Ross H. Andrews

Opisthorchis viverrini: an underestimated parasite in world health

Infection with Opisthorchis viverrini and its associated cholangiocarcinoma (CCA) is an underestimated problem in the Mekong region of Southeast Asia, despite the widespread use of praziquantel and health education measures for parasite control. Although data from Cambodia, Laos and Vietnam are rare, data from Thailand often show wide-ranging variability in epidemiological parameters, including human morbidity and the prevalence and incidence of CCA. The recent discovery of high levels of population genetic variability in O. viverrini in different wetlands in Thailand and Laos, which indicates the presence of sibling species, suggests that we have underestimated the complexity of this epidemiological situation. Future research should determine the relationship between the genetic variability of O. viverrini and patterns of opisthorchiasis-related disease.

Mitochondrial DNA sequence variation among geographical isolates of Opisthorchis viverrini in Thailand and Lao PDR, and phylogenetic relationships with other trematodes

The present study compared the genetic variation among 14 different geographical isolates of Opisthorchis viverrini sensu lato from Thailand and Lao PDR using sequence data for 2 mitochondrial DNA genes, the subunit 1 of NADH dehydrogenase gene (nad1) and cytochrome c oxidase gene (cox1). Four different nad1 haplotypes were detected among isolates, all of which were identical at the amino acid sequence level. Nucleotide sequence variation among 14 isolates ranged from 0 to 0.3% for nad1. Two different cox1 haplotypes were detected among isolates. These two haplotypes differed at 2 nucleotide positions, one of which resulted in a change in the amino acid sequence. Nucleotide sequence variation among isolates for cox1 ranged from 0 to 0.5%. Comparison of cox1 sequences of O. viverrini to those of other trematodes revealed nucleotide differences of 13-31%. A phylogenetic analysis of the cox1 sequence data revealed strong statistical support for a clade containing O. viverrini and 2 other species of opisthorchid trematodes; O. felineus and Clonorchis sinsensis.

Genetic variation in Opisthorchis viverrini (Trematoda: Opisthorchiidae) from northeast Thailand and Laos PDR based on random amplified polymorphic DNA analyses.

Genetic variation in Opisthorchis viverrini adults originating from different locations in northeast Thailand and Laos, People’s Democratic Republic (PDR), was examined using random amplified polymorphic DNA (RAPD) analyses. In an initial analysis, the genomic DNA of one fluke from each of ten localities was amplified using 15 random primers (10-mers); however, genetic variation among O. viverrini specimens was detected reliably for only four primers. A more detailed RAPD analysis using these four primers was conducted on ten individuals from nine localities. Considerable genetic variation was detected among O. viverrini from different geographical areas and among some individuals from the same collecting locality. Comparison of the RAPD profiles revealed that O. viverrini adults from Laos PDR were genetically distinct from those from northeast Thailand. The taxonomic significance of this finding needs to be explored in more detail. The RAPD markers established in the present study provide opportunities to examine the biology and epidemiology of O. viverrini and fish-borne trematodes within the region. Additionally, application of these markers in such studies could have important implications in relation to the prevalence of cholangiocarcinoma in different regions of Asia.

Microsatellite loci in the carcinogenic liver fluke, Opisthorchis viverrini and their application as population genetic markers

Opisthorchis viverrini is a carcinogenic foodborne trematode endemic in Southeast Asia especially in Thailand and the Lao Peoples Democratic Republic. Opisthorchiasis causes hepatobiliary diseases and cholangiocarcinoma (bile duct cancer). Currently there is substantial evidence on genetic variation of O. viverrini but the information on population genetic structure is lacking. Because microsatellite DNA of this parasite is not available, we for the first time isolated and utilized microsatellite DNA as genetic markers to examine genetic diversity and the population structure of O. viverrini. Partial genomic DNA libraries were constructed by conventional and enrichment methods which yielded microsatellite-containing clones of 0.18-0.25% and 16.84%, respectively. Within 41 microsatellite loci isolated 36.59% were perfect, 60.98% were interrupted and 2.44% were compound microsatellites. The CA repetitions were the most frequent, followed by GT and CAT. Primers specific to the flanking regions of 12 microsatellite loci were developed to genotype 150 O. viverrini individuals from geographical localities in Thailand and Lao PDR. Allele numbers per locus ranged from 2 to 15, with the mean expected heterozygosity of 0.03-0.66. Analyses of O. viverrini from 5 localities revealed a high level of genetic diversity and had significant deviation from Hardy-Weinberg equilibrium. Significant heterozygote deficiency as well as heterozygote excess was detected across all localities indicating the possibility of selfing (inbreeding) as a predominant reproductive mode. Significant genetic differentiation (F(ST)) was also detected between worms from different localities with varying levels of genetic heterogeneity. We discuss our results in terms of what these novel microsatellite markers reveal about the epidemiology and transmission dynamics of this medically important parasite, both in terms of the current study and their potential for future comprehensive population genetic studies O. viverrini sensu lato in Southeast Asia.

Dams and disease triggers on the lower Mekong river.

Ongoing and proposed construction of several large hydropower dams along the mainstream Mekong River and various tributaries has created a number of unanswered environmental and societal questions for governments and communities in Cambodia, China, Lao PDR, Myanmar, Thailand, and Vietnam [1]–[3]. Most concern over the controversial dam-building projects focuses on the extent to which river health and food security will be affected negatively. Foremost, the 85 or more proposed dams threaten to reduce the diversity and abundance of freshwater fish, the major animal protein source for many of the 67 million inhabitants of the Mekong River basin [4]–[7].

Cohort profile: cholangiocarcinoma screening and care program (CASCAP)

BackgroundCholangiocarcinoma (CCA) is an extremely aggressive cancer that is usually fatal. Although globally morbidity and mortality are increasing, knowledge of the disease remains limited. The Mekong region of Southeast Asia, and particularly the northeast of Thailand, has by far the highest incidence of CCA worldwide with 135.4 per 100,000 among males and 43.0 per 100,000 among females being reported in Khon Kaen Province. Most patients are first seen during late stage disease with 5-year survival being less than 10xa0%. Starting in 1984, control and prevention strategies have been focused on health education. Although early detection can substantially increase 5-year survival, there are currently no strategies to increase early diagnosis.Methods/designThe Cholangiocarcinoma Screening and Care Program (CASCAP) is a prospective cohort study comprising two cohorts- the screening and the patient cohorts. For the screening cohort, ultrasound examination will be carried out regularly at least annually to determine whether there is current bile duct and/or liver pathology so that the optimal screening program for early diagnosis can be established. This cohort is expected to include at least 150,000 individuals coming from high-risk areas for CCA. For the patient cohort, it is estimated that about 25,000 CCA patients will be included during the 5-year recruitment period. All CCA patients will be treated according to routine clinical care and followed so that effective surgical treatment can be formulated. This cohort is indeed a conventional cancer registry. Thus, CASCAP is an ongoing project in which the number of participants changes dynamically.DiscussionsThis is the first project on CCA that involves screening the at risk population at the community level. At the time of preparing this report, a total of 85,927 individuals have been enrolled in the screening cohort, 55.0xa0% of whom have already undergone ultrasound screening, and 2661 CCA cases have been enrolled in the patient cohort. Among the participants of the screening, whose mean age was 53.8u2009±u20099.8xa0years, 55.6xa0% were female, 77.5xa0% attained primary school as the highest level of education, 79.9xa0% were farmers, 29.9xa0%, reported having relatives with CCA, 89.1xa0% had eaten uncooked fish, and 42.2xa0% of those who had been tested for liver fluke were found to be infected.

Genetic markers to distinguish Moniezia expansa from M. benedeni (Cestoda: Anoplocephalidae) and evidence of the existence of cryptic species in Australia

Multilocus enzyme electrophoresis (MEE) was used to genetically compare Moniezia spp. collected from sheep and cattle in southern Australia. Fifteen enzyme loci were established for the genetic characterisation of individual specimens of Moniezia, which represents a significant increase in the number of loci that was previously available for these cestodes. Another four enzyme loci were also identified as providing potential genetic markers. There were no differences in staining activity or banding patterns of different body regions of individual cestodes. Fixed genetic differences between Moniezia expansa and Moniezia benedeni were detected at 10 (77%) loci. Individuals that lacked the diagnostic morphological characters (i.e. the arrangement pattern of the interproglottidal glands) could be assigned to either morphospecies based on their MEE profiles. Genetic differences were also detected among individuals of the two morphospecies. One specimen of M. expansa had fixed genetic differences at 33% of enzyme loci when compared to all other specimens of M. expansa. Fixed genetic differences were also detected between the two groups of M. benedeni at 12 of 13 (92%) enzyme loci. This magnitude of genetic difference was greater than that detected between the two morphospecies. This provides evidence for the existence of cryptic species in the genus Moniezia.

Impact of temporal changes and host factors on the genetic structure of a population of Opisthorchis viverrini sensu lato in Khon Kaen Province (Thailand)

The population genetics of 317 individual Opisthorchis viverrini from Khon Kaen Province Thailand, from 4 different years and 4 cyprinid fish species was examined using multilocus enzyme electrophoresis of enolase (Enol), phosphoglucomutase (Pgm) and triose phosphate isomerase (Tpi). Allele and genotype frequencies for Enol and Pgm were consistent irrespective of year or host species. No heterozygote deficiency was detected for Enol. Significant heterozygote deficiencies were detected in 3 of 4 years for Pgm. For Tpi, allele frequencies of the most common allele and genotype frequency varied between years and among individuals from different host species. Heterozygote deficiencies for Tpi were detected in 2 years. No significant heterozygous deficiencies were detected among O. virerrini from different fish species in 2005, except at Pgm and Tpi from Puntioplites protozsron. There was no statistical significance in pairwise FST values between O. viverrini from Cyclocheilichthys armatus in different years or different host species in 2005. Significant departures from Hardy-Weinberg expectations and a high rate of gene flow in a population of O. viverrini are discussed in terms of self- and cross-fertilisation, natural selection, non-random mating, the Wahlund effect, presence of null alleles, intensity of infection, biology and ecology of their intermediate cyprinid hosts.

Opisthorchis viverrini: Implications of the systematics of first intermediate hosts, Bithynia snail species in Thailand and Lao PDR.

Bithynia snails are important hosts of the human liver fluke, Opisthorchis viverrini. Allozyme markers were used to examine genetic variation and the systematics of Bithynia snails from different geographical localities in Thailand and Lao PDR. Our results show that Bithynia siamensis goniomphalos contains two major evolutionary clades with at least nine morphologically similar but genetically distinct cryptic species (taxa). Clade A consists of snails from the Chi and the Mun River wetlands, Thailand and clade B consists of snails from the Nam Ngum River wetland, Lao PDR, as well as snails from some localities in the Songkram River wetland, Thailand. There is an association between O. viverrini cryptic taxa with B. s. goniomphalos cryptic taxa within the defined wetlands. Bithynia species are the critical amplifying component of O. viverrini and they control transmission from humans to fish second intermediate hosts. These findings are particularly important for understanding the transmission dynamics of O. viverrini. They are, in addition, a link in the life cycle of O. viverrini which can be used for developing and implementing programs to eliminate and control opisthorchiasis and its associated cholangiocarcinoma.

Argas persicus sensu stricto does occur in Australia

Abstract The status of Argas persicus in Australia has been a matter of controversy for over 25xa0years. Although early records of A. persicus sensu stricto are common, a major revision of the genus indicated that these records refer to A. robertsi, first described in 1968 from northern Australia, and to an “undescribed” member of the complex occurring in the south. Here, we show that A. persicus sensu stricto does occur in southern Australia and is the only species of Argas in the area recorded from poultry. Another undescribed species belonging to the A. persicus complex, from crows’ nests near Lake Eyre in South Australia, was also discovered. This information is of considerable epidemiological significance, as A. persicus sensu stricto is a major vector for a number of highly pathogenic diseases of poultry, not all of which have yet been recorded from Australia.