A common physiological response of organisms to environmental stresses is the increase in expression of heat shock proteins (Hsps). In insects, this process has been widely examined for heat stress, but the response to cold stress has been far less studied. In the present study, we focused on 11 Drosophila melanogaster Hsp genes during the stress exposure and recovery phases. The temporal gene expression of adults was analyzed during 9 h of cold stress at 0 °C and during 8 h of recovery at 25 °C. Increased expression of some, but not all, Hsp genes was elicited in response to cold stress. The transcriptional activity of Hsp genes was not modulated during the cold stress, and peaks of expression occurred during the recovery phase. On the basis of their response, we consider that Hsp60, Hsp67Ba and Hsc70‐1 are not cold‐inducible, whereas Hsp22, Hsp23, Hsp26, Hsp27, Hsp40, Hsp68, Hsp70Aa and Hsp83 are induced by cold. This study suggests the importance of the recovery phase for repairing chilling injuries, and highlights the need to further investigate the contributions of specific Hsp genes to thermal stress responses. Parallels are drawn between the stress response networks resulting from heat and cold stress.

The maternally inherited intracellular bacteria Wolbachia can manipulate host reproduction in various ways that foster frequency increases within and among host populations. Manipulations involving cytoplasmic incompatibility (CI), where matings between infected males and uninfected females produce non-viable embryos, are common in arthropods and produce a reproductive advantage for infected females. CI was associated with the spread of Wolbachia variant wRi in Californian populations of Drosophila simulans, which was interpreted as a bistable wave, in which local infection frequencies tend to increase only once the infection becomes sufficiently common to offset imperfect maternal transmission and infection costs. However, maternally inherited Wolbachia are expected to evolve towards mutualism, and they are known to increase host fitness by protecting against infectious microbes or increasing fecundity. We describe the sequential spread over approximately 20 years in natural populations of D. simulans on the east coast of Australia of two Wolbachia variants (wAu and wRi), only one of which causes significant CI, with wRi displacing wAu since 2004. Wolbachia and mtDNA frequency data and analyses suggest that these dynamics, as well as the earlier spread in California, are best understood as Fisherian waves of favourable variants, in which local spread tends to occur from arbitrarily low frequencies. We discuss implications for Wolbachia-host dynamics and coevolution and for applications of Wolbachia to disease control. Wolbachia are bacteria that live within the cells of arthropod hosts and are widespread in many groups of insects. These bacteria can rapidly spread through a population through a process of cytoplasmic incompatibility whereby females uninfected by Wolbachia show embryo death when they mate with males carrying the bacteria. Because the infected females pass on Wolbachia to their offspring, this places them at a reproductive advantage, ensuring that the infection spreads through insect populations once it reaches a high enough frequency to overcome any negative fitness effects on its host. Yet while such a rapid spread has been predicted, it has rarely been observed in nature. Here we show that a Wolbachia infection of Drosophila simulans flies has spread very rapidly in eastern Australia, replacing another Wolbachia infection that has also spread in recent years. These invasions appear to have taken place from a very low frequency, implying that both infections are likely to have had a benefit to their hosts rather than a cost. These results have implications for the spread of Wolbachia infections currently being introduced into populations of mosquitoes and other insects for disease suppression.

The use of conventional chemical insecticides and bacterial toxins to control lepidopteran pests of global agriculture has imposed significant selection pressure leading to the rapid evolution of insecticide resistance. Transgenic crops (e.g., cotton) expressing the Bt Cry toxins are now used world wide to control these pests, including the highly polyphagous and invasive cotton bollworm Helicoverpa armigera. Since 2004, the Cry2Ab toxin has become widely used for controlling H. armigera, often used in combination with Cry1Ac to delay resistance evolution. Isolation of H. armigera and H. punctigera individuals heterozygous for Cry2Ab resistance in 2002 and 2004, respectively, allowed aspects of Cry2Ab resistance (level, fitness costs, genetic dominance, complementation tests) to be characterised in both species. However, the gene identity and genetic changes conferring this resistance were unknown, as was the detailed Cry2Ab mode of action. No cross-resistance to Cry1Ac was observed in mutant lines. Biphasic linkage analysis of a Cry2Ab-resistant H. armigera family followed by exon-primed intron-crossing (EPIC) marker mapping and candidate gene sequencing identified three independent resistance-associated INDEL mutations in an ATP-Binding Cassette (ABC) transporter gene we named HaABCA2. A deletion mutation was also identified in the H. punctigera homolog from the resistant line. All mutations truncate the ABCA2 protein. Isolation of further Cry2Ab resistance alleles in the same gene from field H. armigera populations indicates unequal resistance allele frequencies and the potential for Bt resistance evolution. Identification of the gene involved in resistance as an ABC transporter of the A subfamily adds to the body of evidence on the crucial role this gene family plays in the mode of action of the Bt Cry toxins. The structural differences between the ABCA2, and that of the C subfamily required for Cry1Ac toxicity, indicate differences in the detailed mode-of-action of the two Bt Cry toxins. Transgenic crops expressing the insecticidal protein Cry2Ab from Bacillus thuringiensis (Bt) are used worldwide to suppress damage by lepidopteran pests, often used in combination with Cry1Ac toxin to delay resistance evolution. Until now, the Cry2Ab mode of action and the mechanism of resistance were unknown, with field-isolated Cry2Ab resistant Helicoverpa armigera showing no cross-resistance to Cry1Ac. In this study, biphasic linkage analysis of a Cry2Ab-resistant H. armigera family followed by EPIC marker mapping and candidate gene sequencing identified three independent INDEL mutations in an ATP-Binding Cassette transporter subfamily A gene (ABCA2). A deletion mutation was identified in the same gene of resistant H. punctigera. All four mutations are predicted to truncate the ABCA2 protein. This is the first molecular genetic characterization of insect resistance to the Cry2Ab toxin, and detection of diverse Cry2Ab resistance alleles will contribute to understanding the micro-evolutionary processes that underpinned lepidopteran Bt-resistance.

We have developed and validated two new fluorescence-based PCR assays to detect the Wolbachia w Mel strain in Aedes aegypti and the w Ri and w Au strains in Drosophila simulans . The new assays are accurate, informative, and cost-efficient for large-scale Wolbachia screening.