Genetic diversity and population structure of Haemonchus contortus in goats from Thailand.

Haemonchus contortus is one of the most economically important parasitic nematodes affecting small ruminant livestock worldwide. This study was conducted to elucidate the genetic diversity and population structure of this nematode in Thailand based on mitochondrial DNA markers, the nicotinamide dehydrogenase subunit 4 (nad4) and the cytochrome c oxidase subunit 1 (cox1) genes. One hundred and thirty-six adult worms were obtained from 86 abomasa of slaughtered goats from 13 different localities in 5 regions of Thailand. Identification to the genus Haemonchus was done using morphology. DNA sequences of the nuclear ribosomal second internal transcribed spacer (ITS2) identified each specimen to species: three fixed nucleotide (SNP) differences distinguished H. contortus from H. placei. Genetic analysis defined 118 and 122 unique haplotypes in partial sequences of nad4 (alignment length 723 bp) and cox1 (645 bp) genes, respectively. Nucleotide diversities were 0.031 and 0.043 for nad4 and cox1 genes, respectively. Low genetic differentiation was observed among H. contortus samples from various provinces in Thailand. This is the first study on the genetic diversity and population structure of H. contortus of goats in Thailand. This study has provided insights into the transmission dynamics of this parasitic nematode, information which is essential for farm management and parasite control.

The first molecular identification of benzimidazole resistance in Haemonchus contortus from goats in Thailand.

BACKGROUND AND AIM: Haemonchus contortus is one of the major trichostrongyloid nematodes affecting small ruminant production worldwide, especially in tropical and subtropical regions. Adult H. contortus suck the blood from the host abomasum leading to anemia and often death in heavily infected animals. The mainstay of parasitic control is an anthelmintic drug, but long-term drug use may cause drug resistance. The aim of this study was to examine benzimidazole resistance in H. contortus of goats from different regions in Thailand by detecting the frequency of the F200Y polymorphism in the ß-tubulin isotype 1 gene. MATERIALS AND METHODS: A total of 121 H. contortus adults were obtained from 31 naturally infected out of 37 slaughtered goats from city abattoirs in five regions of Thailand. The frequency of the F200Y polymorphism in the ß-tubulin isotype 1 gene was detected following the allele-specific polymerase chain reaction protocol. RESULTS: The overall genotype frequencies in Thailand were homozygous resistant (RR: 24%), heterozygous (SR: 44.6%), and homozygous susceptible (SS: 31.4%). The allele frequencies were resistant allele (R: 46%) and susceptible allele (S: 54%). The R allele frequency and the RR genotype varied from 30% to 65% and 0% to 43.9%, respectively. The frequency of R alleles was significantly higher in the southern region (0.65) as compared to northern (0.30, p=0.001), western (0.38, p=0.04), and central regions (0.30, p=0.03). The RR genotype was also significantly higher in the southern region (43.9%) versus the northern (0 %, p=0.001), western (11.8%, p=0.012), and central regions (17.4%, p=0.001). CONCLUSION: This is the first study of the detection of single-nucleotide polymorphisms in codon 200 of the ß-tubulin isotype 1 gene of H. contortus from goats in Thailand. These findings are essential and imply that an integrated approach is needed for issues such as drug treatment, farm management, prevention, and control strategies. This is of interest to farmers, veterinarians, and the department of livestock.

Effects of aestivation on survival of Bithynia siamensis goniomphalos snails and the infection of Opisthorchis viverrini in the irrigation area of wet- and dry-season rice paddy.

The snail Bithynia siamensis goniomphalos acts as the first intermediate host of the human liver fluke, Opisthorchis viverrini, a major cause of cholangiocarcinoma in Northeast Thailand. In this study, we investigated host-parasite interactions in wet- and dry-season rice paddy plantation. The snails that aestivated through 4-month-dried period after rice harvest had average mortality rate of 36.45% and monthly increasing from 17.76% to 54.57%. Surprisingly, at depths greater than 5 cm experienced a higher mortality rate than upper level. Average mortality rate at the depth more than 5 cm, was 42.97%. In the initial month of aestivation, mortality rate of 0-5 and 5-10 cm depth was 9.13% and 26.39% then increase to 57.58% and 51.97%, respectively in the last 4th month. The average prevalence O. viverrini infection in snails during the study period was 0.44%. The highest prevalence of O. viverrini infection was found in the cool dry and wet season. The odds of O. viverrini infection in female snails and large snails were higher relative to male snails and medium-sized snails. The physicochemical factors involved in increased prevalence of O. viverrini infection in snail hosts were mean daily air temperature and average monthly rainfall. Short aestivation period made lower mortality in irrigation area than the previous report of non-irrigation area, that is, the aestivation is one of snail population control.

Effects of physicochemical factors on development and survival of Opisthorchis viverrini uterine eggs

Objective: To investigate the maturity development of miracidia in uterine eggs from four portions of the Opisthorchis viverrini uterus and environmental factors possibly affected in maturation and infectivity of distal part uterine eggs. Methods: Uteri of adult worms were divided into 4 equal parts. Development of eggs was determined under light microscope. Only embryonated eggs were used to evaluate the effects of physicochemical factors: temperature, salinity, acidity, ultraviolet A, B, C. Infection success was evaluated by feeding treated eggs to intermediate host snails and determining by using a PCR approach. Results: Eggs obtained from the uterus closest to the ovary (regions 1 and 2) failed to develop in vitro. Eggs from region 4 of the uterus (close to the genital pore) were used to study effects of physicochemical factors. The highest survival and infection success was in groups of eggs kept at 30 ℃ (95.20％). The calculated period of loss infection success (LI50 and LI95) on miracidia in distal uterine eggs by exposure to UV-A, UV-B and UV-C were 73 and 1523 d; 8 and 20 d; 1 and 2 d, respectively. Lethal concentrations (LC50 and LC95) of salinity, HCl and NaOH on miracidia in distal uterine eggs were 45.43 and 120.09 ppt, 0.01 and 0.25 M; 0.01 and 0.11 M, respectively, after 24 h exposure. Conclusions: Opisthorchis viverrini eggs display a high tolerance to environmental conditions, especially after snail host eating for infection.

Effects of albendazole, artesunate, praziquantel and miltefosine, on Opisthorchis viverrini cercariae and mature metacercariae.

OBJECTIVE: To explore larvicidal effects of anthelmintic drugs on Opisthorchis viverrini (O. viverrini) for alternative approach to interrupting its cycle for developing a field-based control program. METHODS: The larvicidal activities of albendazole (Al), artesunate (Ar), praziquantel (Pzq) and miltefosine (Mf) on O. viverrini cercariae and mature metacercariae were investigated. Lethal concentrations (LC50 and LC95) of these drugs were determined. Mature metacercariae previously exposed to various concentrations of the drugs were administered to hamsters. Worms were harvested 30 d post infection and worm recovery rates calculated. Al, Ar, Pzq and Mf produced morphological degeneration and induced shedding tails of cercariae after 24 h exposure. RESULTS: The LC50 and LC95 of Al, Ar, Pzq and Mf on cercariae were 0.720 and 1.139, 0.350 and 0.861, 0.017 and 0.693, and 0.530 and 1.134 ppm, respectively. LC50 and LC95 of Ar on mature metacercariae were 303.643 and 446.237 ppm and of Mf were 289.711 and 631.781 ppm, respectively but no lethal effect in Pzq- and Al-treated groups (up to 1 ppt). No worms were found in hamsters administered Pzq-treated metacercariae. The adult worms from Al-treated metacercariae were significantly bigger in size compared to the control group (P < 0.05). Fecundity and body width were greater in adults from Mf-treated metacercariae compared to the control group (P < 0.05). CONCLUSIONS: The larvicidal effects of these drugs were high efficacy to O. viverrini cercariae but lesser efficacy to metacercariae. It should be further studied with the eventual aim of developing a field-based control program.