Conifer defence against insects: microarray gene expression profiling of Sitka spruce (Picea sitchensis) induced by mechanical wounding or feeding by spruce budworms (Choristoneura occidentalis) or white pine weevils (Pissodes strobi) reveals large

Conifers are resistant to attack from a large number of potential herbivores or pathogens. Previous molecular and biochemical characterization of selected conifer defence systems support a model of multigenic, constitutive and induced defences that act on invading insects via physical, chemical, biochemical or ecological (multitrophic) mechanisms. However, the genomic foundation of the complex defence and resistance mechanisms of conifers is largely unknown. As part of a genomics strategy to characterize inducible defences and possible resistance mechanisms of conifers against insect herbivory, we developed a cDNA microarray building upon a new spruce (Picea spp.) expressed sequence tag resource. This first-generation spruce cDNA microarray contains 9720 cDNA elements representing c. 5500 unique genes. We used this array to monitor gene expression in Sitka spruce (Picea sitchensis) bark in response to herbivory by white pine weevils (Pissodes strobi, Curculionidae) or wounding, and in young shoot tips in response to western spruce budworm (Choristoneura occidentalis, Lepidopterae) feeding. Weevils are stem-boring insects that feed on phloem, while budworms are foliage feeding larvae that consume needles and young shoot tips. Both insect species and wounding treatment caused substantial changes of the host plant transcriptome detected in each case by differential gene expression of several thousand array elements at 1 or 2 d after the onset of treatment. Overall, there was considerable overlap among differentially expressed gene sets from these three stress treatments. Functional classification of the induced transcripts revealed genes with roles in general plant defence, octadecanoid and ethylene signalling, transport, secondary metabolism, and transcriptional regulation. Several genes involved in primary metabolic processes such as photosynthesis were down-regulated upon insect feeding or wounding, fitting with the concept of dynamic resource allocation in plant defence. Refined expression analysis using gene-specific primers and real-time PCR for selected transcripts was in agreement with microarray results for most genes tested. This study provides the first large-scale survey of insect-induced defence transcripts in a gymnosperm and provides a platform for functional investigation of plant-insect interactions in spruce. Induction of spruce genes of octadecanoid and ethylene signalling, terpenoid biosynthesis, and phenolic secondary metabolism are discussed in more detail.

The complete mitochondrial genome of the gall-forming fly, Fergusonina taylori(Nelson and Yeates) (Diptera: Fergusoninidae)

The monogeneric family Fergusoninidae consists of gall-forming flies that, together with Fergusobia (Tylenchida: Neotylenchidae) nematodes, form the only known mutualistic association between insects and nematodes. In this study, the entire 16,000 bp mitochondrial genome of Fergusonina taylori Nelson and Yeates was sequenced. The circular genome contains one encoding region including 27 genes and one non-coding A þT-rich region. The arrangement of the proteincoding, ribosomal RNA (rRNA) and transfer RNA (tRNA) genes was the same as that found in the ancestral insect. Nucleotide composition is highly A þ T biased. All of the protein initiation codons are ATN, except for nad1 which begins with TTT. All 22 tRNA anticodons of F. taylori match those observed in Drosophila yakuba, and all form the typical cloverleaf structure except for tRNA-Ser (AGN) which lacks a dihydrouridine (DHU) arm. Secondary structural features of the rRNA genes of Fergusonina are similar to those proposed for other insects, with minor modifications. The mitochondrial genome of Fergusonina presented here may prove valuable for resolving the sister group to the Fergusoninidae, and expands the available mtDNA data sources for acalyptrates overall.

Evolutionary genetics and human assisted movement of a globally invasive pest (Russian wheat aphid : Diuraphis noxia)

This PhD study has examined the population genetics of the Russian wheat aphid (RWA, Diuraphis noxia), one of the world’s most invasive agricultural pests, throughout its native and introduced global range. Firstly, this study investigated the geographic distribution of genetic diversity within and among RWA populations in western China. Analysis of mitochondrial data from 18 sites provided evidence for the long-term existence and expansion of RWAs in western China. The results refute the hypothesis that RWA is an exotic species only present in China since 1975. The estimated date of RWA expansion throughout western China coincides with the debut of wheat domestication and cultivation practices in western Asia in the Holocene. It is concluded that western China represents the limit of the far eastern native range of this species. Analysis of microsatellite data indicated high contemporary gene flow among northern populations in western China, while clear geographic isolation between northern and southern populations was identified across the Tianshan mountain range and extensive desert regions. Secondly, this study analyzed the worldwide pathway of invasion using both microsatellite and endosymbiont genetic data. Individual RWAs were obtained from native populations in Central Asia and the Middle East and invasive populations in Africa and the Americas. Results indicated two pathways of RWA invasion from 1) Syria in the Middle East to North Africa and 2) Turkey to South Africa, Mexico and then North and South America. Very little clone diversity was identified among invasive populations suggesting that a limited founder event occurred together with predominantly asexual reproduction and rapid population expansion. The most likely explanation for the rapid spread (within two years) from South Africa to the New World is by human movement, probably as a result of the transfer of wheat breeding material. Furthermore, the mitochondrial data revealed the presence of a universal haplotype and it is proposed that this haplotype is representative of a wheat associated super-clone that has gained dominance worldwide as a result of the widespread planting of domesticated wheat. Finally, this study examined salivary gland gene diversity to determine whether a functional basis for RWA invasiveness could be identified. Peroxidase DNA sequence data were obtained for a selection of worldwide RWA samples. Results demonstrated that most native populations were polymorphic while invasive populations were monomorphic, supporting previous conclusions relating to demographic founder effects in invasive populations. Purifying selection most likely explains the existence of a universal allele present in Middle Eastern populations, while balancing selection was evident in East Asian populations. Selection acting on the peroxidase gene may provide an allele-dependent advantage linked to the successful establishment of RWAs on wheat, and ultimately their invasion potential. In conclusion, this study is the most comprehensive molecular genetic investigation of RWA population genetics undertaken to date and provides significant insights into the source and pathway of global invasion and the potential existence of a wheat-adapted genotype that has colonised major wheat growing countries worldwide except for Australia. This research has major biosecurity implications for Australia’s grain industry.

The mitochondrial genome of the Russian wheat aphid Diuraphis noxia : large repetitive sequences between trnE and trnF in aphids

To characterize aphid mitochondrial genome (mitogenome) features, we sequenced the complete mitogenome of the Russian wheat aphid, Diuraphis noxia. The 15,784-bp mitogenome with a high A + T content (84.76%) and strong C skew (− 0.26) was arranged in the same gene order as that of the ancestral insect. Unlike typical insect mitogenomes, D. noxia possessed a large tandem repeat region (644 bp) located between trnE and trnF. Sequencing partial mitogenome of the cotton aphid (Aphis gossypii) further confirmed the presence of the large repeat region in aphids, but with different repeat length and copy number. Another motif (58 bp) tandemly repeated 2.3 times in the control region of D. noxia. All repeat units in D. noxia could be folded into stem-loop secondary structures, which could further promote an increase in copy numbers. Characterization of the D. noxia mitogenome revealed distinct mitogenome architectures, thus advancing our understanding of insect mitogenomic diversities and evolution.

A multi-genome analysis approach enables tracking of the invasion of a single Russian wheat aphid (Diuraphis noxia) clone throughout the New World

This study investigated the population genetics, demographic history and pathway of invasion of the Russian wheat aphid (RWA) from its native range in Central Asia, the Middle East and Europe to South Africa and the Americas. We screened microsatellite markers, mitochondrial DNA and endosymbiont genes in 504 RWA clones from nineteen populations worldwide. Following pathway analyses of microsatellite and endosymbiont data, we postulate that Turkey and Syria were the most likely sources of invasion to Kenya and South Africa, respectively. Furthermore, we found that one clone transferred between South Africa and the Americas was most likely responsible for the New World invasion. Finally, endosymbiont DNA was found to be a high resolution population genetic marker, extremely useful for studies of invasion over a relatively short evolutionary history time frame. This study has provided valuable insights into the factors that may have facilitated the recent global invasion by this damaging pest.