Identification of small juvenile scombrids from northwest tropical Australia using mitochondrial DNA cytochrome b sequences

Small juveniles of the nine species of scombrids in Australian waters are morphologically similar to one another and, consequently, difficult to identify to species level. We show that the sequence of the mitochondrial DNA cytochrome b gene region is a powerful tool for identification of these young fish. Using this method, we identified 50 juvenile scombrids collected from Exmouth Bay, Western Australia. Six species of scombrids were apparent in this sample of fish: narrow-barred Spanish mackerel (Scomberomorus commerson), Indian mackerel (Rastrelliger kanagurta), frigate tuna (Auxis thazard), bullet tuna (Auxis rochei), leaping bonito (Cybiosarda elegans), and kawakawa (Euthynnus affinis). The presence of Indian mackerel, frigate tuna, leaping bonito, and kawakawa is the first indication that coastal waters may be an important spawning habitat for these species, although offshore spawning may also occur. The occurrence of small juvenile S. commerson was predicted from the known spawning patterns of that species, but other mackerel species (Scomberomorus munroi, Scomberomorus queenslandicus, Scomberomorus semifasiciatus) likely to be spawning during the sampling period were not detected among the 50 small juveniles analyzed here.

Evolutionary relationships between bat coronaviruses and their hosts

Recent studies have suggested that bats are the natural reservoir of a range of coronaviruses (CoVs), and that rhinolophid bats harbor viruses closely related to the severe acute respiratory syndrome (SARS) CoV, which caused an outbreak of respiratory illness in humans during 2002-2003. We examined the evolutionary relationships between bat CoVs and their hosts by using sequence data of the virus RNA-dependent RNA polymerase gene and the bat cytochrome b gene. Phylogenetic analyses showed multiple incongruent associations between the phylogenies of rhinolophid bats and their CoVs, which suggested that host shifts have occurred in the recent evolutionary history of this group. These shifts may be due to either virus biologic traits or host behavioral traits. This finding has implications for the emergence of SARS and for the potential future emergence of SARS-CoVs or related viruses.

Evidence for extensive population structure in the white-spotted eagle ray within the Indo-Pacific inferred from mitochondrial gene sequences

The white-spotted eagle ray Aetobatus narinari is a species complex that occurs circumglobally throughout warm-temperate waters. Aetobatus narinari is semi-pelagic and large (up to 300 cm disc width), suggesting high dispersal capabilities and gene flow on a wide spatial scale. Sequence data from two mitochondrial genes, cytochrome b (cytb) and NADH dehydrogenase subunit 4 (ND4), were used to determine the genetic variability within and among 18 sampling locations in the central Indo-Pacific biogeographical region. Populations in the Indo-Pacific were highly genetically structured with c. 70% of the total genetic variation found among three geographical regions (East China Sea, Southeast Asia and Australia). FST was 0.64 for cytb and 0.53 for ND4, with φST values being even larger, that is, 0.78 for cytb and 0.65 for ND4. This high-level genetic partitioning provides strong evidence against extensive gene flow in A. narinari. The degree of genetic population structuring in the Indo-Pacific was similar to that found on a global scale. Global FST was 0.63 for cytb and 0.57 for ND4, and global φST values were 0.94 for cytb and 0.82 for ND4. This suggests that the A. narinari complex may be more speciose than the two or three species proposed to date. Further sampling and genetic analyses are likely to uncover the ‘evolutionarily significant’ and ‘management’ units that are critical to determine the susceptibilities of individual populations to regional fishing pressures and to provide advice on management options. Network analyses showed a close genetic relationship between haplotypes from the central Indo-Pacific and South Africa, providing support for a proposed dispersal pathway from the possible centre of origin of the A. narinari species complex in the Indo-Pacific into the Atlantic Ocean.

Cytochrome P450 aromatase in grey mullet: cDNA and promoter isolation; brain, pituitary and ovarian expression during puberty.

In a study towards elucidating the role of aromatases during puberty in female grey mullet, the cDNAs of the brain (muCyp19b) and ovarian (muCyp19a) aromatase were isolated by RT-PCR and their relative expression levels were determined by quantitative real-time RT-PCR. The muCyp19a ORF of 1515 bp encoded 505 predicted amino acid residues, while that of muCyp19b was 1485 bp and encoded 495 predicted amino acid residues. The expression level of muCyp19b significantly increased in the brain as puberty advanced; however, its expression level in the pituitary increased only slightly with pubertal development. In the ovary, the muCyp19a expression level markedly increased as puberty progressed. The promoter regions of the two genes were also isolated and their functionality evaluated in vitro using luciferase as the reporter gene. The muCyp19a promoter sequence (650 bp) contained a consensus TATA box and putative transcription factor binding sites, including two half EREs, an SF-1, an AhR/Arnt, a PR and two GATA-3s. The muCyp19b promoter sequence (2500 bp) showed consensus TATA and CCAAT boxes and putative transcription binding sites, namely: a PR, an ERE, a half ERE, a SP-1, two GATA-binding factor, one half GATA-1, two C/EBPs, a GRE, a NFkappaB, three STATs, a PPAR/RXR, an Ahr/Arnt and a CRE. Basal activity of serially deleted promoter constructs transiently transfected into COS-7, [alpha]T3 and TE671 cells demonstrated the enhancing and silencing roles of the putative transcription factor binding sites. Quinpirole, a dopamine agonist, significantly reduced the promoter activity of muCyp19b in TE671. The results suggest tissue-specific regulation of the muCyp19 genes and a putative alternative promoter for muCyp19b.

Refined Global Analysis of Bemisia tabaci (Hemiptera: Sternorrhyncha: Aleyrodoidea: Aleyrodidae) Mitochondrial Cytochrome Oxidase 1 to Identify Species Level Genetic Boundaries.

Identifying species boundaries within morphologically indistinguishable cryptic species complexes is often contentious. For the whitefly Bemisia tabaci (Gennadius) (Hemiptera: Sternorrhyncha: Aleyrodoidea: Aleyrodidae), the lack of a clear understanding about the genetic limits of the numerous genetic groups and biotypes so far identified has resulted in a lack of consistency in the application of the terms, the approaches use to apply them and in our understanding of what genetic structure within B. tabaci means. Our response has been to use mitochondrial gene cytochrome oxidase one to consider how to clearly and consistently define genetic separation. Using Bayesian phylogenetic analysis and analysis of sequence pairwise divergence we found a considerably higher to number of genetic groups than had been previously determined with two breaks in the distribution, one at 11% and another at 3.5%. At >11% divergence, 11 distinct groups were resolved, whereas at >3.5% divergence 24 groups were identified. Consensus sequences for each of these groups were determined and were shown to be useful in the correct assignment of sequences of unknown origin. The 3.5% divergence bound is consistent with species level separations in other insect taxa and Suggests that B. tabaci is it cryptic species composed of at least 24 distinct species. We further show that the placement of Bemesia atriplex (Froggatt) within the B. tabaci in, group adds further weight to the argument for species level separation within B. tabaci. This new analysis, which constructs consensus sequences and uses these its a standard against which unknown sequences call be compared, provides for the first time it consistent means of identifying the genetic hounds of each species with it high degree of certainty.

Floating row covers and pyriproxyfen help control silverleaf whitefly Bemisia tabaci (Gennadius) Biotype B (Homoptera: Aleyrodidae) in zucchini

In zucchini, the use of row covers until flowering and the insect growth regulator (IGR) pyriproxyfen are effective methods of reducing the number of insects, especially silverleaf whitefly (Bemisia tabaci (Gennadius) Biotype B), on plants. We compared floating row covers (FRCs) up until flowering with silverleaf whitefly (SLW) introduced (FRC + SLW), or not introduced (FRC-only), or with introduction of SLW in open plots (SLW-only), or with introduction of SLW in open plots with IGR (SLW + IGR). FRC increased temperature and humidity compared with the uncovered treatments. Average fruit weight was less (P < 0.01) for the FRC + SLW treatment compared with the other treatments and the percentage of marketable fruit was less for the FRC + SLW than for the other three treatments. This result indicates that the use of either row covers or IGR controls whiteflies, reduces fruit damage and increases the size, weight, and quality of fruit, and may also control other sap-sucking insects. However, if SLW are already present on plants, the use of FRC may reduce predation and favour build up of SLW. Thus, FRC and IGR, if used judiciously, may provide an effective alternative to broad-spectrum pesticides in small-scale cucurbit production.

Sampling and management of Bemisia tabaci (Genn.) biotype B in Australian cotton

Data on seasonal population abundance of Bemisia tabaci biotype B (silverleaf whitefly (SLW)) in Australian cotton fields collected over four consecutive growing seasons (2002/2003-2005/2006) were used to develop and validate a multiple-threshold-based management and sampling plan. Non-linear growth trajectories estimated from the field sampling data were used as benchmarks to classify adult SLW field populations into six density-based management zones with associated control recommendations in the context of peak flowering and open boll crop growth stages. Control options based on application of insect growth regulators (IGRs) are recommended for high-density populations (>2 adults/leaf) whereas conventional (non-IGR) products are recommended for the control of low to moderate population densities. A computerised re-sampling program was used to develop and test a binomial sampling plan. Binomial models with thresholds of T=1, 2 and 3 adults/leaf were tested using the field abundance data. A binomial plan based on a tally threshold of T=2 adults/leaf and a minimum sample of 20 leaves at nodes 3, 4 or 5 below the terminal is recommended as the most parsimonious and practical sampling protocol for Australian cotton fields. A decision support guide with management zone boundaries expressed as binomial counts and control options appropriate for various SLW density situations is presented. Appropriate use of chemical insecticides and tactics for successful field control of whiteflies are discussed.

Pyriproxyfen controls silverleaf whitefly, Bemisia tabaci (Gennadius), biotype B (Homoptera: Aleyrodidae) (SLW) better than buprofezin in bitter melons Momordica charantia L. (Cucurbitaceae)

To improve compatibility between chemical and biological controls, the use of selective insecticides such as insect growth regulators (IGRs) is crucial. In cucurbits, the use of pyriproxyfen (an IGR) has been shown by others to be an effective method of reducing the number of sap-sucking insects, especially silverleaf whitefly, Bemisia tabaci (Gennadius) Biotype B (SLW). Therefore, we compared pyriproxyfen and buprofezin (an IGR) with that of no treatment (control) in a bitter melon crop for the control of populations of SLW and for their effects on fruit production. Pyriproxyfen controlled SLW and tended to have heavier fruits than the control treatment and reduced the abundance of nymphs and exuvia. Buprofezin showed no evidence in controlling SLW compared with the pyriproxyfen and control treatments. Neither pyriproxyfen nor buprofezin had any effect on the number of harvested fruit or overall fruit yield, but the average weight per fruit was higher than the control treatment. Pyriproxyfen was effective in controlling whitefly populations in bitter melons, and both pyriproxyfen and buprofezin may have the potential to increase yield. Their longer-term use may increase predation by natural enemies as they are species-specific and could favour build up of natural enemies of SLW. Thus, the judicious use of pyriproxyfen may provide an effective alternative to broad-spectrum insecticides in small-scale cucurbit production.

New efforts at biological control of Mikania micrantha H.B.K. (Asteraceae) in Papua New Guinea and Fiji.

New efforts at biological control of Miconia calvescens (Melastomataceae) is a serious invader in the tropical Pacific, including the Hawaiian and Tahitian Islands, and currently poses a major threat to native biodiversity in the Wet Tropics of Australia. The species is fleshy-fruited, small-seeded and shade tolerant, and thus has the potential to be dispersed widely and recruit in relatively intact rainforest habitats, displacing native species. Understanding and predicting the rate of spread is critical for the design and implementation of effective management actions. We used an individual-based model incorporating a dispersal function derived from dispersal curves for similar berry-fruited native species, and life-history parameters of fecundity and mortality to predict the spatial structure of a Miconia population after a 30 year time period. We compared the modelled population spatial structure to that of an actual infestation in the rainforests of north Queensland. Our goal was to assess how well the model predicts actual dispersion and to identify potential barriers and conduits to seed movement and seedling establishment. The model overpredicts overall population size and the spatial extent of the actual infestation, predicting individuals to occur at a maximum 1,750 m from the source compared with the maximum distance of any detected individual in the actual infestation of 1,191 m. We identify several characteristic features of managed invasive populations that make comparisons between modelled outcomes and actual infestations difficult. Our results suggest that the model’s ability to predict both spatial structure and spread of the population will be improved by incorporating a spatially explicit element, with dispersal and recruitment probabilities that reflect the relative suitability of different parts of the landscape for these processes. Mikania micrantha H.B.K. (Asteraceae) in Papua New Guinea and Fiji.

Inter-seasonal population dynamics and pest status of Bemisia tabaci (Gennadius) biotype B in an Australian cropping system

Bemisia tabaci, biotype B, commonly known as the silverleaf whitefly (SLW) is an alien species that invaded Australia in the mid-90s. This paper reports on the invasion ecology of SLW and the factors that are likely to have contributed to the first outbreak of this major pest in an Australian cotton cropping system, population dynamics of SLW within whitefly-susceptible crop (cotton and cucurbit) and non-crop vegetation (sowthistle, Sonchus spp.) components of the cropping system were investigated over four consecutive growing seasons (September-June) 2001/02-2004/05 in the Emerald Irrigation Area (EIA) of Queensland, Australia. Based on fixed geo-referenced sampling sites, variation in spatial and temporal abundance of SLW within each system component was quantified to provide baseline data for the development of ecologically sustainable pest management strategies. Parasitism of large (3rd and 4th instars) SLW nymphs by native aphelinid wasps was quantified to determine the potential for natural control of SLW populations. Following the initial outbreak in 2001/02, SLW abundance declined and stabilised over the next three seasons. The population dynamics of SLW is characterised by inter-seasonal population cycling between the non-crop (weed) and cotton components of the EIA cropping system. Cotton was the largest sink for and source of SLW during the study period. Over-wintering populations dispersed from weed host plant sources to cotton in spring followed by a reverse dispersal in late summer and autumn to broad-leaved crops and weeds. A basic spatial source-sink analysis showed that SLW adult and nymph densities were higher in cotton fields that were closer to over-wintering weed sources throughout spring than in fields that were further away. Cucurbit fields were not significant sources of SLW and did not appear to contribute significantly to the regional population dynamics of the pest. Substantial parasitism of nymphal stages throughout the study period indicates that native parasitoid species and other natural enemies are important sources of SLW mortality in Australian cotton production systems. Weather conditions and use of broad-spectrum insecticides for pest control are implicated in the initial outbreak and on-going pest status of SLW in the region.

Evaluation of wood characteristics of tropical post-mid rotation plantation Eucalyptus cloeziana and E. pellita: Part (b) Accelerated seasoning of sawn timber.

Drying trials were conducted using two species of plantation grown eucalypt timbers: 19-year-old Eucalyptus cloeziana (Gympie messmate) and 15-year-old Eucalyptus pellita (red mahogany). The objective of this study was to gain an understanding of the drying potential of young plantation grown material using accelerated seasoning methods, a process expected to be critcal to the success of plantation hardwood products entering value added markets. The findings are encouraging, indicating that both species can be dried using conventional drying techniques much faster than industry is currently achieving when drying native forest timber. The results suggest that there is a definite drying time advantatge in vacuum drying over conventional methods for 19-year-old E. cloeziana. The findings have shown that through careful schedule manipulation and adjustment, the grade quality can be optimised to suit the desired expectation. As this study was limited to only a small number of trials, time and quality improvements are expected to be realised for both conventional and vacuum drying methods as more research is conducted.

Effectiveness of Various Artificial Larval Diets for Rearing Bactrocera passiflorae (Froggatt) and B. xanthodes (Broun) in the Laboratory in Fiji

Laboratory colonies of Bactrocera passiflorae (Froggatt) and B. xanthodes (Broun) were established at Koronivia Research Station, Fiji in 1991. Laboratory rearing of the two economically important species was a prerequisite to studies conducted on protein bait spray and quarantine treatment development. To increase the production of laboratory reared fruit flies for this research and also to have a substitute larval diet available, replicated comparisons of the effectiveness of larval diets were carried out using B. passiflorae and B. xanthodes. The diets compared were pawpaw/bagasse, dehydrated carrot and diets used for culturing Mediterranean fruit fly (Ceratitis capitata Wiedemann), Oriental fruit fly (B. dorsalis Hendel), melon fly (B. cucurbitae Coquillett) and B. latifrons (Hendel), pawpaw diet and breadfruit diet. B. passiflorae and B. xanthodes eggs seeded onto the various diets were allowed to develop into larvae, pupae and adults. The percentage egg hatch, number of pupae recovered, percentage pupal mortality, weight of 100 pupae, number of adults and percentage eclosion were used to determine the effectiveness of the diets. Results showed that pawpaw/bagasse and dehydrated carrot diets performed favorably for both species. The pawpaw diet currently used as standard larval diets for both species is the most readily available and easiest to use. Breadfruit diet was tested on B. xanthodes only and showed that it was a suitable substitute for the pawpaw-based diets. Other larval diets, cassava/pawpaw and banana diets, that have been developed and used in the South Pacific areas are also discussed in this paper. When pawpaw or breadfruit are not available, dehydrated carrot diet may be substituted for fruit-based larval diets.

Bovine cysticercosis-Development of a real-time PCR to enhance classification of suspect cysts identified at meat inspection

Laboratory confirmation methods are important in bovine cysticerosis diagnosis as other pathologies can result in morphologically similar lesions resulting in false identifications. We developed a probe-based real-time PCR assay to identify Taenia saginata in suspect cysts encountered at meat inspection and compared its use with the traditional method of identification, histology, as well as a published nested PCR. The assay simultaneously detects T. saginata DNA and a bovine internal control using the cytochrome c oxidase subunit 1 gene of each species and shows specificity against parasites causing lesions morphologically similar to those of T. saginata. The assay was sufficiently sensitive to detect 1 fg (Ct 35.09 +/- 0.95) of target DNA using serially-diluted plasmid DNA in reactions spiked with bovine DNA as well as in all viable and caseated positive control cysts. A loss in PCR sensitivity was observed with increasing cyst degeneration as seen in other molecular methods. In comparison to histology, the assay offered greater sensitivity and accuracy with 10/19 (53%) T. saginata positives detected by real-time PCR and none by histology. When the results were compared with the reference PCR, the assay was less sensitive but offered advantages of faster turnaround times and reduced contamination risk. Estimates of the assay's repeatability and reproducibility showed the assay is highly reliable with reliability coefficients greater than 0.94. Crown Copyright (C) 2013 Published by Elsevier B.V. All rights reserved.

Mitochondrial DNA diversity of Cleruchoides noackae (Hymenoptera: Mymaridae): a potential biological control agent for Thaumastocoris peregrinus (Hemiptera: Thaumastocoridae)

Carpintero and Dellap, (Hemiptera: Thaumastocoridae) is a native Australian sap-feeding insect that has become invasive and seriously damaging to commercially grown in the Southern Hemisphere. Lin and Huber (Hymenoptera: Mymaridae) was recently discovered as an egg parasitoid of the Thaumastocoridae in Australia. Mitochondrial DNA (mtDNA; cytochrome oxidase subunit I, COI) sequence diversity amongst 104 individuals from these native populations revealed 24 sequence haplotypes. The COI haplotypes of individuals collected from the Sydney and Southeast Queensland clustered in distinct groups, indicating limited spread of the insect between the regions. Individuals collected from Perth in Western Australia were represented by four COI haplotypes. Although this population is geographically more isolated from other populations, two COI haplotypes were identical to haplotypes found in the Sydney region. The results suggest that has recently been introduced into Perth, possibly from the Sydney area. The high mtDNA diversity and limited spread that is suggested for is in contrast to the lack of geographic associated mtDNA diversity and extensive spread of . If implemented as a biological control agent, this factor will need to be considered in collecting and releasing .

Continuous combined microwave and hot air treatment of apples for fruit fly (Bactrocera tryoni and B. jarvisi) disinfestation

Apples at 24 ± 2 °C were heated in a pilot scale hot air assisted (40 °C) continuous pentagonal microwave system, to evaluate the effectiveness of this treatment on insect mortality (variety Mutsu) and fruit quality (variety Granny Smith). An average temperature of 53.4 ± 1.3 °C at core, bottom and flesh of the apple was recorded at the end of the treatment. One hundred percent mortality of the most tolerant stage of Queensland fruit fly (Bactrocera tryoni, Froggatt) and Jarvis's fruit fly (Bactrocera jarvisi, Tryon), were observed when the Mortality value (M52, equivalent time of isothermal treatment at 52 °C) at the slowest heating point applicable for each experiment was ≥ 50 min and ≥ 37 min, respectively. Results showed that microwave heat treatment is effective for insect disinfestation without any adverse impact on total soluble solids, flesh or peel firmness of the treated apples. The treated apples recorded a significantly higher pH and lower ion leakage than the untreated apples after 3 or 4 weeks. Therefore, the microwave heat treatment has the potential to be developed as an alternative chemical free quarantine treatment against economically significant insect pests. Industrial relevance Hot air assisted microwave heating of fruits and vegetables, is more cost effective compared to vapour heat treatment and ionising radiation for disinfestation of insects. Microwave treatment is environmentally friendly compared to fumigation and chemical treatments. Hot air assisted microwave disinfestation can be performed at farms or centralised pack houses since the capital cost would be comparatively lower than vapour heat or ionising radiation treatments.