On the impact of re-mating and residual fertility on the Sterile Insect Technique efficacy: Case study with the medfly, Ceratitis capitata.

The sterile insect technique (SIT) can be an efficient solution for reducing or eliminating certain insect pest populations. It is widely used in agriculture against fruit flies, including the Mediterranean fruit fly (medfly), Ceratitis capitata. The re-mating tendency of medfly females and the fact that the released sterile males may have some residual fertility could be a challenge for the successful implementation of the SIT. Obtaining the right balance between sterility level and sterile male quality (competitiveness, longevity, etc) is the key to a cost-efficient program. Since field experimental approaches can be impacted by many environmental variables, it is difficult to get a clear understanding on how specific parameters, alone or in combination, may affect the SIT efficiency. The use of models not only helps to gather knowledge, but it allows the simulation of a wide range of scenarios and can be easily adapted to local populations and sterile male production. In this study, we consider single- and double-mated females. We first show that SIT can be successful only if the residual fertility is less than a threshold value that depends on the basic offspring number of the targeted pest population, the re-mating rates, and the parameters of double-mated females. Then, we show how the sterile male release rate is affected by the parameters of double-mated females and the male residual fertility. Different scenarios are explored with continuous and periodic sterile male releases, with and without ginger aromatherapy, which is known to enhance sterile male competitiveness, and also taking into account some biological parameters related to females that have been mated twice, either first by a wild (sterile) male and then a sterile (wild) male, or by two wild males only. Parameter values were chosen for peach as host fruit to reflect what could be expected in the Corsican context, where SIT against the medfly is under consideration. Our results suggest that ginger aromatherapy can be a decisive factor determining the success of SIT against medfly. We also emphasize the importance of estimating the duration of the refractory period between matings depending on whether a wild female has mated with a wild or sterile male. Further, we show the importance of parameters, like the (hatched) eggs deposit rate and the death-rate related to all fertile double-mated females. In general, re-mating is considered to be detrimental to SIT programs. However, our results show that, depending on the parameter values of double-mated females, re-mating may also be beneficial for SIT. Our model can be easily adapted to different contexts and species, for a broader understanding of release strategies and management options.

The maxillary palps of Tephritidae are selectively tuned to food volatiles and diverge with ecology.

The maxillary palp is an auxiliary olfactory organ in insects, which, different from the antennae, is equipped with only a few olfactory sensory neuron (OSN) types. We postulated that these derived mouthpart structures, positioned at the base of the proboscis, may be particularly important in mediating feeding behaviors. As feeding is spatio-temporally segregated from oviposition in most Tephritidae, this taxonomic group appears quite suitable to parse out sensory breadth and potential functional divergence of palps and antennae. Scanning electron microscopy and anterograde staining underlined the limited palpal olfactory circuit in Tephritidae: only three morphological subtypes of basiconic sensilla were found, each with two neurons, and project to a total of six antennal lobe glomeruli in Bactrocera dorsalis. Accordingly, the palps detected only few volatiles from the headspace of food (fermentation and protein lures) and fruit (guava and mango) compared to the antennae (17 over 77, using gas-chromatography coupled electrophysiology). Interestingly, functionally the antennae were more tuned to fruit volatiles, detecting eight times more fruit than food volatiles (63 over 8), whereas the number of fruit and food volatile detection was more comparable in the palps (14 over 8). As tephritids diverge in oviposition preferences, but converge on food substrates, we postulated that the receptive ranges of palpal circuits would be more conserved compared to the antennae. However, palpal responses of three tephritid species that differed in phylogenetic relatedness and ecologically niche, diverged across ecological rather than phylogenetic rifts. Two species with strongly overlapping ecology, B. dorsalis and Ceratitis capitata, showed inseparable response profiles, whereas the cucurbit specialist Zeugodacus cucurbitae strongly diverged. As Z. cucurbitae is phylogenetically placed between B. dorsalis and C. capitata, the results indicate that ecology overrides phylogeny in the evolution of palpal tuning, in spite of being predisposed to detecting food volatiles.

Mediterranean fruit fly population phenological patterns are strongly affected by elevation and host presence.

The Mediterranean fruit fly (medfly) (Ceratitis capitata, Diptera: Tephritidae), is an extremely polyphagous pest that threatens the fruit production and trading industry worldwide. Monitoring C. capitata populations and analysing its dynamics and phenology is considered of outmost importance for designing and implementing sound management approaches. The aim of this study was to investigate the factors regulating the population dynamics of the C. capitata in a coastal and semi-mountainous area. We focused on effects of topography (e.g. elevation), host presence and seasonal patterns of ripening on the phenological patterns considering data collected in 2008. The experimental area is characterized by mixed fruit orchards, and Mediterranean climate with mild winters. Two trap types were used for population monitoring. The female targeted McPhail type and the male targeted Jackson type. Traps were placed in farms located at different elevations and landscape morphology (coastal and semi-mountainous areas). The main crops included citrus, apples, peaches, plums, pears, figs, quinces and apricots. Adult captures were first recorded in May, peaked in mid-summer and mid-autumn and almost ceased at the end of the season (January 2008). Captures in the coastal areas preceded that of highlands by 15 days. Most of the adults detected during the fruit ripening of late stone fruit cultivars (first peak) and citrus (second peak). The probability of capturing the first adults preceded almost three weeks the peak of adult captures either considering the elevation or host focus analyses. The results provide valuable information on the seasonal population trend of C. capitata in mixed fruit Mediterranean orchards and can support the set-up of IPM systems in areas with various landscapes and different hosts throughout the fruit growing season.

Deep orange gene editing triggers temperature-sensitive lethal phenotypes in Ceratitis capitata.

BACKGROUND: The Mediterranean fruit fly, Ceratitis capitata, is a significant agricultural pest managed through area-wide integrated pest management (AW-IPM) including a sterile insect technique (SIT) component. Male-only releases increase the efficiency and cost-effectiveness of SIT programs, which can be achieved through the development of genetic sexing strains (GSS). The most successful GSS developed to date is the C. capitata VIENNA 8 GSS, constructed using classical genetic approaches and an irradiation-induced translocation with two selectable markers: the white pupae (wp) and temperature-sensitive lethal (tsl) genes. However, currently used methods for selecting suitable markers and inducing translocations are stochastic and non-specific, resulting in a laborious and time-consuming process. Recent efforts have focused on identifying the gene(s) and the causal mutation(s) for suitable phenotypes, such as wp and tsl, which could be used as selectable markers for developing a generic approach for constructing GSS. The wp gene was recently identified, and efforts have been initiated to identify the tsl gene. This study investigates Ceratitis capitata deep orange (Ccdor) as a tsl candidate gene and its potential to induce tsl phenotypes. RESULTS: An integrated approach based on cytogenetics, genomics, bioinformatics, and gene editing was used to characterize the Ccdor. Its location was confirmed on the right arm of chromosome 5 in the putative tsl genomic region. Knock-out of Ccdor using CRISPR/Cas9-NHEJ and targeting the fourth exon resulted in lethality at mid- and late-pupal stage, while the successful application of CRISPR HDR introducing a point mutation on the sixth exon resulted in the establishment of the desired strain and two additional strains (dor 12del and dor 51dup), all of them expressing tsl phenotypes and presenting no (or minimal) fitness cost when reared at 25 °C. One of the strains exhibited complete lethality when embryos were exposed at 36 °C. CONCLUSIONS: Gene editing of the deep orange gene in Ceratitis capitata resulted in the establishment of temperature-sensitive lethal mutant strains. The induced mutations did not significantly affect the rearing efficiency of the strains. As deep orange is a highly conserved gene, these data suggest that it can be considered a target for the development of tsl mutations which could potentially be used to develop novel genetic sexing strains in insect pests and disease vectors.

Survey on Drosophila suzukii and Ceratitis capitata (Diptera: Drosophilidae, Tephritidae) and Associated Eucoilinae Species (Hymenoptera: Figitidae) in Northwestern Argentina. First Record of Dicerataspis grenadensis and Leptopilina boulardi as Parasitoids of D. suzukii.

The Southeast Asian-native Drosophila suzukii (Matsumura) (Diptera: Drosophilidae), also known as "spotted-wing Drosophila," is one of the most globally invasive agricultural species. Although D. suzukii is a pest spread throughout all the Argentinian fruit-growing regions, few information has been published on its impact on local fruit production. Parasitoid species associated with D. suzukii in Argentina belong to Pteromalidae (Chalcidoidea), Diapriidae (Diaprioidea), both attacking host pupae, and Figitidae (Cynipoidea), which attack host larvae. Nine Eucoilinae (Figitidae) species, belonging to Dicerataspis, Dieucoila, Euxestophaga, Ganaspis, Hexacola, and Leptopilina genera, have been associated with D. suzukii in Argentina. Ceratitis capitata (Wiedemann), commonly known as "medfly," is native to Africa and has a worldwide distribution, covering many tropical, subtropical, and temperate regions. In Argentina, C. capitata has been associated with several native hymenopterous parasitoids belonging to Braconidae (Ichneumonioidea), Eulophidae (Chalcidoidea), Pteromalidae, Diapriidae, and Figitidae families. Only two eucoline species, Ganaspis pelleranoi (Brèthes) and Leptopilina haywardi (Blanchard) have been related to medfly in Argentina. We report new trophic associations between the parasitoids Dicerataspis grenadensis Ashmead and Leptopilina boulardi (Barbotin, Carton and Kelner-Pillault) and D. suzukii, and between the parasitoid Odontosema albinerve Kieffer and C. capitata, after surveys conducted in Tucumán, northwestern Argentina. An annotated checklist and a taxonomic key of Eucoilinae associated with both invasive pests, in Argentina, are also provided.

Gene drive and genetic sex conversion in the global agricultural pest Ceratitis capitata.

Homing-based gene drives are recently proposed interventions promising the area-wide, species-specific genetic control of harmful insect populations. Here we characterise a first set of gene drives in a tephritid agricultural pest species, the Mediterranean fruit fly Ceratitis capitata (medfly). Our results show that the medfly is highly amenable to homing-based gene drive strategies. By targeting the medfly transformer gene, we also demonstrate how CRISPR-Cas9 gene drive can be coupled to sex conversion, whereby genetic females are transformed into fertile and harmless XX males. Given this unique malleability of sex determination, we modelled gene drive interventions that couple sex conversion and female sterility and found that such approaches could be effective and tolerant of resistant allele selection in the target population. Our results open the door for developing gene drive strains for the population suppression of the medfly and related tephritid pests by co-targeting female reproduction and shifting the reproductive sex ratio towards males. They demonstrate the untapped potential for gene drives to tackle agricultural pests in an environmentally friendly and economical way.

Latitudinal variation in survival and immature development of Ceratitis capitata populations reared in two key overwintering hosts.

Ceratitis capitata, a major agricultural pest, is currently expanding its geographic distribution to northern, temperate areas of Europe. Its seasonal biology and invasion success depend on temperature, humidity and host availability. In coastal warmer Mediterranean regions and cooler temperature areas, bitter oranges and apples serve as overwintering hosts during the larval stage. We assessed the overwintering capacity of C. capitata populations obtained from different areas of the northern hemisphere by studying the survival and development rates of immature stages in both fruits under laboratory conditions. Eggs from each population were artificially inserted in the flesh of the two hosts and kept at 15, 20, or 25 °C until pupation and adult emergence. Climatic analysis of the area of the population origin showed combined effects of latitude, host and macroclimatic variables on immature survival and development rates. Egg to adult survival rates and developmental duration were longer in apples than in bitter oranges. For populations originated from southern-warmer areas, egg to adult developmental duration was prolonged and adult emergence reduced at 15 °C compared to those populations obtained from northern regions. Our findings reveal varying plastic responses of medfly populations to different overwintering hosts and temperatures highlighting the differential overwintering potential as larvae within fruits. This study contributes towards better understanding the medfly invasion dynamics in temperate areas of Northern Europe and other parts of the globe with similar climates.

Probiotic consortium modulating the gut microbiota composition and function of sterile Mediterranean fruit flies.

The sterile insect technique (SIT) remains a successful approach in managing pest insects. However, the long-term mass rearing and sterilizing radiation associated with SIT have been observed to induce physiological and ecological fitness decline in target insects. This decline may be attributed to various factors, including commensal microbiota dysbiosis, selection procedures, loss of heterozygosity, and other complex interactions.. There is evidence that the bacterial symbiont of insects may play critical roles in digestion, development, reproduction, and behavior. Probiotics are an increasingly common approach for restoring the intestinal microbiota structure and fitness parameters of sterile insects, particularly in the Vienna 8 genetic sexing strain (V8-GSS) of the Mediterranean fruit fly (medfly), Ceratitis capitata. Here, we explore the influence of the previously isolated bacterial strain, Lactococcus lactis, Enterobacter sp., and Klebsiella oxytoca, administration as probiotic consortia (LEK-PC) to the larvae and/or adult diet over the course of 20 rearing generations on fitness parameters. The experiment was carried out in four colonies: a control colony (C), one to which probiotics were not added, one to which probiotics were added to the larval medium (L+), one to which probiotics were added to the adult medium (A+), and one to which probiotics were added to both the larval and adult mediums (AL+). Emergence, flight ability, survival under stress conditions, and mating competitiveness, were all significantly improved by the LEK-PC treatment independently of the administration stage. The intestinal microbiota structure of various medfly V8-GSS colonies also underwent a significant shift, despite the fact that the core microbial community was unaffected by the LEK-PC administration stage, according to 16S metagenomics sequencing. Comparison of the metabolic function prediction and associated carbohydrate enzymes among colonies treated with "LEK-PC" showed an enrichment of metabolic functions related to carbohydrates, amino acids, cofactors, and vitamins metabolism, as well as, glycoside hydrolase enzymes in the AL+ colony compared to the control. This study enriches the knowledge regarding the benefits of probiotic treatment to modulate and restore the intestinal microbiota of C. capitata sterile males for a better effectiveness of the SIT.

Evidence that recent climatic changes have expanded the potential geographical range of the Mediterranean fruit fly.

The species distributions migration poleward and into higher altitudes in a warming climate is especially concerning for economically important insect pest species, as their introduction can potentially occur in places previously considered unsuitable for year-round survival. We explore the expansion of the climatically suitable areas for a horticultural pest, the Mediterranean fruit fly (medfly) Ceratitis capitata (Diptera, Tephritidae), with an emphasis on Europe and California. We reviewed and refined a published CLIMEX model for C. capitata, taking into consideration new records in marginal locations, with a particular focus on Europe. To assess the model fit and to aid in interpreting the meaning of the new European distribution records, we used a time series climate dataset to explore the temporal patterns of climate suitability for C. capitata from 1970 to 2019. At selected bellwether sites in Europe, we found statistically significant trends in increasing climate suitability, as well as a substantial northward expansion in the modelled potential range. In California, we also found a significant trend of northward and altitudinal expansion of areas suitable for C. capitata establishment. These results provide further evidence of climate change impacts on species distributions and the need for innovative responses to increased invasion threats.

An 18-day, 3 °C cold treatment effectively kills Ceratitis capitata (Diptera: Tephritidae) in kiwifruit (Actinidia spp.).

A series of experiments were carried out to develop a phytosanitary disinfestation protocol to kill Ceratitis capitata (Weidemann) (Mediterranean fruit fly, Diptera: Tephritidae) in 'Hayward' kiwifruit (Actinidia deliciosa (A. Chev.) C.F. Liang and A.R. Ferguson) and 'Zesy002' kiwifruit (Actinidia chinensis Planch.). Experiments on 4 immature life stages (eggs and 3 larval instars) with treatment durations of between 5 and 18 days showed that third instars were the most tolerant to temperatures around 3 °C, with the lethal time to 99.9968% (probit 9) mortality (LT99.9968) estimated to be 17.3 days (95% confidence interval (CI) 16.4-18.2). Larvae reared and treated in 'Zesy002' were significantly more susceptible to cold treatment than those reared in 'Hayward'. A large-scale trial testing a disinfestation protocol of 3 ±â€…0.5 °C for 18 days treated over 500,000 third-instar C. capitata with no survivors. These results demonstrate that a cold treatment of 3.5 °C or below for 18 days induces C. capitata mortality in kiwifruit at a rate that exceeds 99.9968% with a degree of confidence greater than 99%.

Daily activity profiles over the lifespan of female medflies as biomarkers of aging and longevity.

The relationship between the early-age activity of Mediterranean fruit flies (medflies) or other fruit flies and their lifespan has not been much studied, in contrast to the connections between lifespan and diet, sexual signaling, and reproduction. The objective of this study is to assess intra-day and day-to-day activity profiles of female Mediterranean fruit flies and their role as biomarker of longevity as well as to explore the relationships between these activity profiles, diet, and age-at-death throughout the lifespan. We use advanced statistical methods from functional data analysis (FDA). Three distinct patterns of activity variations in early-age activity profiles can be distinguished. A low-caloric diet is associated with a delayed activity peak, while a high-caloric diet is linked with an earlier activity peak. We find that age-at-death of individual medflies is connected to their activity profiles in early life. An increased risk of mortality is associated with increased activity in early age, as well as with a higher contrast between daytime and nighttime activity. Conversely, medflies are more likely to have a longer lifespan when they are fed a medium-caloric diet and when their daily activity is more evenly distributed across the early-age span and between daytime and nighttime. The before-death activity profile of medflies displays two characteristic before-death patterns, where one pattern is characterized by slowly declining daily activity and the other by a sudden decline in activity that is followed by death.

RNAi-mediated silencing of Mediterranean fruit fly (Ceratitis capitata) endogenous genes using orally-supplied double-stranded RNAs produced in Escherichia coli.

BACKGROUND: The Mediterranean fruit fly (medfly), Ceratitis capitata Wiedemann, is a major pest affecting fruit and vegetable production worldwide, whose control is mainly based on insecticides. Double-stranded RNA (dsRNA) able to down-regulate endogenous genes, thus affecting essential vital functions via RNA interference (RNAi) in pests and pathogens, is envisioned as a more specific and environmentally-friendly alternative to traditional insecticides. However, this strategy has not been explored in medfly yet. RESULTS: Here, we screened seven candidate target genes by injecting in adult medflies gene-specific dsRNA hairpins transcribed in vitro. Several genes were significantly down-regulated, resulting in increased insect mortality compared to flies treated with a control dsRNA targeting the green fluorescent protein (GFP) complementary DNA (cDNA). Three of the dsRNAs, homologous to the beta subunit of adenosine triphosphate (ATP) synthase (ATPsynbeta), a vacuolar ATPase (V-ATPase), and the ribosomal protein S13 (RPS13), were able to halve the probability of survival in only 48 h after injection. We then produced new versions of these three dsRNAs and that of the GFP control as circular molecules in Escherichia coli using a two-self-splicing-intron-based expression system and tested them as orally-delivered insecticidal compounds against medfly adults. We observed a significant down-regulation of V-ATPase and RPS13 messenger RNAs (mRNAs) (approximately 30% and 90%, respectively) compared with the control medflies after 3 days of treatment. No significant mortality was recorded in medflies, but egg laying and hatching reduction was achieved by silencing V-ATPase and RPS13. CONCLUSION: In sum, we report the potential of dsRNA molecules as oral insecticide in medfly. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Automated image-based analysis unveils acute effects due to sub-lethal pesticide doses exposure.

Pesticides are still abused in modern agriculture. The effects of their exposure to even sub-lethal doses can be detrimental to ecosystem stability and human health. This work aims to validate the use of machine learning techniques for recognizing motor abnormalities and to assess any effect post-exposure to a minimal dosage of these substances on a model organism, gaining insights into potential risks for human health. The test subject was the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae), exposed to food contaminated with the LC30 of Carlina acaulis essential oil. A deep learning approach enabled the pose estimation within an arena. Statistical analysis highlighted the most significant features between treated and untreated groups. Based on this analysis, two learning-based algorithms, Random Forest (RF) and XGBoost were employed. The results were compared through different metrics. RF algorithm generated a model capable of distinguishing treated subjects with an area under the receiver operating characteristic curve of 0.75 and an accuracy of 0.71. Through an image-based analysis, this study revealed acute effects due to minimal pesticide doses. So, even small amounts of these biocides drifted far from distribution areas may negatively affect the environment and humans.

Ecotone effect on the fruit fly assemblages (Diptera: Tephritidae) in natural and anthropized environments.

Various species of fruit flies are important pests of fruit cultures and in some crop of vegetables worldwide. Studies of their population patterns, ecological processes and mechanisms that influence their sampling and distribution in the ecosystems, provides important information to support researches on species diversity and ecologically based pest control programs. The aims of this paper were to analyze the patterns of fruit fly species: diversity, abundance and composition in the margin and inside of a fragment of native forest (35 ha); on the margin and in the inside a commercial orchard (2.5 ha). This research was carried out in transects in a fragment of semideciduous forest and in a commercial orchard in the region of Dourados, Mato Grosso do Sul, Brazil. A total of 1,918 adult fruit flies: 1,350 Ceratitis capitata (Wiedemann) (♂♂+♀♀) and 568 of the genus Anastrepha Schiner (275♂♂ + 293♀♀) from six infrageneric groups and 12 different species were captured. C. capitata (80%), and Anastrepha sororcula Zucchi were the most abundant species, being this last one representing 50.68% of individuals in the genus Anastrepha. There are significant differences in species diversity of fruit flies caught in the traps installed in the edge and inside of both environments: traps installed in the forest edge had higher diversity index (H' = 2.13) in compare to the inside of forest (H' = 1.67), with the same pattern repeated in the orchard: edge (H' = 0.55) and inside (H' = 0.41). The results in this paper corroborate with the prediction that in ecotonal areas between environments there are higher diversity in compare with the inside of each of the confronting ecosystems. The technique proposed here saves time, effort and resources in rapid inventories for sampling fruit fly species richness in natural forests and large fruit tree orchards.

The use of toxic baits for the suppression of Mediterranean fruit fly in mango orchards.

Ceratitis capitata (Wiedemann, 1824) is an insect of major economic importance in the mango orchards of the submedium of the São Francisco River Valley, the main area of mango production and exportation in Brazil. To provide alternatives for the management of C. capitata, toxic baits based on alpha-cypermethrin (Gelsura®) and spinosad (Success® 0.02 CB) were evaluated in three commercial mango experiments during two consecutive harvests: 2016/2017 (experiment 1 - area 1) and 2017/2018 (experiment 2 -area 2 and experiment 3 - area 3). According to the results, there was a large reduction in the infestation of C. capitata after five sequential applications of the alpha-cypermethrin (6 g.ha-1) and spinosad (0.38 g.ha-1) toxic baits performed at seven-day intervals during mango fruit ripening in all experiments and years (harvest) evaluated. Compared with the untreated plots, the plots with alpha-cypermethrin and spinosad applications showed a significant reduction in the damage induced (fallen fruits and/or on trees) by C. capitata. The management of C. capitata in mango orchards can include the use of the toxic bait based on alpha-cypermethrin, which represents an alternative to rotate with spinosad toxic bait in the São Francisco River Valley.

Interference of tritrophic (grape × medfly × parasitoid) interactions by mineral and biomaterial films.

Fruit fly infestation is one of the main obstacles to the exportation of fresh agricultural produce. Films of mineral particles and biomaterials have the potential to protect fruits against tephritid fruit fly infestation. The present study evaluated the effects of particle films on the tritrophic interactions of grape (Vitis vinifera L.), the fruit fly Ceratitis capitata (Wiedemann) and the parasitoid Diachasmimorpha longicaudata (Ashmead) under semi-field conditions. Grapes were biometrically characterised (i.e. colour, firmness, mass, length and diameter), treated with mineral particles, biomaterials or distilled water (control), and then used in oviposition and parasitism bioassays. In the oviposition bioassay, the treated grapes were exposed to 50 C. capitata pairs in field cages, and after 48 h, the punctures and eggs on each fruit were counted. In the parasitism bioassay, treated grapes were artificially infested with third-instar C. capitata larvae (two per fruit), exposed (2 h) to 50 D. longicaudata pairs in field cages to determine parasitism index, larval and pupal viabilities and number of flies and parasitoids emerged. Treatment with the mineral film affected fruit colour and reduced C. capitata oviposition but failed to significantly affect the parasitism capacity of D. longicaudata. The ability of the parasitoid to locate and parasitise C. capitata larvae in kaolin-coated fruits suggests that kaolin films could be used in conjunction with biological agents to control fruit flies.

Next-generation genetic sexing strain establishment in the agricultural pest Ceratitis capitata.

Tephritid fruit fly pests pose an increasing threat to the agricultural industry due to their global dispersion and a highly invasive nature. Here we showcase the feasibility of an early-detection SEPARATOR sex sorting approach through using the non-model Tephritid pest, Ceratitis capitata. This system relies on female-only fluorescent marker expression, accomplished through the use of a sex-specific intron of the highly-conserved transformer gene from C. capitata and Anastrepha ludens. The herein characterized strains have 100% desired phenotype outcomes, allowing accurate male-female separation during early development. Overall, we describe an antibiotic and temperature-independent sex-sorting system in C. capitata, which, moving forward, may be implemented in other non-model Tephritid pest species. This strategy can facilitate the establishment of genetic sexing systems with endogenous elements exclusively, which, on a wider scale, can improve pest population control strategies like sterile insect technique.

Development of a novel genetic sexing strain of Ceratitis capitata based on an X-autosome translocation.

Genetic sexing strains (GSS), such as the Ceratitis capitata (medfly) VIENNA 8 strain, facilitate male-only releases and improve the efficiency and cost-effectiveness of sterile insect technique (SIT) applications. Laboratory domestication may reduce their genetic diversity and mating behaviour and hence, refreshment with wild genetic material is frequently needed. As wild males do not carry the T(Y;A) translocation, and wild females do not easily conform to artificial oviposition, the genetic refreshment of this GSS is a challenging and time-consuming process. In the present study, we report the development of a novel medfly GSS, which is based on a viable homozygous T(XX;AA) translocation using the same selectable markers, the white pupae and temperature-sensitive lethal genes. This allows the en masse cross of T(XX;AA) females with wild males, and the backcrossing of F1 males with the T(XX;AA) females thus facilitating the re-establishment of the GSS as well as its genetic refreshment. The rearing efficiency and mating competitiveness of the novel GSS are similar to those of the T(Y;A)-based VIENNA 8 GSS. However, its advantage to easily allow the genetic refreshment is of great importance as it can ensure the mass production of high-quality males and enhanced efficacy of operational SIT programs.

Differential responses of Ceratitis capitata to infection by the entomopathogenic fungus Purpureocillium lilacinum.

The medfly Ceratitis capitata is one of the most damaging fruit pests with quarantine significance due to its extremely wide host range. The use of entomopathogenic fungi constitutes a promising approach with potential applications in integrated pest management. Furthermore, developing insect control methods can involve the use of fungal machinery to cause metabolic disruption, which may increase its effectiveness by impairing insect development. Insect species, including C. capitata, relies on reproduction potential, nutrient reserves, metabolic activities, and immune response for survival. Accordingly, the purpose of this study was to investigate the impacts of the entomopathogenic fungus Purpureocillium lilacinum on C. capitata pre-mortality. The medfly V8 strain was subjected to laboratory bioassays, which consisted on determining the virulence of P. lilacinum on the medfly. Purpureocillium lilacinum was applied on abdominal topical of 5-day-old males and females. Following the fungal inoculation, we have confirmed (i) a significant increase in tissue sugar content, (ii) a significant decrease in carbohydrase activities, digestive glycosyl hydrolase, and proteinase activities in whole midguts of treated flies, (iii) the antimicrobial peptides (AMPs) genes expression profile was significantly influenced by fly gender, fly status (virgin, mature, and mated), and time after infection, but infection itself had no discernible impact on the AMPs for the genes that were examined. This study provides the first insight into how P. lilacinum could affect C. capitata physiological mechanisms and provides the foundation for considering P. lilacinum as a novel, promising biocontrol agent.

Natural Oil Lure Outperforms Trimedlure in Capturing Males of the Mediterranean Fruit Fly, Ceratitis capitata (Diptera: Tephritidae).

Females of certain tephritid fruit fly species (Diptera: Tephritidae) pose an enormous agricultural threat, as they oviposit in commercially important fruits and vegetables. Trapping networks are often operated in fruit fly-free areas to detect incipient infestations. Trapping relies largely on male attractants, so-called male lures, with trimedlure (TML) being used to detect invasive Ceratitis spp. Operating large-scale surveillance programs incurs substantial costs for both supplies and labor, and the problem is exacerbated by the fact that trimedlure (as well as other male lures) is effective for relatively short intervals in the field (6-8 weeks). Because frequent servicing increases costs, there is considerable interest in modifying existing lures or developing new formulations to extend their effective field longevity. Here, we present results of a field study in Hawaii on a wild population of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann), that compared male captures in traps baited with (i) fresh liquid TML, (ii) TML plugs, (iii) a novel controlled-release TML sachet, and (iv) a novel natural oil blend dispensed from a sachet. Catch was recorded weekly for 12 weeks and then at 16 and 20 weeks, with 12 traps deployed per treatment. The natural oil formulation, which contains the natural plant product α-copaene, was as effective as the fresh liquid TML even after weathering for 20 weeks. Future work will focus on developing a dispenser for this formulation that is compatible with standard trap design and deployment.