Evaluation of endogenous references for gene expression profiling in different tissues of the oriental fruit fly Bactrocera dorsalis (Diptera: Tephritidae)

<p>Abstract</p> <p>Background</p> <p>Quantitative real-time reverse transcriptase PCR (RT-qPCR) has been widely used for quantification of mRNA as a way to determine key genes involved in different biological processes. For accurate gene quantification analysis, normalization of RT-qPCR data is absolutely essential. To date, normalization is most frequently achieved by the use of internal controls, often referred to as reference genes. However, several studies have shown that the reference genes used for the quantification of mRNA expression can be affected by the experimental set-up or cell type resulting in variation of the expression level of these key genes. Therefore, the evaluation of reference genes is critical for gene expression profiling, which is often neglected in gene expression studies of insects. For this purpose, ten candidate reference genes were investigated in three different tissues (midgut, Malpighian tubules, and fat body) of the oriental fruit fly, <it>Bactrocera dorsalis </it>(Hendel).</p> <p>Results</p> <p>Two different programs, <it>geNorm </it>and <it>Normfinder</it>, were used to analyze the data. According to <it>geNorm</it>, α-TUB + ACT5 are the most appropriate reference genes for gene expression profiling across the three different tissues in the female flies, while ACT3 + α-TUB are considered as the best for males. Furthermore, we evaluated the stability of the candidate reference genes to determine the sexual differences in the same tissue. In the midgut and Malpighian tubules, ACT2 + α-TUB are the best choice for both males and females. However, α-TUB + ACT1 are the best pair for fat body. Meanwhile, the results calculated by <it>Normfinder </it>are quite the same as the results with <it>geNorm</it>; α-TUB is always one of the most stable genes in each sample validated by the two programs.</p> <p>Conclusions</p> <p>In this study, we validated the suitable reference genes for gene expression profiling in different tissues of <it>B. dorsalis. </it>Moreover, appropriate reference genes were selected out for gene expression profiling of the same tissues taking the sexual differences into consideration. This work not only formed a solid basis for future gene expression study in <it>B. dorsalis</it>, but also will serve as a resource to screen reference genes for gene expression studies in any other insects.</p

Incidence of competitors and landscape structure as predictors of woodland-dependent birds

Globally, modification of landscapes for agriculture has had a strong influence on the distribution and abundance of biota. In particular, woodland-dependent birds are under threat across agricultural landscapes in Britain, North America and Australia, with their decline and extirpation attributed to the loss and fragmentation of habitat. Other native species have become over-abundant in response to anthropogenic landscape change and have strong interactive effects on avian assemblage structure. In eastern Australia, the hyper-aggressive noisy miner (Manorina melanocephala) often dominates woodlands in agricultural landscapes through interspecific competition, resulting in declines of species richness of woodland-dependent birds. We aimed to determine the relative influence and importance of interspecific competition, in situ habitat structure and landscape structure for woodland-dependent bird species at the landscape level. We recorded species-specific landscape incidence of woodland-dependent birds in 24 agricultural-woodland mosaics (25 km(2)) in southern Queensland, Australia. We selected extensively cleared landscapes (10-23 % woodland cover) where fragmentation effects are expected to be greatest. We applied generalised linear models and hierarchical partitioning to quantify the relative importance of the landscape-level incidence of the noisy miner, mistletoe abundance, shrub cover, woodland extent, woodland subdivision and land-use intensity for the incidence of 46 species of woodland birds at the landscape-scale. The landscape-level incidence of the noisy miner was the most important explanatory variable across the assemblage. Both in situ habitat structure and landscape structure were of secondary importance to interspecific aggression, although previous research suggests that the increasing incidence of the noisy miner in fragmented agricultural landscapes is itself a consequence of anthropogenic changes to landscape structure. Species' responses to fragmentation varied from positive to negative, but complex habitat structure had a consistently positive effect, suggesting in situ restoration of degraded habitats could be a conservation priority. Landscape wide conservation of woodland-dependent bird populations in agricultural landscapes may be more effective if direct management of noisy miner populations is employed, given the strong negative influence of this species on the incidence of woodland-dependent birds among landscapes