Fig 1.
A flow diagram illustrating the methods used for fecal examination, Opisthorchis-like egg positive samples and available fecal samples for DNA extraction.
Fig 2.
The PCR-RFLP products of O. viverrini after digestion with FuaI restriction enzyme.
Amplicons of 247 bp and 129 bp from ITS2-PCR products of O. viverrini were generated after digestion. M was 100 bp DNA ladder, P1 was undigested positive and P2 was digested PCR products of positive samples. Numbers 1–6 showed undigested and digested ITS2-PCR products from fecal samples.
Table 1.
The prevalences of Opisthorchis-like egg, ITS2-PCR sensitivities and PCR-RFLP profiles for detection Opisthorchis-like eggs in Chachoengsao Province, central Thailand.
Table 2.
The indicators of genetic diversity and neutrality tests in the populations of O. viverrini from three villages using cox1 sequences.
Table 3.
The indicators of genetic diversity and neutrality tests in the populations of O. viverrini from three villages using nad1 sequences.
Fig 3.
The haplotype networks of cox1 and nad1 sequences of O. viverrini isolated from three villages in central Thailand, reference isolates from Thailand, Lao PDR, Vietnam and Cambodia.
(A) The haplotype network of 29 haplotypes based on cox1 sequences. Haplotype M contains 20 isolates and shows highest frequency; seven isolates from Thoong-Heang Village (TH), 12 isolates from Na-Ngam Village (NN), and one isolate from Lao PDR. Twenty two singletons are obtained as shown in smallest circles; 10 singletons (Haplotype B, D, E, F, G, H, I, J, K and L) from Na-Yao Village (NY), seven singletons (Haplotype N, O, P, Q, R, S, and T) from Na-Ngam Village and five singletons (V, W, Y Z and A1) from Thoong-Heang Village. The reference isolates from Thailand and Lao PDR are grouped in Haplotype B1 and C1. (B) The haplotype network of 31 haplotypes using nad1 sequences. Haplotype G presents the highest frequency of 9 isolates; 7 isolates from Na-Ngam Village, one isolate from Na-Yao Village and one isolate from Lao PDR. Twenty singletons are demonstrated; seven singletons (Haplotype B, C, D, E, F, H and I) from Na-Yao Village, three singletons (Haplotype I, K, M and N) from Thoong-Heang Village, four singletons (Haplotype O, P R and S) from Na-Ngam Village, five singletons (Haplotype T, U, V, Y and Z) from Vietnam and one singletons (Haplotype E1) isolate from Lao PDR. The reference isolates from Thailand, Lao PDR, Vietnam and Cambodia are clustered in Haplotype X, A1, B1, C1 and D1. Each open circle represents each haplotype and the numbers of each haplotype were placed in parentheses. Each singleton is marked with the names for villages and isolates. The solid line shows the network relationship of the haplotypes. The frequency of nucleotide change between haplotypes is shown as black bullets.
Fig 4.
The Randomized Axelerated Maximum Likelihood tree based on the cox1 sequences of O. viverrini.
The alignment of 393 nucleotide sequences without gaps and 27 variants from three villages (NY = Na-Yao, NN = Na-Ngam, TH = Thoong-Heang) were analyzed. O. viverrini Thai isolates (CP, BR, KBp, KPv, SK, NG, MS, KLp, KS) and Lao PDR isolates (SV, VT, O. viverrini mitochondrion partial genome) were used as the reference isolates. The percentages of 1,000 replications (bootstraps) of more than 50% are shown at the nodes. The number of samples is in parentheses.
Fig 5.
The Randomized Axelerated Maximum Likelihood tree based on the nad1 sequences of O. viverrini.
The alignment of 688 nucleotide sequences without gaps and 18 variants from three villages (NY = Na-Yao, NN = Na-Ngam, TH = Thoong-Heang) were analyzed. O. viverrini Thai isolates (BR, KS, LP, NP, KPv, MS, KLp, KBp, SK, NADH1), Lao PDR isolates (CP, NG, SV, VT, KM, O. viverrini mitochondrion partial genome), Vietnamese isolates (BD, OvBD1, OvL, PY3, OvPY3, QN, DL3) and a Cambodian isolate (KD) were used as the reference isolates. The percentages of 1,000 replications (bootstraps) of more than 50% are shown at the nodes. The number of samples is in parentheses.