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Development and optimisation of PCR-based techniques in predator gut analysis

Dodd, Ciara Siobhan 2004. Development and optimisation of PCR-based techniques in predator gut analysis. PhD Thesis, Cardiff University.

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Abstract

Pterostichus melanarius (Illiger) has been established as an important predator of slugs in the laboratory, semi-natural environments and in the field. The current method of choice for studying these predator-prey interactions is using monoclonal antibodies (MAbs) coupled with enzyme linked immunosorbent assay (ELISA). Recently, DNA-based detection methods have been suggested as a viable alternative in this area of research. DNA-based detection methods proved suitable for investigating predation by P. melanarius on economically damaging slug species. Mitochondrial DNA primers were designed for the 12S ribosomal RNA gene that were specific for Deroceras reticulatum, members of the Arion hortensis aggregate and the Arion genus. The detection limit and rate of decay of slug DNA in the beetle gut was determined using the slug-specific primers in laboratory-based feeding trials. Slug remains could be reliably detected within beetle guts for at least 24 hours following ingestion, suggesting that this technique would be suitable for detecting predation in the field. Direct comparison of immunological and DNA-based detection methods revealed that the sensitivity of MAb-ELISA is greater than amplification of prey DNA using the polymerase chain reaction (PCR). The implications of these findings are discussed. DNA-based detection methods were suitable for detecting predation by beetles on slugs in the field. The number of slug-positive beetles identified using MAb-ELISA was greater than when PCR and prey-specific primers were used. These results are discussed in the context of the availability of target and alternative prey in the field. Sequence data was also analysed to reveal the population structure and demographic history of A. hortensis aggregate in the British Isles. A high level of differentiation between populations was revealed. The rate of molecular evolution in these species is rapid with high levels of inter-specific divergence. The 12s rRNA gene proved especially useful in phylogenetic reconstruction and corroborated previous results based on morphology and enzyme electrophoresis that the congener, Arion intermedius belongs to the same subgenus (Kobelita) as the aggregate species.

Item Type: Thesis (PhD)
Status: Unpublished
Schools: Biosciences
Subjects: Q Science > QH Natural history > QH301 Biology
ISBN: 9781303157943
Date of First Compliant Deposit: 30 March 2016
Last Modified: 10 Jan 2018 00:52
URI: http://orca-mwe.cf.ac.uk/id/eprint/55533

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