: 27207134 DOI: 10.1016/j.actatropica.2016.05.002

Angiostrongylus malaysiensis is a nematode parasite of various rat species. When first documented in Malaysia, it was referred to as A. cantonensis. Unlike A. cantonensis, the complete mitochondrial genome of A. malaysiensis has not been documented. We report here its complete mitogenome, its differentiation from A. cantonensis, and the phylogenetic relationships with its congeners and other Metastrongyloid taxa. The whole mitogenome of A. malaysiensis had a total length of 13,516bp, comprising 36 genes (12 PCGs, 2 rRNA and 22 tRNA genes) and a control region. It is longer than that of A. cantonensis (13,509bp). Its control region had a long poly T-stretch of 12bp which was not present in A. cantonensis. A. malaysiensis and A. cantonensis had identical start codon for the 12 PCGs, but four PCGs (atp6, cob, nad2, nad6) had different stop codon. The cloverleaf structure for the 22 tRNAs was similar in A. malaysiensis and A. cantonensis except the TΨC-arm was absent in trnV for A. malaysiensis but present in A. cantonensis. The Angiostrongylus genus was monophyletic, with A. malaysiensis and A. cantonensis forming a distinct lineage from that of A. costaricensis and A. vasorum. The genetic distance between A. malaysiensis and A. cantonensis was p=11.9% based on 12 PCGs, p=9.5% based on 2 rRNA genes, and p=11.6% based on 14 mt-genes. The mitogenome will prove useful for studies on phylogenetics and systematics of Angiostrongylus lungworms and other Metastrongyloid nematodes.
Fulltext Immunochromatographic test for rapid serological diagnosis of human angiostrongyliasis

Eamsobhana P, Tungtrongchitr A, Wanachiwanawin D, Yong HS

Int J Infect Dis, 2018 Aug;73:69-71.
PMID: 29908250 DOI: 10.1016/j.ijid.2018.06.005

OBJECTIVES: The serological diagnosis of human infection with Angiostrongylus cantonensis remains problematic because there are no commercially available validated tests. Most laboratories use domestically prepared tests such as the enzyme-linked immunosorbent assay (ELISA) or immunoblotting. Since laboratory facilities are not always available in endemic areas, we developed and assessed a rapid lateral flow immunochromatographic assay (AcQuickDx Test) to detect anti-A. cantonensis antibodies in human serum.
METHODS: The test device was assembled with purified 31-kDa glycoprotein as diagnostic antigen and with gold-labelled anti-human immunoglublin-G as the detector reagent. A total of 97 serum samples were tested - 19 samples from clinically diagnosed patients with detectable A. cantonensis-specific antibody in immunoblotting; 43 samples from patients with other parasitic diseases, i.e. gnathostomiasis (n=13), toxocariasis (n=2), trichinellosis (n=2), hookworm infection (n=4), filariasis (n=5), cysticercosis (n=9), paragonimiasis (n=2), opisthorchiasis (n=3), and malaria (n=3); and 35 samples from normal healthy subjects.
RESULTS: The sensitivity, specificity, positive predictive value and negative predictive value of AcQuickDx Test to detect anti-A. cantonensis specific antibodies in serologically confirmed angiostrongyliasis cases, were 100%, 98.72%, 95% and 100%, respectively. Positive AcQuickDx was observed in 1 of 4 cases with hookworm infections. No positive AcQuickDx was observed in cases with other parasitic diseases, and the individual healthy subjects.
CONCLUSIONS: AcQuickDx Test is rapid, highly sensitive and specific, and easy to perform without additional equipment or ancillary supplies. It yields results that are interpreted visually, and possesses a long shelf-life at room temperature. Thus, it can be applied as an additional test for clinical diagnostic support of angiostrongyliasis either in conventional laboratories or for remote areas where laboratory infrastructure is not available.
Dot immunogold filtration assay (DIGFA) for the rapid detection of specific antibodies against the rat lungworm Angiostrongylus cantonensis (Nematoda: Metastrongyloidea) using purified 31-kDa antigen

Eamsobhana P, Gan XX, Ma A, Wang Y, Wanachiwanawin D, Yong HS

J Helminthol, 2014 Dec;88(4):396-401.
PMID: 23710755 DOI: 10.1017/S0022149X13000321

A rapid dot immunogold filtration assay (DIGFA) was adopted for specific immunodiagnosis of human cerebral angiostrongyliasis, using purified 31-kDa glycoprotein specific to Angiostrongylus cantonensis as diagnostic antigen and protein A colloidal gold conjugate as antigen-antibody detector. A total of 59 serum samples were assayed - 11 samples from clinically diagnosed patients with detectable A. cantonensis-specific antibody in immunoblotting; 23 samples from patients with other related parasitic diseases, i.e. gnathostomiasis (n= 8), cysticercosis (n= 5), toxocariasis (n= 2), filariasis (n= 4), paragonimiasis (n= 2) and malaria (n= 2); and 25 samples from normal healthy subjects. The sensitivity and specificity of DIGFA to detect anti-A. cantonensis specific antibodies in serologically confirmed angiostrongyliasis cases, were both 100%. No positive DIGFA was observed in cases with other parasitic diseases, and the healthy control subjects. The 3-min DIGFA is as sensitive and specific as the 3-h immunoblot test in angiostrongyliasis confirmed cases that revealed a 31-kDa reactive band. The gold-based DIGFA is more rapid and easier to perform than the traditional enzyme-linked immunosorbent assay (ELISA). The test utilizing purified A. cantonensis antigen is reliable and reproducible for specific immunodiagnosis of human infection with A. cantonensis - thus can be applied as an additional routine test for clinical diagnostic support. Large-scale sero-epidemiological studies in endemic communities in north-east Thailand are under way to evaluate its usefulness under field conditions.
Cytochrome c oxidase subunit I haplotype reveals high genetic diversity of Angiostrongylus malaysiensis (Nematoda: Angiostrongylidae)

Eamsobhana P, Yong HS, Song SL, Prasartvit A, Boonyong S, Tungtrongchitr A

J Helminthol, 2018 Mar;92(2):254-259.
PMID: 28330511 DOI: 10.1017/S0022149X17000244

The rat lungworm Angiostrongylus malaysiensis is a metastrongyloid nematode parasite. It has been reported in Malaysia, Thailand, Laos, Myanmar, Indonesia and Japan. In this study, A. malaysiensis adult worms recovered from the lungs of wild rats in different geographical regions/provinces in Thailand were used to determine their haplotype by means of the mitochondrial partial cytochrome c oxidase subunit I (COI) gene sequence. The results revealed high COI haplotype diversity of A. malaysiensis from Thailand. The geographical isolates of A. malaysiensis from Thailand and other countries formed a monophyletic clade distinct from the closely related A. cantonensis. In the present study, five new haplotypes were identified in addition to the four haplotypes reported in the literature. Phylogenetic analysis revealed that four of these five new haplotypes - one from Mae Hong Song (northern region), two from Tak (western region) and one from Phang Nga (southern region) - formed a distinct clade with those from Phatthalung (southern region) and Malaysia. The haplotype from Malaysia was identical to that of Phatthalung (haplotype AM1). In general, the COI sequences did not differentiate unambiguously the various geographical isolates of A. malaysiensis. This study has confirmed the presence of high COI genetic diversity in various geographical isolates of A. malaysiensis. The COI gene sequence will be suitable for studying genetic diversity, population structure and phylogeography.
Genetic diversity of infective larvae of Gnathostoma spinigerum (Nematoda: Gnathostomatidae) in freshwater swamp eels from Thailand

Eamsobhana P, Wanachiwanawin D, Roongruangchai K, Song SL, Yong HS

J Helminthol, 2017 Nov;91(6):767-771.
PMID: 27890039 DOI: 10.1017/S0022149X16000857

Human gnathostomiasis is a food-borne zoonosis caused by a tissue nematode of the genus Gnathostoma. The disease is highly endemic in Asia, including Thailand. The freshwater swamp eel (Monopterus albus), the second intermediate host of the gnathostome nematode, has an important role in transmitting the infection in Thailand. Surveys on the infective larvae of Gnathostoma spinigerum based on morphological features in freshwater swamp eels have been performed continuously and reported in Thailand. However, there is still limited molecular data on intra-species variations of the parasite. In this study, a total of 19 third-stage larvae of morphologically identified G. spinigerum were collected from 437 liver samples of freshwater swamp eels purchased from a large wholesale market in Bangkok, Thailand. Molecular characterization based on mitochondrial cytochrome c oxidase subunit I (COI) sequences was performed to elucidate their genetic variations and phylogenetic relationship. Among the 19 infective larvae recovered from these eels, 16 were sequenced successfully. Phylogenetic analyses inferred from the partial COI gene showed the presence of three distinct COI haplotypes. Our findings confirm the presence of G. spinigerum as the main species in Thailand.
Fulltext Distinct genetic clades of Malaysian Copera damselflies and the phylogeny of platycnemine subfamilies

Lim PE, Tan J, Eamsobhana P, Yong HS

Sci Rep, 2013 Oct 17;3:2977.
PMID: 24131999 DOI: 10.1038/srep02977

The phylogenetic relationships of some taxa in the Platycnemidinae at the species and generic levels have been investigated. Phylogenetic trees were generated from both individual mitochondrial encoded COI, COII, 16S rDNA and nuclear encoded 28S rDNA and also combined sequences; these data indicate that the component taxa of the genus Copera belong to two distinct genetic clades - the marginipes group and the annulata group. There was no distinct genetic difference between the red-legged and yellow-legged morphs of C. vittata. Molecular data showed that the annulata group is considered a member of the genus Platycnemis, as originally proposed. The genus Coeliccia, a member of the subfamily Calicnemiinae (Platycnemididae), is not grouped with the Platycnemidinae. The Disparoneurinae of the 'Protoneuridae' showed a closer relationship to the Platycnemidinae than the Calicnemiinae. The dataset supports the placement of the Disparoneurinae as a subfamily of the Platycnemididae. This resolves the monophyly of Platycnemididae.
Fulltext Complete mitochondrial genome of Zeugodacus tau (Insecta: Tephritidae) and differentiation of Z. tau species complex by mitochondrial cytochrome c oxidase subunit I gene

Yong HS, Song SL, Lim PE, Eamsobhana P

PLoS One, 2017;12(12):e0189325.
PMID: 29216281 DOI: 10.1371/journal.pone.0189325

The tephritid fruit fly Zeugodacus tau (Walker) is a polyphagous fruit pest of economic importance in Asia. Studies based on genetic markers indicate that it forms a species complex. We report here (1) the complete mitogenome of Z. tau from Malaysia and comparison with that of China as well as the mitogenome of other congeners, and (2) the relationship of Z. tau taxa from different geographical regions based on sequences of cytochrome c oxidase subunit I gene. The complete mitogenome of Z. tau had a total length of 15631 bp for the Malaysian specimen (ZT3) and 15835 bp for the China specimen (ZT1), with similar gene order comprising 37 genes (13 protein-coding genes-PCGs, 2 rRNA genes, and 22 tRNA genes) and a non-coding A + T-rich control region (D-loop). Based on 13 PCGs and 15 mt-genes, Z. tau NC_027290 (China) and Z. tau ZT1 (China) formed a sister group in the lineage containing also Z. tau ZT3 (Malaysia). Phylogenetic analysis based on partial sequences of cox1 gene indicates that the taxa from China, Japan, Laos, Malaysia, Bangladesh, India, Sri Lanka, and Z. tau sp. A from Thailand belong to Z. tau sensu stricto. A complete cox1 gene (or 13 PCGs or 15 mt-genes) instead of partial sequence is more appropriate for determining phylogenetic relationship.
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.
Fulltext Distinct genetic lineages of Bactrocera caudata (Insecta: Tephritidae) revealed by COI and 16S DNA sequences

Lim PE, Tan J, Suana IW, Eamsobhana P, Yong HS

PLoS One, 2012;7(5):e37276.
PMID: 22615962 DOI: 10.1371/journal.pone.0037276

The fruit fly Bactrocera caudata is a pest species of economic importance in Asia. Its larvae feed on the flowers of Cucurbitaceae such as Cucurbita moschata. To-date it is distinguished from related species based on morphological characters. Specimens of B. caudata from Peninsular Malaysia and Indonesia (Bali and Lombok) were analysed using the partial DNA sequences of cytochrome c oxidase subunit I (COI) and 16S rRNA genes. Both gene sequences revealed that B. caudata from Peninsular Malaysia was distinctly different from B. caudata of Bali and Lombok, without common haplotype between them. Phylogenetic analysis revealed two distinct clades, indicating distinct genetic lineage. The uncorrected 'p' distance for COI sequences between B. caudata of Malaysia-Thailand-China and B. caudata of Bali-Lombok was 5.65%, for 16S sequences from 2.76 to 2.99%, and for combined COI and 16S sequences 4.45 to 4.46%. The 'p' values are distinctly different from intraspecific 'p' distance (0-0.23%). Both the B. caudata lineages are distinctly separated from related species in the subgenus Zeugodacus - B. ascita, B. scutellata, B. ishigakiensis, B. diaphora, B. tau, B. cucurbitae, and B. depressa. Molecular phylogenetic analysis indicates that the B. caudata lineages are closely related to B. ascita sp. B, and form a clade with B. scutellata, B. ishigakiensis, B. diaphora and B. ascita sp. A. This study provides additional baseline for the phylogenetic relationships of Bactrocera fruit flies of the subgenus Zeugodacus. Both the COI and 16S genes could be useful markers for the molecular differentiation and phylogenetic analysis of tephritid fruit flies.
Complete mitochondrial genome reveals genetic diversity of Angiostrongylus cantonensis (Nematoda: Angiostrongylidae)

Yong HS, Song SL, Eamsobhana P, Goh SY, Lim PE

Acta Trop, 2015 Dec;152:157-164.
PMID: 26348256 DOI: 10.1016/j.actatropica.2015.09.001

Angiostrongylus cantonensis is a zoonotic parasite that causes eosinophilic meningitis in humans. Earlier work on its mitochondrial genome was based on long polymerase chain reaction method. To date, only the mitogenome of the isolates from China has been studied. We report here the complete mitogenome of the Thailand isolate based on next generation sequencing and compare the genetic diversity with other isolates. The mitogenome of the Thailand isolate (13,519bp) is longer than those of the China isolates (13,497-13,502bp). Five protein-coding genes (atp6, cox1, cox2, cob, nad2) show variations in length among the isolates. The stop codon of the Thailand isolate differs from the China and Taiwan isolates in 4 genes (atp6, cob, nad2, nad6). Additionally, the Thailand isolate has 4 incomplete T stop codon compared to 3 in the China and Taiwan isolates. The control region is longer in the Thailand isolate (258bp) than the China (230-236bp) and Taiwan (237bp) isolates. The intergenic sequence between nad4 and cox1 genes in the Thailand isolate lacks 2bp (indels) at the 5'-end of the sequence as well as differs at 7 other sites compared to the China and Taiwan isolates. In the Thailand isolate, 18 tRNAs lack the entire TΨC-arm, compared to 17 in the China isolate and 16 in the Taiwan isolate. Phylogenetic analyses based on 36 mt-genes, 12 PCGs, 2 rRNA genes, 22 tRNA genes and control region all indicate closer genetic affinity between the China and Taiwan isolates compared to the Thailand isolate. Based on 36 mt-genes, the inter-isolate genetic distance varies from p=3.2% between China and Taiwan isolates to p=11.6% between Thailand and China isolates. The mitogenome will be useful for population, phylogenetics and phylogeography studies.
Fulltext Complete mitochondrial genome of Bactrocera arecae (Insecta: Tephritidae) by next-generation sequencing and molecular phylogeny of Dacini tribe

Yong HS, Song SL, Lim PE, Chan KG, Chow WL, Eamsobhana P

Sci Rep, 2015;5:15155.
PMID: 26472633 DOI: 10.1038/srep15155

The whole mitochondrial genome of the pest fruit fly Bactrocera arecae was obtained from next-generation sequencing of genomic DNA. It had a total length of 15,900 bp, consisting of 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes and a non-coding region (A + T-rich control region). The control region (952 bp) was flanked by rrnS and trnI genes. The start codons included 6 ATG, 3 ATT and 1 each of ATA, ATC, GTG and TCG. Eight TAA, two TAG, one incomplete TA and two incomplete T stop codons were represented in the protein-coding genes. The cloverleaf structure for trnS1 lacked the D-loop, and that of trnN and trnF lacked the TΨC-loop. Molecular phylogeny based on 13 protein-coding genes was concordant with 37 mitochondrial genes, with B. arecae having closest genetic affinity to B. tryoni. The subgenus Bactrocera of Dacini tribe and the Dacinae subfamily (Dacini and Ceratitidini tribes) were monophyletic. The whole mitogenome of B. arecae will serve as a useful dataset for studying the genetics, systematics and phylogenetic relationships of the many species of Bactrocera genus in particular, and tephritid fruit flies in general.
Fulltext Complete Mitochondrial Genome of Three Bactrocera Fruit Flies of Subgenus Bactrocera (Diptera: Tephritidae) and Their Phylogenetic Implications

Yong HS, Song SL, Lim PE, Eamsobhana P, Suana IW

PLoS One, 2016;11(2):e0148201.
PMID: 26840430 DOI: 10.1371/journal.pone.0148201

Bactrocera latifrons is a serious pest of solanaceous fruits and Bactrocera umbrosa is a pest of Artocarpus fruits, while Bactrocera melastomatos infests the fruit of Melastomataceae. They are members of the subgenus Bactrocera. We report here the complete mitochondrial genome of these fruit flies determined by next-generation sequencing and their phylogeny with other taxa of the subgenus Bactrocera. The whole mitogenomes of these three species possessed 37 genes namely, 13 protein-coding genes (PCGs), 2 rRNA and 22 tRNA genes. The mitogenome of B. latifrons (15,977 bp) was longer than those of B. melastomatos (15,954 bp) and B. umbrosa (15,898 bp). This difference can be attributed to the size of the intergenic spacers (283 bp in B. latifrons, 261 bp in B. melastomatos, and 211 bp in B. umbrosa). Most of the PCGs in the three species have an identical start codon, except for atp8 (adenosine triphosphate synthase protein 8), which had an ATG instead of GTG in B. umbrosa, whilst the nad3 (NADH dehydrogenase subunit 3) and nad6 (NADH dehydrogenase subunit 6) genes were characterized by an ATC instead of ATT in B. melastomatos. The three species had identical stop codon for the respective PCGs. In B. latifrons and B. melastomatos, the TΨC (thymidine-pseudouridine-cytidine)-loop was absent in trnF (phenylalanine) and DHU (dihydrouracil)-loop was absent in trnS1 (serine S1). In B. umbrosa, trnN (asparagine), trnC (cysteine) and trnF lacked the TψC-loop, while trnS1 lacked the DHU-stem. Molecular phylogeny based on 13 PCGs was in general concordant with 15 mitochondrial genes (13 PCGs and 2 rRNA genes), with B. latifrons and B. umbrosa forming a sister group basal to the other species of the subgenus Bactrocera which was monophyletic. The whole mitogenomes will serve as a useful dataset for studying the genetics, systematics and phylogenetic relationships of the many species of Bactrocera genus in particular, and tephritid fruit flies in general.

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