Low-dose methyl bromide fumigation as a quarantine disinfestation treatment for nectarines against Queensland fruit fly (Diptera:Tephritidae)

White nectarines (Prunus persica var. nucipersica) were fumigated with methyl bromide (MB) at a nominal treatment dose of 18 g m-3 at 18°C for 5 h and 30 min as a quarantine disinfestation treatment against Bactrocera tryoni, the Queensland fruit fly. Three large scale trials were conducted against each of the four immature lifestages, eggs and first, second and third instars. There were no survivors from the estimated 43,614 eggs, 41,873 first instars, 41,345 second instars and 33,549 third instars treated, thereby resulting in an efficacy of GROTERDAN99.99% mortality at the 95% confidence level for each lifestage. Of the 12 trials reported herein, the highest concentration of MB, sampled from the chamber headspace analysed by gas chromatography, was 18.7 g m-3. The maximum chamber temperature from 5 min readings was 19.7°C and the maximum fruit core temperature was 19.5°C. The treatment time for all trials was exactly 5.5 h. Thus the recommended treatment dose to disinfest nectarines from B. tryoni is 19.0 g m-3 MB at 20.0°C for 5.5 h. Fruit quality trials were conducted on white nectarines at three combinations of treatment parameters: 15 g m-3 MB at 19°C for 5.25 h; 18 g m-3 MB at 19°C for 5.5 h and 21 g m-3 MB at 19°C for 5.5 h. The fruit were stored at 0, 4 and 8 days at 4°C and 8 days at 4°C followed by 4 d at 22°C. They were then were assessed for skin colour, flesh colour, skin defects, flesh defects, fruit weight loss, flesh firmness, total soluble solids, titratable acidity and rots. There was no significant difference between untreated control and MB treated fruits in any of the parameters measured. Thus the treatments did not have adverse effects on fruit quality.

Area-wide releases and evaluation of the parasitoid Eretmocerus hayati(Hymenoptera:Aphelinidae) for silverleaf whitefly control

Silverleaf whitefly (SLW), Bemisia tabaci biotype B, is a major horticultural pest that costs Queensland vegetable growers millions of dollars in lost production and control measures each year. In the Bowen and Burdekin districts of North Queensland, the major cultivated SLW host crops are tomatoes, melons, green beans, pumpkins, eggplants, and cucumbers, which cover a total production area of approximately 6500 ha. Eretmocerus hayati, an effective SLW parasitoid, was imported into Australia by CSIRO in 2002 and released from quarantine in 2004. In 2006, DAFF established a mass-rearing unit for E. hayati at Bowen Research Station to provide E. hayati for release on vegetable farms within its SLW integrated pest management research program. A total of 1.3 million E. hayati were released over three seasons on 34 vegetable farms in the Bowen and Burdekin districts (October 2006 to December 2008). Post-release samplings were conducted across the release area over this time period with parasitism levels recorded in tomatoes, melons, beans, eggplants, pumpkins, and various SLW weed hosts. Sample data show that E. hayati established at most release sites as well as some non-release sites, indicating natural spread. Overall results from these three years of evaluation clearly demonstrated that E hayati releases played a significant role in SLW control. In most crops sampled, E hayati exerted between 30 and 80% parasitism. Even in regularly sprayed crops, such as tomato and eggplant, E. hayati was able to achieve an overall average parasitism of 45%.

Open source computer-vision based guidance system for UAVs on-board decision making

The use of UAVs for remote sensing tasks; e.g. agriculture, search and rescue is increasing. The ability for UAVs to autonomously find a target and perform on-board decision making, such as descending to a new altitude or landing next to a target is a desired capability. Computer-vision functionality allows the Unmanned Aerial Vehicle (UAV) to follow a designated flight plan, detect an object of interest, and change its planned path. In this paper we describe a low cost and an open source system where all image processing is achieved on-board the UAV using a Raspberry Pi 2 microprocessor interfaced with a camera. The Raspberry Pi and the autopilot are physically connected through serial and communicate via MAVProxy. The Raspberry Pi continuously monitors the flight path in real time through USB camera module. The algorithm checks whether the target is captured or not. If the target is detected, the position of the object in frame is represented in Cartesian coordinates and converted into estimate GPS coordinates. In parallel, the autopilot receives the target location approximate GPS and makes a decision to guide the UAV to a new location. This system also has potential uses in the field of Precision Agriculture, plant pest detection and disease outbreaks which cause detrimental financial damage to crop yields if not detected early on. Results show the algorithm is accurate to detect 99% of object of interest and the UAV is capable of navigation and doing on-board decision making.

Causes and consequences of inbreeding in the ant Formica exsecta

Human actions cause destruction and fragmentation of natural habitats, predisposing populations to loss of genetic diversity and inbreeding, which may further decrease their fitness and survival. Understanding these processes is a main concern in conservation genetics. Yet data from natural populations is scarce, particularly on invertebrates, owing to difficulties in measuring both fitness and inbreeding in the wild. Ants are social insects, and a prime example of an ecologically important group for which the effects of inbreeding remain largely unstudied. Social insects serve key roles in all terrestrial ecosystems, and the division of labor between the females in the colonies queens reproduce, workers tend to the developing brood probably is central to their ecological success. Sociality also has important implications for the effects of inbreeding. Despite their relative abundance, the effective population sizes of social insects tend to be small, owing to the low numbers of reproductive individuals relative to the numbers of sterile workers. This may subject social insects to loss of genetic diversity and subsequent inbreeding depression. Moreover, both the workers and queens can be inbred, with different and possibly multiplicative consequences. The aim of this study was to investigate causes and consequences of inbreeding in a natural population of ants. I used a combination of long-term field and genetic data from colonies of the narrow-headed ant Formica exsecta to examine dispersal, mating behavior and the occurrence of inbreeding, and its consequences on individual and colony traits. Mating in this species takes place in nuptial flights that have been assumed to be population-wide and panmictic. My results, however, show that dispersal is local, with queens establishing new colonies as close as 60 meters from their natal colony. Even though actual sib-mating was rare, individuals from different but related colonies pair, which causes the population to be inbred. Furthermore, multiple mates of queens were related to each other, which also indicates localized mating flights. Hence, known mechanisms of inbreeding avoidance, dispersal and multiple mating, were not effective in this population, as neither reduced inbreeding level of the future colony. Inbreeding had negative consequences both at the individual and colony level. A queen that has mated with a related male produces inbred workers, which impairs the colony s reproductive success. The inbred colonies were less productive and, specifically, produced fewer new queens, possibly owing to effects of inbreeding on the caste determination of female larvae. A striking finding was that males raised in colonies with inbred workers were smaller, which reflects an effect of the social environment as males, being haploid, cannot be inbred themselves. The queens produced in the inbred colonies, in contrast, were not smaller, but their immune response was up-regulated. Inbreeding had no effect on queen dispersal, but inbred queens had a lower probability of successfully founding a new colony. Ultimately, queens that survived through the colony founding phase had a shorter lifespan. This supports the idea that inbreeding imposes a genetic stress, leading to inbreeding depression on both the queen and the colony level. My results show that inbreeding can have profound consequences on insects in the wild, and that in social species the effects of inbreeding may be multiplicative and mediated through the diversity of the social environment, as well as the genetic makeup of the individuals themselves. This emphasizes the need to take into account all levels of organization when assessing the effects of genetic diversity in social animals.

Natural variation and Biogeography of the melon fruit fly, Zeugodacus cucurbitae (Diptera : Tephritidae), in Southeast-Asia and the west-pacific

This thesis used multidisciplinary approaches which greatly enhance our understanding of population structure and can be particularly powerful tools for resolving variation of melon fly over geographic and temporal scales, and for determining invasive pathways. The results from this thesis reinforce the value of integrating multiple data sets to better understand and resolve natural variation within an important pest to determine whether there are cryptic species, discrete lineages or host races, and to identify dispersal pathways in an invasive pest. These results are instructive for regional biosecurity, trade and quarantine, and provide important background for future area-wide management programmes. The integrative methodology adopted in this thesis is applicable to a variety of other insect pests.

Molecular modeling of Bt Cry1Ac (DI–DII)–ASAL (Allium sativum lectin)–fusion protein and its interaction with aminopeptidase N (APN) receptor of Manduca sexta

Genetic engineering of Bacillus thuringiensis (Bt) Cry proteins has resulted in the synthesis of various novel toxin proteins with enhanced insecticidal activity and specificity towards different insect pests. In this study, a fusion protein consisting of the DI–DII domains of Cry1Ac and garlic lectin (ASAL) has been designed in silico by replacing the DIII domain of Cry1Ac with ASAL. The binding interface between the DI–DII domains of Cry1Ac and lectin has been identified using protein–protein docking studies. Free energy of binding calculations and interaction profiles between the Cry1Ac and lectin domains confirmed the stability of fusion protein. A total of 18 hydrogen bonds was observed in the DI–DII–lectin fusion protein compared to 11 hydrogen bonds in the Cry1Ac (DI–DII–DIII) protein. Molecular mechanics/Poisson–Boltzmann (generalized-Born) surface area [MM/PB (GB) SA] methods were used for predicting free energy of interactions of the fusion proteins. Protein–protein docking studies based on the number of hydrogen bonds, hydrophobic interactions, aromatic–aromatic, aromatic–sulphur, cation–pi interactions and binding energy of Cry1Ac/fusion proteins with the aminopeptidase N (APN) of Manduca sexta rationalised the higher binding affinity of the fusion protein with the APN receptor compared to that of the Cry1Ac–APN complex, as predicted by ZDOCK, Rosetta and ClusPro analysis. The molecular binding interface between the fusion protein and the APN receptor is well packed, analogously to that of the Cry1Ac–APN complex. These findings offer scope for the design and development of customized fusion molecules for improved pest management in crop plants.

Ecological consequences of genetic modifications - an invasion analysis approach

Biological invasions are considered as one of the greatest threats to biodiversity, as they may lead to disruption and homogenization of natural communities, and in the worst case, to native species extinctions. The introduction of gene modified organisms (GMOs) to agricultural, fisheries and forestry practices brings them into contact with natural populations. GMOs may appear as new invasive species if they are able to (1) invade into natural habitats or (2) hybridize with their wild relatives. The benefits of GMOs, such as increased yield or decreased use of insecticides or herbicides in cultivation, may thus be reduced due the potential risks they may cause. A careful ecological risk analysis therefore has to precede any responsible GMO introduction. In this thesis I study ecological invasion in relation to GMOs, and what kind of consequences invasion may have in natural populations. A set of theoretical models that combine life-history evolution, population dynamics, and population genetics were developed for the hazard identification part of ecological risks assessment of GMOs. In addition, the potential benefits of GMOs in management of an invasive pest were analyzed. In the first study I showed that a population that is fluctuating due to scramble-type density dependence (due to, e.g., nutrient competition in plants) may be invaded by a population that is relatively more limited by a resource (e.g., light in plants) that is a cause of contest-type density dependence. This result emphasises the higher risk of invasion in unstable environments. The next two studies focused on escape of a growth hormone (GH) transgenic fish into a natural population. The results showed that previous models may have given too pessimistic a view of the so called Trojan gene -effect, where the invading genotype is harmful for the population as a whole. The previously suggested population extinctions did not occur in my studies, since the changes in mating preferences caused by the GH-fish were be ameliorated by decreased level of competition. The GH-invaders may also have to exceed a threshold density before invasion can be successful. I also showed that the prevalence of mature parr (aka. sneaker) strategy among GH-fish may have clear effect on invasion outcome. The fourth study assessed the risks and developed methods against the invasion of the Colorado Potato Beetle (CPB, Leptinotarsa decemlineata). I showed that the eradication of CPB is most important for the prevention of their establishment, but the cultivation of transgenic Bt-potato could also be effective. In general, my results emphasise that invasion of transgenic species or genotypes to be possible under certain realistic conditions and resulting in competitive exclusion, population decline through outbreeding depression and genotypic displacement of native species. Ecological risk assessment should regard the decline and displacement of the wild genotype by an introduced one as a consequence that is as serious as the population extinction. It will also be crucial to take into account different kinds of behavioural differences among species when assessing the possible hazards that GMOs may cause if escaped. The benefits found of GMO crops effectiveness in pest management may also be too optimistic since CPB may evolve resistance to Bt-toxin. The models in this thesis could be further applied in case specific risk assessment of GMOs by supplementing them with detailed data of the species biology, the effect of the transgene introduced to the species, and also the characteristics of the populations or the environments in the risk of being invaded.

Population structure and evolution in the ant Plagiolepis pygmaea and its two social parasites Plagiolepis xene and Plagiolepis grassei

Social behaviour affects dispersal of animals and is an important modifier of genetic population structures. The female sex is often philopatric, which maintains coancestry within the breeding groups and promotes cooperative behaviours. This enables also inclusive fitness returns from altruism and explains why some individuals sacrifice personal reproduction for the good of others in social insects such as ants. However, reduced dispersal and population substructuring at the level of colonies may also entail inbreeding, loss of genetic diversity, and vulnerability. In addition, the most vulnerable ants are species that are evolved to parasitize colonies of other ants, and which compromise between abilities to disperse and the efficiency to parasitize the host. On the other hand, certain social organisations of ant colonies may facilitate a species to disperse outside its natural range and become a pest. Altogether, knowledge on genetic structuring of ant populations, as well as the evolution of their life histories can contribute to conservation biology and population management. The aim of this thesis was to investigate population structures and phylogenetic evolution of the ant Plagiolepis pygmaea and its two obligatory, workerless social parasites (inquilines) P. xene and P. grassei with genetic markers and DNA sequence data. The results support the general assumption that populations of inquiline parasites are highly fragmented and genetically vulnerable. Comparison of the two parasites suggests that differences in their relative abundance may follow from their interaction with the host, i.e. how well the species is adapted to reproduce in the host colonies. The results also indicate that the most recent free living ancestor to these two parasite species is their common host. This is considered to provide evidence for the controversial issue of sympatric speciation. Further, given that the level of adaptations to parasitic life history depends on the evolutionary time since the free-living ancestor, the results establish a link between species rarity and its evolutionary age. The populations of the host species P. pygmaea displayed significantly reduced dispersal both among the females (queens) and males, and high levels of inbreeding which may enhance worker altruism. In addition, the queens were found to mate with multiple males. Given the high relatedness between the queens and their mates, this occurs probably for non-genetic reasons, e.g. without benefits associated in genetically more diverse offspring. The results hence caution that the contribution of non-genetic factors to the prevailing mating patterns and genetic population structures should not be underestimated.

Using Bayesian networks to model surveillance in complex plant and animal health systems

In this chapter we consider biosecurity surveillance as part of a complex system comprising many different biological, environmental and human factors and their interactions. Modelling and analysis of surveillance strategies should take into account these complexities, and also facilitate the use and integration of the many types of different information that can provide insight into the system as a whole. After a brief discussion of a range of options, we focus on Bayesian networks for representing such complex systems. We summarize the features of Bayesian networks and describe these in the context of surveillance.

Assessment of crop insect damage using unmanned aerial systems: A machine learning approach

Agricultural pests are responsible for millions of dollars in crop losses and management costs every year. In order to implement optimal site-specific treatments and reduce control costs, new methods to accurately monitor and assess pest damage need to be investigated. In this paper we explore the combination of unmanned aerial vehicles (UAV), remote sensing and machine learning techniques as a promising technology to address this challenge. The deployment of UAVs as a sensor platform is a rapidly growing field of study for biosecurity and precision agriculture applications. In this experiment, a data collection campaign is performed over a sorghum crop severely damaged by white grubs (Coleoptera: Scarabaeidae). The larvae of these scarab beetles feed on the roots of plants, which in turn impairs root exploration of the soil profile. In the field, crop health status could be classified according to three levels: bare soil where plants were decimated, transition zones of reduced plant density and healthy canopy areas. In this study, we describe the UAV platform deployed to collect high-resolution RGB imagery as well as the image processing pipeline implemented to create an orthoimage. An unsupervised machine learning approach is formulated in order to create a meaningful partition of the image into each of the crop levels. The aim of the approach is to simplify the image analysis step by minimizing user input requirements and avoiding the manual data labeling necessary in supervised learning approaches. The implemented algorithm is based on the K-means clustering algorithm. In order to control high-frequency components present in the feature space, a neighbourhood-oriented parameter is introduced by applying Gaussian convolution kernels prior to K-means. The outcome of this approach is a soft K-means algorithm similar to the EM algorithm for Gaussian mixture models. The results show the algorithm delivers decision boundaries that consistently classify the field into three clusters, one for each crop health level. The methodology presented in this paper represents a venue for further research towards automated crop damage assessments and biosecurity surveillance.

Revisiting Coptotermes (Isoptera: Rhinotermitidae): a global taxonomic road map for species validity and distribution of an economically important subterranean termite genus

Coptotermes Wasmann (Isoptera: Rhinotermitidae) is one of the most economically important subterranean termite genera and some species are successful invaders. However, despite its important pest status, the taxonomic validity of many named Coptotermes species remains unclear. In this study, we reviewed all named species within the genus and investigated evidence supporting the validity of each named species. Species were systematically scrutinized according to the region of their original description: Southeast Asia, India, China, Africa, the Neotropics, and Australia. We estimate that of the currently 69 named species described by accepted nomenclatural rules, only 21 taxa have solid evidence for validity, 44 names have uncertain status, and the remaining species names should be synonymized or were made unavailable. Species with high degrees of invasiveness may be known under additional junior synonyms due to independent parochial descriptions. Molecular data for a vast majority of species are scarce and significant effort is needed to complete the taxonomic and phylogenetic revision of the genus. Because of the wide distribution of Coptotermes, we advocate for an integrative taxonomic effort to establish the distribution of each putative species, provide specimens and corresponding molecular data, check original descriptions and type specimens (if available), and provide evidence for a more robust phylogenetic position of each species. This study embodies both consensus and contention of those studying Coptotermes and thus pinpoints the current uncertainty of many species. This project is intended to be a roadmap for identifying those Coptotermes species names that need to be more thoroughly investigated, as an incentive to complete a necessary revision process.

'Ladd traps' as a visual trap for male and female Queensland fruit fly, Bactrocera tryoni (Diptera: Tephritidae)

Bactrocera tryoni (Froggatt) is Australia's major horticultural insect pest, yet monitoring females remains logistically difficult. We trialled the ‘Ladd trap’ as a potential female surveillance or monitoring tool. This trap design is used to trap and monitor fruit flies in countries other (e.g. USA) than Australia. The Ladd trap consists of a flat yellow panel (a traditional ‘sticky trap’), with a three dimensional red sphere (= a fruit mimic) attached in the middle. We confirmed, in field-cage trials, that the combination of yellow panel and red sphere was more attractive to B. tryoni than the two components in isolation. In a second set of field-cage trials, we showed that it was the red-yellow contrast, rather than the three dimensional effect, which was responsible for the trap's effectiveness, with B. tryoni equally attracted to a Ladd trap as to a two-dimensional yellow panel with a circular red centre. The sex ratio of catches was approximately even in the field-cage trials. In field trials, we tested the traditional red-sphere Ladd trap against traps for which the sphere was painted blue, black or yellow. The colour of sphere did not significantly influence trap efficiency in these trials, despite the fact the yellow-panel/yellow-sphere presented no colour contrast to the flies. In 6 weeks of field trials, over 1500 flies were caught, almost exactly two-thirds of them being females. Overall, flies were more likely to be caught on the yellow panel than the sphere; but, for the commercial Ladd trap, proportionally more females were caught on the red sphere versus the yellow panel than would be predicted based on relative surface area of each component, a result also seen the field-cage trial. We determined that no modification of the trap was more effective than the commercially available Ladd trap and so consider that product suitable for more extensive field testing as a B. tryoni research and monitoring tool.