Comparative transcriptome sequencing for identification of Australian pest fruit flies

A project undertaken at the School of Biological Earth and Environmental Sciences, University of New South Wales, and supervised by Kathryn Raphael

The tephritid or "true" fruit flies, Bactrocera tryoni (TRYONI) and B. neohumeralis (NEO), originally native to the rainforests of Queensland and Northern NSW, are now Australia’s worst horticultural pests (Figure 1), and a quarantine risk thoughout the Asia-Pacific region. Australian horticulture is also threatened by other native fruit flies, such as B. jarvisi. Furthermore, exotic flies from Papua New Guinea and SE Asia are serious quarantine risks for Australia. Accurate species identification is an imperative for pest control. This proposal makes use of new next-generation sequencing technologies to compare gene expression levels and gene sequence differences between NEO and TRYONI to a much greater depth than has been possible before.

TRYONI and NEO are extremely similar morphologically, ecologically and genetically (Morrow et al., 2000; Raphael et al., 2004). The overlapping distribution of the species on the east coast of Australia has been well characterised over nearly 100 years. However TRYONI but not NEO has spread south from an original endemic habitat – the rainforests of eastern Australia – into temperate fruit production areas. Hence TRYONI is regarded as a more serious pest than NEO. The key differentiating feature between the species is a strong pre-mating isolation mechanism in the wild, involving mating time – TRYONI mates in a narrow window of low light intensity at dusk, whereas NEO mates in bright light during the middle of the day. We have investigated the genetics of the difference in mating time and found that it relates to a difference in the amount of expression, rather than DNA sequence, of genes that control the endogenous biological (circadian) clock (An et al., 2004; Figure 2). However, DNA sequence differences must exist somewhere in the genomes of the two species. The complete genome sequence of TRYONI is currently being completed. In order to locate some of the crucial DNA changes that differentiate TRYONI and NEO, this project will examine all the genes expressed in the head (where the circadian clock is controlled) of the two species, and use differences in the level of expression between the species as a marker of key genes. Next generation sequencing is the method of choice as it can assay for both sequence differences and quantitative differences in gene expression. The genetic differences thus discovered will allow species identification tests to be developed and will be important for future studies of behaviour and pest status, leading to novel control strategies in the future.

References

An, X., Tebo, M., Sunmi, S. Frommer, M. and Raphael, K.A. (2004) The cryptochrome (cry) gene and a mating isolation mechanism in tephritid fruit flies. Genetics 168, 2025-2036.

Morrow, J., Scott, L., Congdon, B., Yeates, D., Frommer, M. and Sved, J. (2000) Close genetic similarity between two sympatric species of tephritid fruit fly reproductively isolated by mating time. Evolution 54, 899-910.

Raphael, K.A., Whyard, S., Shearman, D., An, X. and Frommer, M. (2004) Bactrocera tryoni and closely related pest tephritids – molecular background towards transgenesis. Insect Biochem. Mol. Biol. 34, 167-176.