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Title: Population genetic structure of Fasciola hepatica in Great Britain
Author: Beesley, Nicola Jane
ISNI:       0000 0004 6058 9128
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2016
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The liver fluke, Fasciola hepatica, is a trematode parasite that causes disease of economic and welfare importance to the UK livestock sector. Prevalence of F. hepatica infection in sheep and cattle in the UK is increasing; in part due to changes in climate and farming practice, but compounded by the emergence of resistance to the drug triclabendazole. Adult F. hepatica within the definitive host are hermaphrodite, capable of both self- and cross-fertilisation. There are reports of parasite populations maintained by parthenogenesis. Diploid (2n = 2x = 20) and triploid (2n = 3x = 30) F. hepatica have been reported in the UK, but there is little information to indicate how frequently triploidy occurs. The F. hepatica life cycle is complex requiring a snail intermediate host, Galba truncatula. Clonal expansion of the parasite occurs within the snail followed by release of cercariae which subsequently encyst on pasture. Little is known about the population genetic structure of F. hepatica in the UK, or how the clonal expansion in the snail, the capacity for self-fertilisation, and the potential for aggregation of metacercariae on pasture, influence genetic diversity of F. hepatica populations within the definitive host. Knowledge of the genetic structure of F. hepatica in Great Britain is critical to understanding the level of gene flow within F. hepatica populations and how this impacts on the spread of drug resistance genes. The aim of this thesis was to determine the ploidy and population genetic structure of F. hepatica infecting sheep and cattle in Great Britain, and define how passage of the parasite through the snail intermediate host may influence genetic diversity. To determine ploidy a total of 715 adult parasites were collected from naturally infected sheep and cattle, exceeding the statistically representative sample size of 384 individuals (expected proportion of triploids 50 %; 95 % confidence level; 5 % confidence limits). The ploidy and presence of sperm was determined by aceto-orcein squash. One hundred percent were confirmed as diploid and all contained sperm. A multiplex PCR and capillary electrophoresis approach was validated for a panel of fifteen polymorphic microsatellites to assess the population genetic structure of F. hepatica. Eight of these microsatellites met the criteria for population genetic analyses, and were used to produce a multilocus genotype (MLG) for 950 adult parasites from the livers of 44 naturally infected sheep, and 629 F. hepatica adults from 31 naturally infected cattle livers. The average heterozygosity across all parasites and loci was determined as 0.752 (SD = 0.130), indicating high levels of genetic variation in F. hepatica populations. Of the 1579 samples, 1424 distinct MLGs were observed, which supports high genotypic diversity in the population as a whole. Forty six hosts harboured multiple, genotypically identical parasites, which were shown to have arisen from clonal lineages rather than distinct reproductive events. The FIS value indicates a self-fertilisation rate no higher than 2 %, and the FST value (0.0202) indicates low population structure and high gene flow in the British F. hepatica population. To assess the influence of the snail intermediate host on genetic diversity, MLG profiles were used to address: (i) whether a snail can be infected with, and shed cercariae of, more than one genotype of F. hepatica and (ii) the population structure of F. hepatica within field populations of snails in Great Britain. For experiment (i) 49 G. truncatula, from a colony maintained in the laboratory, were exposed to miracidia of known, but distinct, genotype. Two snails, showed evidence of infection with more than one genotype of F. hepatica, but did not shed cercariae. In experiment (ii) 155 snails, including 52 G. truncatula, were collected from three farms in Scotland and North Wales. None of the snails showed evidence of infection with F. hepatica. The results here show that F. hepatica isolated from British cattle and sheep are diploid. In addition, genetic analysis suggests cross-fertilisation predominates over self-fertilisation and parthenogenesis. The British F. hepatica population shows high genetic diversity within populations, but little genetic differentiation between populations, and is therefore panmictic. Our analyses support high gene flow and identified a proportion of parasites within the definitive host that are genetically identical. These observations suggest that drug resistance genes could spread rapidly in F. hepatica populations.
Supervisor: Hodgkinson, J. ; Paterson, S. ; Williams, D. J. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral