Pre-release dietary supplements of methoprene and raspberry ketone increase field abundance of sterile Queensland fruit flies (Diptera: Tephritidae).

The sterile insect technique (SIT) is a sustainable pest management tool based on the release of millions of sterile insects that suppress reproduction in targeted populations. Success of SIT depends on survival, maturation, dispersal, and mating of released sterile insects. Laboratory and field cage studies have demonstrated that dietary supplements of methoprene and raspberry ketone (RK) promote sexual maturation of adult Queensland fruit fly, Bactrocera tryoni (Froggatt), and may hence shorten the delay between release and maturity in the field. We investigated the effects of methoprene and RK dietary supplements on field abundance of sexually mature sterile Q-flies relative to untreated flies fed only sugar and yeast hydrolysate before release at 2 d of age. Compared with untreated flies, more methoprene- and RK-treated flies were recaptured in cuelure traps to which only sexually mature males are attracted. At distances of 100 and 200 m from the release point, recapture rates were higher for methoprene- and RK-treated flies than for untreated flies, but at 300 m recapture rates were low and were similar for treated and untreated flies. Rainfall, relative humidity, wind speed, and wind direction did not affect recapture rates, but temperature was positively correlated with recapture rates for all treatments. There was a strong correlation between the number of sterile and wild flies caught in traps, indicating co-location in the field. Dietary supplements of methoprene and RK can substantially increase abundance of sexually mature sterile male Q-flies in the field following release as 2-d-old immature adults.

Dietary methoprene treatment promotes rapid development of reproductive organs in male Queensland fruit fly.

Methoprene supplements added to diets of yeast hydrolysate and sugar promote early expression of sexual behaviour and mating in male Queensland fruit fly (Bactrocera tryoni; 'Q-fly') and show promise as a pre-release treatment for sterile insect technique programs. Currently it is not known whether the early mating behaviour of methoprene-treated male Q-flies is only behavioural or is coupled with accelerated development of reproductive organs. Accordingly, the present study investigates whether incorporation of methoprene into diets of yeast hydrolysate and sugar (1:3) or sugar alone, accelerate development of testes, ejaculatory apodeme, and accessory glands in male Q-flies and ovaries in females. All organs increased in size as the flies aged and matured, and development rate of all organs was far greater when the flies were provided yeast hydrolysate in addition to sugar. Incorporation of methoprene into diets containing yeast hydrolysate was found to strongly accelerate development of testes and ejaculatory apodeme, but not accessory glands, in males. In the absence of yeast hydrolysate, methoprene treatment had only a modest effect on male organ development. In contrast to males, development of ovaries in female Q-flies did not respond to dietary methoprene supplements, regardless of whether they were fed yeast hydrolysate and sugar or sugar alone. These findings of diet-dependent effects of methoprene supplements on reproductive organs are a close match to previous studies investigating effects of methoprene supplements on mating behaviour. Overall, methoprene supplements substantially enhance the positive effects of protein rich adult diet on the early expression of sexual behaviour and accelerate development of reproductive organs in male, but not female, Q-flies. Methoprene supplements added to pre-release diets of yeast hydrolysate and sugar show promise as a means of accelerating reproductive development of Q-flies released in sterile insect technique programs, and may also bias operational sex ratio in favour of males.

Ammoides verticillata Essential Oil as Biocontrol Agent of Selected Fungi and Pest of Olive Tree.

BACKGROUND: Ammoides verticillata Briq is an aromatic and medicinal herb that has been widely used in folk medicine for treatment of several types of pathologies such as respiratory problems, colds, fever, headache, gastric disorders and renal infections. OBJECTIVE: Essential oils have been subjected of numerous works for their antimicrobial and insecticidal properties which have been valued by hundreds of publications and patents. The main objective of this work was to evaluate the chemical composition, and the in vitro and in vivo antifungal, and insecticidal properties of essential oil of Ammoides verticillata. METHODS: In this work, the air-dried aerial parts of Ammoides verticillata were hydrodistilled in a Clevenger-type apparatus. The essential oil isolated was analyzed using gas chromatography (GC) and mass spectrometry (GC/MS). The in vitro antifungal activity of the essential oil was investigated against four plant fungi using radial growth technique. The effect of the essential oil on disease development of olive caused by Penicillium crustosum and Aspergillus niger in the in vivo conditions. The fumigation test of the essential oil was evaluated against L3 stages, pupas and adults of Bactrocera oleae known as the olive fly. RESULTS: The essential oil of A. verticillata was characterized mainly by carvacrol (44.3%), limonene (19.3%) and p-cymene (19.2%). The result of in vitro antifungal activity of essential oil showed an interesting antifungal inhibition against Alternaria alternata and Fusarium solani strains with percentage inhibition of 89%. Furthermore, oil of A. verticillata has demonstrated promising in vivo antifungal activity to control infections of olives caused by Aspergillus niger and Penicillium crustosum. In addition, the essential oil exhibited fumigation toxicity against Bactrocera oleae with mortality percentage of 100% at 2 µL/L air. CONCLUSION: The results showed that essential oil of A. verticillata has interesting biological activities, which suggests that oil have the potential to be used as biocide to protect olives of pathogenic fungi and pests.

Postharvest treatment of mandarin fruit using a combination of methyl bromide and phosphine against Bactrocera dorsalis (Diptera: Tephritidae).

BACKGROUND: Chinese Nanfeng mandarin (Citrus reticulate Blanco '2-6') fruit is host to the oriental fruit fly Bactrocera dorsalis Hendel, and therefore requires phytosanitary treatment before exporting abroad. The use of methyl bromide (MB) should be reduced because it has phytotoxic and ozone depleting properties. In the present study, fumigations with mixtures of gaseous phosphine (PH3 ) and MB were conducted to disinfest B. dorsalis. The effect of combined fumigation on postharvest quality of Nanfeng mandarin fruit was then evaluated. RESULTS: There was a clear synergistic effect between MB and PH3 against B. dorsalis at 20 °C. The third-instar larvae of B. dorsalis were the most tolerant to treatment with both MB alone and combined with PH3 . Toxicity assay of third instars indicated that higher PH3 concentrations increased MB toxicity, and 1.42-2.84 g m-3 of PH3 was optimal when combined with MB. Probit analysis showed that, combined with 2.13 g m-3 PH3 , similar mortality was achieved with 50% of the MB required when applied alone. Furthermore, the fruit quality test showed that MB fumigation resulted in a higher respiration rate and >40% fruit rot after storage. The combined treatment reduced these effects and did not have adverse effects on fruit firmness, soluble solid content, titratable acidity, or vitamin C content. CONCLUSION: Our findings demonstrate the synergistic effect between PH3 and MB and indicate that this treatment has potential as a novel strategy for postharvest control of B. dorsalis, especially in MB-sensitive fruit. © 2019 Society of Chemical Industry.

Crustacean cardioactive peptide (CCAP) of the oriental fruit fly, Bactrocera dorsalis (Diptera: Tephritidae): Molecular characterization, distribution and its potential roles in larva-pupa ecdysis.

Insects must undergo ecdysis for successful development and growth, and the crustacean cardioactive peptide (CCAP) is one of the most important hormone in this process. Here we reported a cDNA encoding for the CCAP precursor cloned from the oriental fruit fly, Bactrocera dorsalis, a most destructive insect pest of agriculture. The CCAP mature peptide (PFCNAFTGC-NH2) of B. dorsalis was generated by post-translational processing and found to be highly comparable with other insects. RT-qPCR showed that mRNA of CCAP in B. dorsalis (BdCCAP) was predominantly expressed in the central nervous system (CNS) and midgut of 3rd-instar larvae. By using immunohistochemical analysis, we also localized the endocrine cells that produce CCAP in the CNS, ring gland and midgut of 3rd-instar larvae of B. dorsalis. The synthetic CCAP mature peptide could induce the expression of mRNA of adipokinetic hormone (AKH), the metabolic neuropeptides in insects. The expression of BdCCAP mRNA in the CNS, but not in the midgut, could be upregulated in the response to the challenge of insect molting hormone, 20-hydroxyecdysone.

Comparative Proteomic Profiling between Each of Two Consecutive Developmental Stages of the Solanum Fruit Fly, Bactrocera latifrons (Hendel).

The Solanum fruit fly, Bactrocera latifrons (Hendel), has a complex life cycle including multiple stages (egg, larva, pupa, and adult). Understanding the details of "what", "when", "where", "why", and "how" many hundred thousand proteins operate in this insect, interact, and express between each two consecutive developmental stages at molecular level not only can expand our knowledge, but also lead to the development of novel fruit fly control techniques. We tried to find what, when, and where in this study. Why and how will be presented in upcoming papers. We conducted a proteome profiling using 2-D gel electrophoresis and mass spectrometry. Samples of 3-day-old eggs, 1- and 10-day-old larvae, 1- and 10-day-old pupae, 1- and 9-day-old females and males of B. latifrons were used. A custom peptide database, derived from the de novo B. latifrons whole genome assembly was used for peptide identification. Differentially expressed proteins (DEPs) with significant fold expression and protein functions between two consecutive developmental stages were identified, annotated, described, and listed in gel images and/or charts. With this foundational information, we are not only providing valuable information, but also any impacts due to the biotic or abiotic environmental factors can be identified and manipulated, and lead to further research on gene editing and biomarker discovery.

Tyrosine hydroxylase coordinates larval-pupal tanning and immunity in oriental fruit fly (Bactrocera dorsalis).

BACKGROUND: The oriental fruit fly Bactrocera dorsalis (Hendel), a notorious world pest infesting fruits and vegetables, has evolved a high level of resistance to many commonly used insecticides. In this study, we investigate whether tyrosine hydroxylase (TH) that is required for cuticle tanning (sclerotization and pigmentation) in many insects, could be a potential target in controlling B. dorsalis. RESULTS: We cloned TH cDNA (BdTH) of B. dorsalis. The complete open reading frame of BdTH (KY911196) was 1737 bp in length, encoding a protein of 578 amino acids. Quantitative real-time PCR confirmed that BdTH was highly expressed in the epidermis of 3rd instar larvae, and its expression increased prior to pupation, suggesting a role in larval-pupal cuticle tanning. When we injected dsBdTH or 3-iodo-tyrosine (3-IT) as a TH inhibitor or fed insect diet supplemented with 3-IT, there was significant impairment of larval-pupal cuticle tanning and a severe obstacle to eclosion in adults followed by death in most. Furthermore, injection of Escherichia coli into larvae fed 3-IT resulted in 92% mortality and the expressions of four antimicrobial peptide genes were significantly downregulated. CONCLUSION: These results suggest that BdTH might play a critical role in larval-pupal tanning and immunity of B. dorsalis, and could be used as a potential novel target for pest control. © 2017 Society of Chemical Industry.

Two Gut-Associated Yeasts in a Tephritid Fruit Fly have Contrasting Effects on Adult Attraction and Larval Survival.

Yeast-insect interactions have been well characterized in drosophilid flies, but not in tephritid fruit flies, which include many highly polyphagous pest species that attack ripening fruits. Using the Queensland fruit fly (Bactrocera tryoni) as our model tephritid species, we identified yeast species present in the gut of wild-collected larvae and found two genera, Hanseniaspora and Pichia, were the dominant isolates. In behavioural trials using adult female B. tryoni, a fruit-agar substrate inoculated with Pichia kluyveri resulted in odour emissions that increased the attraction of flies, whereas inoculation with Hanseniaspora uvarum, produced odours that strongly deterred flies, and both yeasts led to decreased oviposition. Larval development trials showed that the fruit-agar substrate inoculated with the 'deterrent odour' yeast species, H. uvarum, resulted in significantly faster larval development and a greater number of adult flies, compared to a substrate inoculated with the 'attractive odour' yeast species, P. kluyveri, and a yeast free control substrate. GC-MS analysis of volatiles emitted by H. uvarum and P. kluyveri inoculated substrates revealed significant quantitative differences in ethyl-, isoamyl-, isobutyl-, and phenethyl- acetates, which may be responsible for the yeast-specific olfactory responses of adult flies. We discuss how our seemingly counterintuitive finding that female B. tryoni flies avoid a beneficial yeast fits well with our understanding of female choice of oviposition sites, and how the contrasting behavioural effects of H. uvarum and P. kluyveri raises interesting questions regarding the role of yeast-specific volatiles as cues to insect vectors. A better understanding of yeast-tephritid interactions could assist in the future management of tephritid fruit fly pests through the formulation of new "attract and kill" lures, and the development of probiotics for mass rearing of insects in sterile insect control programs.

Optimizing methyl-eugenol aromatherapy to maximize posttreatment effects to enhance mating competitiveness of male Bactrocera carambolae (Diptera: Tephritidae).

Methyl-eugenol (ME) (1,2-dimethoxy-4-(2-propenyl)benzene), a natural phytochemical, did enhance male Bactrocera carambolae Drew & Hancock (Diptera: Tephritidae) mating competitiveness 3 d after ingestion. Enhanced male mating competitiveness can significantly increase the effectiveness of the sterile insect technique (SIT). ME application to mass reared sterile flies by feeding is infeasible. ME application by aromatherapy however, would be a very practical way of ME application in fly emergence and release facilities. This approach was shown to enhance mating competitiveness of B. carambolae 3 d posttreatment (DPT). Despite this added benefit, every additional day of delaying release will reduce sterile fly quality and will add cost to SIT application. The present study was planned to assess the effects of ME-aromatherapy on male B. carambolae mating competitiveness 1DPT and 2DPT. ME aromatherapy 1DPT or 2DPT did enhance mating competitiveness of B. carambolae males whereas ME feeding 1DPT and 2DPT did not. Male mating competitiveness was enhanced by the ME aromatherapy irrespective if they received 1DPT, 2DPT or 3DPT. ME aromatherapy, being a viable approach for its application, did enhance mating competitiveness of male B. carambolae 1 d posttreatment as ME feeding did 3 d after ingestion.

Role of α-copaene in the susceptibility of olive fruits to Bactrocera oleae (Rossi).

The influence of α-copaene as a fruit volatile in the susceptibility of Olea europaea L. to the olive fruit fly Bactrocera oleae (Rossi) has been investigated. By studies on the relative area of volatile components from different cultivars, a positive correlation was found between the abundance of α-copaene in the samples and the corresponding degree of fruit infestation. SPME-GC-MS analysis of volatiles from uninfested fruits of O. europaea L. cv. Serrana were performed over two years to determine the variation of α-copaene throughout the different phenological stages. The results suggested that this sesquiterpene has a significant effect on cultivar susceptibility and may act as an oviposition promoter. Further analysis by chiral GC showed that olive fruits release both α-copaene enantiomers. Bioassays on each enantiomer revealed that fruits with increased amounts of (+)-α-copaene favor oviposition of B. oleae females, whereas the increase of (-)-α-copaene affords no statistically significant differences in host preference.

Methoprene application and diet protein supplementation to male melon fly, Bactrocera cucurbitae, modifies female remating behavior.

Methoprene (an analogue of juvenile hormone) application and feeding on a protein diet is known to enhance male melon fly, Bactrocera cucurbitae Coquillett (Diptera: Tephritidae), mating success. In this study, we investigated the effect of these treatments on male B. cucurbitae's ability to inhibit female remating. While 14-d-old females were fed on protein diet, 6-d-old males were exposed to one of the following treatments: (i) topical application of methoprene and fed on a protein diet; (ii) no methoprene but fed on a protein diet; (iii) methoprene and sugar-fed only; and (iv) sugar-fed, 14-d-old males acted as controls. Treatments had no effect on a male's ability to depress the female remating receptivity in comparison to the control. Females mated with protein-deprived males showed higher remating receptivity than females first mated with protein-fed males. Methoprene and protein diet interaction had a positive effect on male mating success during the first and second mating of females. Significantly more females first mated with sugar-fed males remated with protein-fed males and females first mated with methoprene treated and protein-fed males were more likely to remate with similarly treated males. Females mating latency (time to start mating) was significantly shorter with protein-fed males, and mating duration was significantly longer with protein-fed males compared with protein-deprived males. These results are discussed in the context of methoprene and/or dietary protein as prerelease treatment of sterile males in area-wide control of melon fly integrating the sterile insect technique (SIT).