Insect molecular biology: an Australian perspective



One of the biggest advances in biological research has undoubtedly been the development of our capacity to investigate individual phenotypes, species biology and multi-species interactions at the molecular level. This has provided the ability to understand the detailed mechanisms that regulate biological processes and, in many cases, to manipulate them or use them to our advantage. In this Overview we define ‘insect molecular biology’ as the study of gene/protein expression and molecular function and contrast it with ‘traditional entomology’ and ‘comparative molecular entomology’. Obtaining the genomes of various insect species has provided significant advances in our ability to quickly isolate important genes. Study of the proteins they produce is important as they are functionally extremely diverse and are the basis for biological differences in extant species. Australian researchers have contributed significantly to our knowledge of insect molecular biology. Functional insect molecular biology studies undertaken in Australia are summarised, concentrating on the last 15 years, during which time insect molecular research has accelerated, largely due to obtaining key insect genomes and corresponding advances in molecular technologies. Currently, however, in Australia there is minimal collaboration between insect molecular biologists and researchers working in traditional or comparative molecular biology. We propose that an increase in these types of collaborations would benefit the broad field of entomology in Australia and increase the impact of Australian entomology globally.


At a practical level, entomological research can be classified into three (somewhat arbitrary and overlapping) categories. First, ‘traditional entomology’, which generally deals with morphological taxonomy, ecology and pest management/biocontrol. This area of entomology often defines deeper areas of research by identifying interesting biological phenomena. This research may use established biochemical techniques (such as enzyme activity assays) but does not investigate DNA or protein. Second, ‘comparative molecular entomology’, which uses comparisons of one or several molecules (or parts thereof) to conduct research into genetics, comparative genomics, systematics/phylogenetics, population studies, diagnostics/barcoding and monitoring. This type of research is often led by established ‘traditional’ entomologists, who develop the capacity to perform routine molecular techniques such as standard polymerase chain reaction (PCR), DNA sequencing and allozyme analysis. Comparative molecular entomology produces diagnostic data relating to the detection and quantification of biological phenomena, but generally does not deal with the mechanisms underlying these processes. The third category, ‘insect molecular biology’, studies molecular interactions that regulate biological processes. In simple terms, this involves the production of proteins (and non-coding RNAs) from DNA, and the function of these proteins/RNAs. This is fundamental research as it focuses on functional molecular interactions, which provides information on the mechanism and regulation of biological processes, rather than simply their detection/measurement. Scientists with a background in molecular biology generally lead this type of research, rather than those trained in traditional entomology. In this Overview we will concentrate on Australian research falling into what we have defined as ‘insect molecular biology’, as these studies are not often published in the Australian Journal of Entomology. In contrast, there has been a range of papers in this journal that deal with the ‘traditional’ and ‘comparative molecular’ research. For example, DNA barcoding was discussed in a previous Overview article (Mitchell 2008).


The importance of molecular biology is that its major focus is how molecules function to regulate higher-order processes such as development, immunity, olfaction, symbiosis, etc. A key point to consider is that DNA is not a functional molecule in itself but acts as a template from which functional and structural proteins are derived, and provides binding sites for regulatory proteins involved in DNA replication, DNA repair and gene expression. At a functional level, an organism's protein repertoire differentiates it from another, rather than its DNA. In that sense, the term ‘gene function’ is misleading. Whereas DNA is one molecule with a set of biological/physical properties, the proteins it encodes are extremely diverse (of an order similar to the expressed genes) and determine an organism's structure and metabolism. Additionally, similarities/differences in ‘non-coding’ DNA (which are generally not subject to heavy selection pressure) are indicative of ancestral commonality and relative timing of subsequent divergence. They are not, however, necessarily indicative of the current biological differences between extant species.

A good example of this has been the long debated relationship between cockroaches and termites. A recent phylogenetic study applied Bayesian analysis to DNA sequence data from five highly conserved genetic loci with results suggesting that the cockroach family Blattidae is more closely related to the Termitidae than to cockroach families of the Blaberoidea (Inward et al. 2007). The obvious morphological and biological differences between cockroaches and termites are not obviously reflected in genomic DNA of the different taxa but were likely driven by changes to the repertoire of functional genes within their commensal gut flora. Within the lower termites this is further evidenced by the existence of taxon-specific microorganism communities (Yang et al. 2006), within which each species has the potential to provide complete metabolic pathways that benefit a host. In the case of termites, these pathways are crucial to wood degradation. Interestingly, the discovery of shared symbiotic gut flagellates between wood-eating Cryptocercus cockroaches and primitive termites originally raised the possibility that the two groups shared a relatively recent ancestor (Cleveland et al. 1934; Inward et al. 2007). The point of this example is to illustrate the importance of understanding protein function, in terms of developing a detailed understanding of HOW species evolve and WHY extant taxa show functional biological differences.

The first genome sequence for an insect, that of the genetic model, the vinegar fly Drosophila melanogaster Meigen 1830, was published in the year 2000 (Adams et al. 2000). This was a breakthrough in insect molecular biology as it facilitated the identification of large numbers of functional genes (and gene families) for isolation and analysis, both from Drosophila and from other insects. In addition, the fact that Drosophila shares a range of genes that are apparently orthologous to human genes has seen this species become a model for both insect and human molecular biology (as it has been for genetics) (e.g. see Sanokawa-Akakura et al. 2010). Some examples of these shared genes include those encoding neurological signalling proteins (Hirth 2010), developmental proteins (Coulson et al. 1998; Hayward et al. 2002; Wilanowski et al. 2002) and DNA damage repair proteins (Sun et al. 2010). The level of genetic characterisation of Drosophila, and the availability of powerful molecular tools to manipulate it, has seen it used widely to study a variety of genes and proteins from diverse species. Australian scientists have produced various tools that can be used to perform genetic and developmental analyses on D. melanogaster (Lockett et al. 1992; Clarkson & Saint 1999; Saint & Clarkson 2000; Murray & Saint 2007; O'Keefe et al. 2007).

Since the sequencing of D. melanogaster, large portions of the genomes of 31 other insect species have been obtained. These are mainly diptera (including various Drosophila spp. and three mosquito species), but also include pest species from other orders such as the head/body louse (Pediculus humanus (De Geer 1767)), pea aphid (Acyrthosiphon pisum (Harris 1776)), triatomid bug Rhodnius prolixus Stal 1859 and red flour beetle Tribolium castaneum (Herbst 1797), as well as three species that are considered beneficial, namely the silkworm moth (Bombyx mori, Linnaeus 1758), the honeybee (Apis mellifera Linnaeus 1758) and the jewel wasp Nasonia vitripennis (Walker 1836) (Tagu et al. 2010). Australian researchers have been involved in the production and analyses of several of these genomes, including those of A. mellifera (Honeybee Genome Sequencing Consortium 2006), Drosophila spp. (Drosophila 12 Genomes Consortium 2007), A. pisum (International Aphid Genomics Consortium 2010; Tagu et al. 2010) and N. vitripennis (Nasonia Genome Working Group 2010). In terms of global monetary input, the genes/proteins of mosquitoes are the most highly studied, particularly with respect to those regulating disease-vectoring ability. Vast sums are being spent worldwide to understand the molecular basis for interactions between mosquitoes and disease-related microorganisms, mosquito host-finding and mosquito immunity/development (e.g. Ramirez & Dimopoulos 2010). Australian researchers have attracted substantial funds to conduct research into microorganism-based mosquito control (Moreira et al. 2009).


Here we review the insect molecular biology research involving Australian groups, primarily over the previous 15 years, a time that has seen genomics emerge and accelerate molecular research. The sequencing of genomes of species of medical and agricultural importance, illustrates how insect molecular biology is often application-driven, with researchers spread across fields such as medicine, developmental biology, biosecurity and biotechnology, rather than within more traditional entomological fields such as taxonomy, phylogenetics, ecology and pest management/biological control. Furthermore, the funding bodies that service molecular biology and traditional areas of entomological research are often different. In an Australian context, we would expect molecular biology research to receive a much higher proportion of funding through industry-based groups such as industry research and development corporations (RDCs; such as Grains, Cotton, Grape and Wine RDCs), multi-industry levy-based funders (e.g. Horticulture Australia Ltd), cooperative research centres (CRCs; such as National Plant Biosecurity CRC), the Australian Research Council, and medical funders such as National Health and Medical Research Council. In contrast, traditional entomology is funded more through heritage-, conservation- or environment-based grants such as the Australian Biological Resources Study, with the exception of traditional pest management/biological control research, which is largely industry funded.

Mining insect proteins with potential commercial application

Molecular biology research is largely characterised by investigations into protein expression and function. For example, CSIRO has a research program investigating the structure and function of invertebrate proteins, with a focus on those with commercial applications such as silk proteins. One of the most successful examples of this type of research is the characterisation, modification and purification of resilin protein, which has structural similarities with silk proteins and functions to aid insects in fast, repetitive movements involving recoil, such as flying or jumping (Elvin et al. 2005). The resilin project was initially collaboration between CSIRO, the Australian National University (ANU) and the University of Queensland (UQ). The research involved the isolation of Drosophila pro-resilin-like gene (CG15920 gene) from which only the first exon was expressed as a soluble protein, using recombinant bacteria. A fungal enzyme was then used to catalyse a UV-mediated cross linking of the soluble pro-resilin into a solid rubber-like material, but with much greater resilience in maintaining strength and flexibility under repeated use than even high-performance rubbers. Prior expression analysis of the gene had shown that expression was restricted to the pupal phase in Drosophila, indicating that its resilience must be such that it can be maintained throughout the adult life of the insect. The inert recombinant pro-resilin is now being developed for medical applications such as repair or replacement of joints, and joining of severed tissues or sutures.

Another highly studied protein important for hormonal regulation of insect development is the ecdysone receptor (EcR). These receptors are important as they are potential targets for insecticides. CSIRO researchers initially characterised LcEcR1 from Lucilia cuprina and showed it was a functional steroid hormone receptor that could produce ecdysteroid-dependent transcription when expressed in mammalian cells (Hannan & Hill 1997). In addition, the group characterised an ultraspiracle gene product from L. cuprina, which in Drosophila was known to form heterodimers with EcRs to produce a functional signalling complex; this function was confirmed for the L. cuprina gene, also using recombinant mammal cells (Hannan & Hill 2001). Subsequently, the X-ray crystal structure of the ligand-binding domain of EcR from the whitefly Bemisia tabaci (Gennadius 1889) in complex with an ecdysone analogue was obtained and compared with that of the moth, Heliothis virescens Fabricius 1777 (Carmichael et al. 2005).

These studies provided an insight into the apparent insensitivity of hemipterans (relative to lepidopterans and dipterans) to bis-acyl-hydrazine insecticides, which interact with the EcR ligand-binding domain. This work provided the basis for further research that allowed purification, characterisation (including ligand-binding kinetics) and comparison of recombinant ligand-binding domains of the EcRs from the four pest species L. cuprina, Myzus persicae (Sulzer 1776), B. tabaci and Helicoverpa armigera (Hübner 1805) (Graham et al. 2007a,b). This research, in turn, led to pesticide screening assays to detect other compounds that interact with the ligand-binding domains of EcRs, which were patented. These assays facilitated discovery of γ-methylene γ-lactans that showed high affinity for the target site in the sheep body louse Bovicola ovis (Schrank 1781) and L. cuprina (Birru et al. 2010). Although insect esterase proteins have often been studied with regard to insecticide resistance (see below), a different type of esterase, juvenile hormone esterase (responsible for hydrolysis of juvenile hormone), from Drosophila and the cricket Gryllus assimilis Fabricius 1775, has also been isolated (Campbell et al. 1998b, 2001; Crone et al. 2007).

Insect olfaction

Researchers at Monash University are using Drosophila as a model to investigate molecular mechanisms of olfaction, development and disease processes. For example, they use single-sensillum recordings, combined with molecular biology techniques, to examine the way in which insects detect volatile compounds within the environment (Dobritsa et al. 2003; de Bruyne & Warr 2005; Kiely et al. 2007). A recent study showed that Drosophila olfactory receptors (ORs) could detect a range of compounds associated with toxic gasses, explosives and illicit drugs, and that they also had the capacity to detect synthetic compounds (Marshall et al. 2010). The group has also investigated the functional evolution of cells and ORs related to detection of volatile esters by Drosophila spp. (de Bruyne et al. 2010), and in collaboration with UQ and several international scientists, also published work that investigated the currently unresolved signalling mechanisms driving insect olfaction (Smart et al. 2008).

Groups at CSIRO and the South Australian Research and Development Institute (SARDI) are also conducting research into insect ORs particularly with respect to identification and characterisation of important ORs (Anderson et al. 2009; Jordan et al. 2009) and their use as biological detection molecules in olfactory biosensors (or bioelectronic noses) (Leifert et al. 2009; Bailey 2010; Glatz & Bailey-Hill 2011). Such biosensors (of which no commercial examples currently exist) could expect wide use, with countless potential applications in medicine, pest management, agriculture, security and environmental monitoring. Gustatory (taste) receptors in Drosophila have also been studied in Australia (Chyb 2007).

Drosophila development and homeostasis

Australian scientists have published a number of papers on Drosophila development in high-impact journals, particularly with regard to expression and function of proteins associated with cell-cycle regulation during embryonic development (Saint & Patterson 1993), and development of the eye (Brumby et al. 2004). This research began at CSIRO in the late 1980s with a paper in Nature postulating a specific function for the Drosophila rough gene in playing a role in eye pattern formation (Saint et al. 1988), and has endured until the present. During the 1990s, the key researchers were located at the University of Adelaide (UoA) and Melbourne University, with the work being then largely being driven from ANU in the first decade of this century, in collaboration such as with CSIRO and the Peter MacCallum Cancer Centre. Several international groups have also been involved.

The involvement of rough in eye development was further investigated with regard to chromosomal location and relevant adjacent genes (Knibb et al. 1993), and the basis for its regulatory specificity (Lockett et al. 1993). Another important genetic locus involved in Drosophila eye (and other sensory) development is lozenge; Australian scientists conducted a range of mutational studies of lozenge and linked various eye defect phenotypes (Batterham et al. 1996a; Crew et al. 1997; Siddall et al. 2003) and antennal defect phenotypes (Stocker et al. 1993) to mutations within specific regions. Several interacting genes have also been investigated, including D-Pax2 (thought to play a role in eye cell differentiation; Siddall et al. 2003) and Yan (thought to regulate lozenge expression during eye development; Behan et al. 2002).

Cyclin proteins (which also occur in mammals and regulate progression of the cell cycle between various phases) have been studied for their role in Drosophila development, including eye development. Expression studies showed that cyclin E was active in the G1 phase of neural cells (Richardson et al. 1993) and could induce premature entry of eye imaginal disc cells and embryonic epidermal cells, into S phase when expressed ectopically in these cells (Knoblich et al. 1994; Richardson et al. 1995). Cyclin E transcription was shown to be regulated through a series of tissue- and developmental stage-specific transcription factors (Jones et al. 2000). Mutational studies also implicated cyclin E in regulation of cell proliferation in eye imaginal discs (producing a ‘rough eye’ phenotype), and provided tools to screen for interacting proteins; this research confirmed that several of these proteins could modify the mutated cyclin E-derived phenotype (Secombe et al. 1998; Brumby et al. 2002, 2004).

Furthermore, developmental function of two forms of cyclin E were investigated, leading to the discovery that only one of the proteins (cyclin EII) was active in regulating cell phase transitions within the morphogenetic furrow of the imaginal disc, due to inhibition of the cyclin EI in that specific region (Crack et al. 2002). Another study identified other cell-cycle regulatory proteins important for G2 transitions; string is a phosphatase shown to function in eye disc cells but not abdominal histoblasts, and stg, which functions in wing imaginal discs (Kylsten & Saint 1997). String transcription was subsequently shown to be regulated in a cell- and tissue-specific manner by a range of cis-acting elements (Lehman et al. 1999).

Proteins that regulate mitosis and cell division (cytokinesis) during embryogenesis have also been a major field of research (see reviews by Prokopenko et al. 2000b; O'Keefe et al. 2001; Saint & Somers 2003). This research has focused largely on characterisation of the GTP exchange factor pebble (Hime & Saint 1992) and the associated Rho family of small GTPase activators (particularly RacGAP50C), which together are associated with the formation of the contractile ring that mediates the physical cell division (Prokopenko et al. 1999; Prokopenko et al. 2000a). Subsequent research indicated that the pebble gene product is involved in regulation of actomyosin organisation, and that RacGAP50c (in combination with various effectors) regulates bundling of microtubules by interacting directly with anillin (which is a cytoskeleton-related protein found in the contractile ring) (Somers & Saint 2003; Zavortink et al. 2005; Gregory et al. 2008). Pebble-related expression was also linked to the normal developmental process known as the epithelial-mesenchymal transition that occurs during gastrulation (Smallhorn et al. 2004).

Other Drosophila cell-cycle regulators characterised in Australia include the three rows gene required for chromosomal alterations during mitosis (D'Andrea et al. 1993), citron kinase whose correct localisation to the contractile ring is mediated through pebble-mediated signalling (Shandala et al. 2004) and more recently, the Tum and Pav proteins, which are associated with cytokinesis but also with regulated cell death (Jones et al. 2010). In addition, Australian scientists developed a screening system for modifiers of Rho-mediated signalling relating to cytokinesis; the system involved overexpression of putative gene candidates in vivo and the correlation of effects on a defined eye phenotype caused by defective signalling (Gregory et al. 1997). This system allowed identification of four candidate genes not previously implicated in cytokinesis in Drosophila, although orthologues of several of the genes had been implicated in other organisms.

Other Drosophila developmental proteins have been studied in Australia. Research at UoA and ANU identified a novel sequence-specific DNA-binding protein (encoded by the dead ringer gene), which was shown to be involved in transcriptional regulation of proteins affecting embryonic developmental processes including central nervous system development, anterior–posterior patterning and muscle development (Gregory et al. 1996; Shandala et al. 1999, 2002, 2003). Some of the genes regulated by dead ringer were also identified (Shandala et al. 1999) and dead ringer itself was shown to be a member of a broader family of ARID (AT-rich interacting domain) proteins, found in fungi and all metazoans (Kortschak et al. 2000). The Polycomb group (PcG) genes of D. melanogaster (e.g. polycomblike gene) were also investigated and shown to regulate important developmental gene expression, through interaction with other PcG proteins (Lonie et al. 1994; O'Connell et al. 2001). Mechanisms involved in establishment of the dorsoventral axis in fly embryos have also been studied, including the interplay (including receptor tyrosine kinase signalling) between the transcriptional modulator Dorsal, a corepressor required for downregulated gene expression (Groucho), and an accessory ‘silencer’ locus that contained a dead ringer gene (Valentine et al. 1998; Hader et al. 2000). A further high impact publication reported the characterisation of a development-related Drosophila histone protein, His2AvD, in relation to important functional regions of the protein (Clarkson et al. 1999).

Australian scientists also have published articles characterising proteins associated with Drosophila copper homeostasis (Southon et al. 2004, 2008, 2010), including tissue-specific effects (Burke et al. 2008; Binks et al. 2010) and the role in Drosophila development (Norgate et al. 2006). Other developmental signalling pathways have also been investigated (e.g. Burke et al. 1999; Behan et al. 2005; Milton et al. 2005; Siddall et al. 2009). A protein involved in a different type of homeostasis in Drosophila (maintenance of muscle), the cochaperone Starvin, was characterised with respect to coordination of smaller proteins that degrade damaged protein (Amdt et al. 2010). Further, Drosophila ion channels have also been characterised in Australia (Warr & Kelly 1996), and a specific type of ribonuclease gene has been isolated (Hime et al. 1995).

Adaptation to stress and toxins

The Centre for Environmental Stress and Adaptation Research (CESAR, at the University of Melbourne) has used Drosophila as a model to investigate the basis for adaptation to stresses such as environmental changes (Hoffmann & Hercus 2000; Hoffmann & Willi 2008), particularly temperature (Hoffmann et al. 2003), but also starvation (Harshman et al. 1999) and infection with the intracellular bacterial parasite, Wolbachia (Carrington et al. 2009). Much of this work sits largely within what we have defined here as comparative molecular entomology as it has generally used a combination of traditional quantitative genetics to correlate putative stress-related genetic polymorphisms (that may occur in a natural cline) with adaptive physiological and morphological phenotypes (Hoffmann et al. 2004, 2005; Umina et al. 2005; Hoffmann & Weeks 2007; Sgro et al. 2008). A key challenge of this research is determining the functional basis for adaptive changes as many of the genetic polymorphisms used are either markers to the relevant functional genes or are not necessarily associated with functional proteins (Frydenberg et al. 2003). Indeed, when putative functional gene loci associated with thermal adaptation of Drosophila in Europe were examined for variation along an established Australian cline, none was detected (Weeks et al. 2006). In order to unravel the functional basis for these adaptive changes, molecular biology techniques have been used to examine correlations with gene expression levels (Johnson et al. 2009; Telonis-Scott et al. 2009; Jensen et al. 2010; McKechnie et al. 2010).

A recent paper used various genetic stains of Drosophila with respect to a locus encoding a heat-shock protein 90 (HSP90) variant to show that it was associated with ‘release’ of temperature-dependent phenotypic variability in some canalised (phenotypically rigid) traits (Sgro et al. 2010). Previous research indicated that a HSP90-related locus was involved in maintaining canalised morphological traits by buffering associated genotypic variation, although other processes were also involved (Milton et al. 2003). Expression of the heat shock gene hrs-omega had also been correlated with a Drosophila thermal phenotype (McKechnie et al. 1998). Another recent paper has reported that Drosophila ROQUIN family proteins (also found in humans and Caenorhabditis elegans Maupas 1900) are associated with cytoplasmic stress granules, which are formed when eukaryotic cells respond to environmental stress, including heat (Athanasopoulos et al. 2010). The research showed that the ROQUIN proteins bind to specific mRNAs in the cytoplasm to disrupt expression of the associated proteins. Interestingly, it is thought that these stress granules may also function by facilitating preferential expression of stress proteins such as heat-shock proteins. Adaptive proteins associated with aphid–plant interactions have also been briefly examined; the salivary transcriptome and proteome of A. pisum were analysed, leading to the isolation of a salivary gland protein (denoted C002) that is crucial in allowing the aphid to feed on fava bean (Mutti et al. 2008).

Genomic techniques have been extremely important in identifying putative resistance genes across a range of insects (Oakeshott et al. 2003; Teese et al. 2010). Detoxification and insecticide-resistance mechanisms in Drosophila have been the focus of several groups, such as CSIRO, University of Melbourne and CESAR, St. Vincent's Institute of Medical Research, Victorian Department of Primary Industries and ANU. This research investigated several key areas including the expression of resistance-related genes (Odgers et al. 2002; Pyke et al. 2004; Willoughby et al. 2006, 2007), and the expression and function of relevant Drosophila proteins. These proteins include glutathione S-transferases (GSTs) (Low et al. 2007, 2010), cytochrome P450s including that encoded by the Cyp6g1 locus (Bogwitz et al. 2005; Russell et al. 1996; Daborn et al. 2002, 2007; Chung et al. 2007, 2009; Schmidt et al. 2010a), a kinase (Chen et al. 2006), esterases (Campbell et al. 2003) and the acetylcholine receptor (Perry et al. 2007, 2008). These genes have also been a priority in terms of analysing the N. vitripennis genome (Oakeshott et al. 2010). Interestingly, expression of a Drosophila P450 enzyme (among other candidate genes) has been correlated with male aggression (Robin et al. 2006).

Gene targets for the Drosophila-based resistance work were partially informed by earlier studies of chemical resistance mechanisms in the Australian sheep blowfly, L. cuprina (Wiedemann 1830), which has been investigated by CESAR, ANU and CSIRO. A GST (Board et al. 1994) and esterases (Smyth et al. 1994; Newcomb et al. 1996, 1997b) were identified (and isolated) as putative resistance genes in L. cuprina. The esterase proteins were further analysed for mutations that confer resistance (Newcomb et al. 1997a; Campbell et al. 1998a; Chen et al. 2001), with one mutation found to be responsible for organophosphorus resistance in L. cuprina and the house fly Musca domestica L. 1758 (Claudianos et al. 1999). Interestingly, the developmental Scalloped wings gene in L. cuprina (homologous to the Drosophila Notch cell receptor) was shown to be expressed in embryonic and pupal flies, and it was postulated to suppress asymmetry-related adaptive phenotypes in flies exhibiting organophosphate resistance (Chen et al. 1998), mediated by the Rop-1 carboxylesterase (Batterham et al. 1996b). Some of the Drosophila and Lucilia esterases have also been modified and compared with wild-type proteins in their ability to hydrolyse pyrethroids and pyrethroid analogues (Heidari et al. 2004, 2005; Devonshire et al. 2007). L. cuprina resistance to the growth regulators diflubenzuron and cyromazine has also been investigated, with important chromosomal regions identified (Batterham et al. 2006).

Lepidopteran insecticide resistance has also been studied, especially in the most economically important pest of Australian agriculture, the cotton bollworm H. armigera. Similar to research on Drosophila resistance, Australian researchers have used genomic approaches to identify putative resistance genes of H. armigera (Gordon et al. 2007; Wee et al. 2007; Teese et al. 2010). They are also studying this species with respect to validating tools for genetic manipulation, so that gene function can be studied either by targeted gene expression or gene silencing (Collinge et al. 2007; Williams et al. 2007). In addition, subtractive amplification of cDNA library fragments has been used to identify putative fenvalerate-resistance genes in H. armigera (Wee et al. 2008).

Resistance mechanisms in other insects have been investigated at the molecular level. CSIRO and ANU conducted an analysis of changes to the acetylcholinesterase protein in mosquitoes and the green peach aphid, showing that various changes to amino acids with the enzyme's active site were responsible for varying levels of resistance against carbamates and organophosphates (Russell et al. 2004). Additionally, there have been further reports published from Australian research, relating to resistance mechanisms in other pest species including the Mediterranean flour moth Ephestia kuehniella Zeller 1879 (Rahman et al. 2007), T. castaneum (Campbell 2010) and the lesser grain borer Rhyzopertha dominica (Fabricius 1792) (Campbell 2008).

A new collaborative project involving SARDI, CSIRO and Macquarie University is using various molecular approaches, including laser-scanning cytometry, to investigate genes and proteins involved in the cellular response to irradiation in the Queensland fruit fly Bactrocera tryoni (Froggatt 1897). Comparisons with Drosophila and human genomes will be important in this project for developing irradiation dosimetry assays for fruit flies and potentially for humans, given that similar genes and metabolic pathways are thought to be involved in both organisms (O'Keefe et al. 2005).

Insect pathology and immunity

Molecular biology is also being widely applied to the closely aligned fields of insect pathology (including parasitoid–host interactions) and insect immunity. For example, the potential role of silk proteins in the lepidopteran immune response has been investigated (Korayem et al. 2007). The unusual symbiosis between some ichneumonid wasps and their genomically encoded polydnaviruses have also been studied at the Universities of Adelaide and Queensland (Glatz et al. 2004b). These viruses are injected into a host caterpillar during parasitisation and are known to disrupt the immune function and development of hosts, allowing the wasp larva to develop. Australian researchers used the braconid wasp, Cotesia rubecula (Marshall 1885) as a model to isolate several immune-suppressive and structural viral proteins that were partially characterised (Asgari et al. 1996, 2003b; Glatz et al. 2003, 2004a; Cooper et al. 2011). In addition, they have studied some of the venom proteins of this wasp (reviewed in Asgari 2006) and shown that the proteins had various effects on the host immune system (Asgari et al. 2003a; Zhang et al. 2004a, 2006) and the expression of polydnavirus genes in the host (Zhang et al. 2004b). Expressing the C. rubecula Vn50 venom protein in Drosophila showed a range of immune, developmental and reproductive effects on recombinant flies (Thomas et al. 2010). Another project is investigating the venom proteins of an important biological control agent of the diamondback moth (Plutella xylostella, Linnaeus 1777), the ichneumonid wasp Diadegma semiclausum (Hellén 1949).

In addition to these immune-suppressive proteins from wasps, the proteins involved in mounting the lepidopteran immune defence have also been studied in Australia (see review by Schmidt et al. 2010b). In order to investigate novel controls for lepidopteran pests, the H. virescens ascovirus (HvAV-3, a pathogen of economically important Noctuidae) has been sequenced by the Australian Genome Research Facility at UQ (Asgari et al. 2007). Subsequent to this, aspects of HvAV-3 molecular biology have been investigated, including the isolation and characterisation of proteins and non-coding RNAs (microRNAs) associated with virus replication and pathology (Hussain & Asgari 2008; Hussain et al. 2009, 2010; Smede et al. 2009).

Australian scientists are also conducting world-leading research into Wolbachia, and sequenced the genome of a species occurring as a pathogenic symbiont in Drosophila, wMel (Wu et al. 2004). This has provided scientists with their first detailed insights into the genes that regulate Wolbachia–host interactions (Brownlie & O'Neill 2005; Iturbe-Ormaetxe et al. 2005) and has allowed the targeted characterisation of potentially useful proteins (Kurz et al. 2008, 2009). Various groups based in Queensland and New South Wales have collaborated on investigations into the effect of Wolbachia on immune modulation and life-cycle disruption of infected insects, particularly with regard to the use of the parasite as a mosquito control agent (McMeniman et al. 2009; Moreira et al. 2009). They have also shown that the bacterium reduced mortality of virus-infected Drosophila by inhibiting viral replication (Hedges et al. 2008), a finding with implications for reducing the impact of arboviruses.

Jumper ant venom

The venom of the jumper ant Myrmecia pilosula F. Smith 1858 has also been partially characterised. The reason for this research was to explore the basis for human anaphylaxis with which this species has been associated (McGain & Winkel 2002; Brown et al. 2003). Initial experiments were conducted in the mid- to late 1990s and involved the electrophoretic separation of the native venom proteins (Donovan et al. 1995), expression and analysis of recombinant allergens (Donovan et al. 1996; Street et al. 1996) and analysis of cytotoxicity of the minor allergen, pilosulin 1 (King et al. 1998; Wu et al. 1998). Subsequent research subjected the venom to more advanced proteomic analysis (Davies et al. 2004; Wiese et al. 2006) and led to a more detailed understanding of the various allergens and their importance in anaphylaxis (Wiese et al. 2007). In collaboration with Japanese researchers, it was subsequently demonstrated that the novel peptide, pilosulin 5, caused dose-dependent release of histamine from rat peritoneal mast cells (Inagaki et al. 2008).

Honeybee research

The honeybee is a highly studied insect in Australia, both scientifically due to its eusociality (Honeybee Genome Sequencing Consortium 2006), and commercially due to being the key pollinator of our exotic crops plants. The Queensland Brain Institute (part of UQ) is a well-known research centre that has collaborated with various other Australian groups to produce a range of high-impact research papers attempting to characterise the neurological basis for how insects sense their environment using honeybees as a model. A mixture of behavioural and molecular studies has shed light on the neural processing associated with honeybee vision and chemosensation (e.g. Biswas et al. 2008, 2010). The ability to investigate and compare the protein repertoire of biological samples (proteomics) has been used for various entomological studies in Australia. The seminal fluid proteome of A. mellifera was examined by scientists from the ARC Centre of Excellence in Plant Energy Biology, located at University of Western Australia (Baer et al. 2009). The honeybee genome has been mined by Australian researchers for the presence of various important regulatory proteins; a good example is an analysis of detoxification enzymes published in 2006 (Claudianos et al. 2006). Some work has been conducted on honeybee embryonic development at ANU; this work used gene-silencing and transcription analyses to implicate a chemosensory protein in regulation of integument development in bee embryos (Maleszka et al. 2007).


Although the above discussion does not cover all of the insect molecular biology that has been conducted in Australia, it does cover most of the major projects and provides an insight into the key research groups. In addition, we have made efforts to cite a significant percentage of the recent Australian insect molecular biology publications so that this article acts as a portal to much of the Australian science and scientists in this field, as we have defined it in the introduction. We have not discussed in detail the ‘comparative molecular’ studies conducted in Australia (which are numerous) and as such, there are few references for such articles in this Overview.

Given the speed at which molecular technologies are advancing and the speed/ease with which transcriptome, proteome and genome data can now be obtained, it is likely we will see several global trends that will be reflected in Australia. For example, we would expect to see an decrease in the cost of relevant molecular technologies, a greater number of students trained in molecular biology and an increase in the value of insect molecular biology research undertaken (particularly by industry funding bodies).

Despite the significant amount of entomology undertaken in Australia, there are very few projects involving collaboration between molecular biologists and traditional/comparative molecular entomologists. This is likely due to key differences that we highlighted earlier between these areas of research, namely the background of the researchers involved, researcher focus and funding sources. Indeed, insect molecular biologists in Australia tend to present their work at conferences such as the annual Insect Molecular Biology Conference and/or the meeting of the Australian Society for Chemosensory Science, whereas those working in traditional and comparative molecular entomology are more likely to attend the meetings of the Australian Entomological Society or ecology/systematics-based conferences. There are very few researchers involved in both of these groups and there is limited collaboration between the groups.

Hopefully, in the future we will see much greater collaboration between molecular biology researchers and those researchers in the other areas of entomology. Such a trend would be beneficial to each area of entomological research in Australia, and Australian entomology as a whole, as neither of the fields stands alone and each can play a role in informing the others. Such collaborative projects that combine functional molecular and ecological/phylogenetic data would also increase the competitiveness of Australia on the global stage (particularly with respect to the USA where such projects are far more common) and lead to improvements in the global relevance, research outcomes and impact, of Australian entomology.


We thank Dr Sassan Asgari (UQ) for critical review of this manuscript.