Two heat shock cognate 70 genes involved in spermatogenesis regulate the male fertility of Zeugodacus cucurbitae, as potential targets for pest control.

The melon fly Zeugodacus cucurbitae Coquillett (Diptera: Tephritidae) is an agricultural quarantine pest threatening fruit and vegetable production. Heat shock cognate 70 (Hsc70), which is a homolog of the heat shock protein 70 (Hsp70), was first discovered in mice testes and plays an important role in spermatogenesis. In this study, we identified and cloned five Hsc70 genes from melon fly, namely ZcHsc70_1/2/3/4/5. Phylogenetic analysis showed that these proteins are closely related to Hsc70s from other Diptera insects. Spatiotemporal expression analysis showed that ZcHsc70_1 and ZcHsc70_2 are highly expressed in Z. cucurbitae testes. Fluorescence in situ hybridization further demonstrated that ZcHsc70_1 and ZcHsc70_2 are expressed in the transformation and maturation regions of testes, respectively. Moreover, RNA interference-based suppression of ZcHsc70_1 or ZcHsc70_2 resulted in a significant decrease of 74.61% and 63.28% in egg hatchability, respectively. Suppression of ZcHsc70_1 expression delayed the transformation of sperm cells to mature sperms. Meanwhile, suppression of ZcHsc70_2 expression decreased both sperm cells and mature sperms by inhibiting the meiosis of spermatocytes. Our findings show that ZcHsc70_1/2 regulates spermatogenesis and further affects the male fertility in the melon fly, showing potential as targets for pest control in sterile insect technique by genetic manipulation of males.

Mixed insect pest populations of Diaspididae species under control of oligonucleotide insecticides: 3'-end nucleotide matters.

Diaspididae are one of the most serious small herbivorous insects with piercing-sucking mouth parts and are major economic pests as they attack and destroy perennial ornamentals and food crops. Chemical control is the primary management approach for armored scale infestation. However, chemical insecticides do not possess selectivity in action and not always effective enough for the control of armored scale insects. Our previous work showed that green oligonucleotide insecticides (olinscides) are highly effective against armored and soft scale insects. Moreover, olinscides possess affordability, selectivity in action, fast biodegradability, and a low carbon footprint. Insect pest populations undergo microevolution and olinscides should take into account the problem of insecticide resistance. Using sequencing results, it was found that in the mixed populations of insect pests Dynaspidiotus britannicus Newstead and Aonidia lauri Bouche, predominates the population of A. lauri. Individuals of A. lauri comprised for 80% of individuals with the sequence 3'-ATC-GTT-GGC-AT-5' in the 28S rRNA site, and 20% of the population comprised D. britannicus individuals with the sequence 3'-ATC-GTC-GGT-AT-5'. We created olinscides Diasp80-11 (5'-ATG-CCA-ACG-AT-3') and Diasp20-11 (5'-ATA-CCG-ACG-AT-3') with perfect complementarity to each of the sequences. Mortality of insects on the 14th day comprised 98.19 ± 3.12% in Diasp80-11 group, 64.66 ± 0.67% in Diasp20-11 group (p < 0.05), and 3.77 ± 0.94% in the control group. Results indicate that for maximum insecticidal effect it is necessary to use an oligonucleotide insecticide that corresponds to the dominant species. Mortality in Diasp80-11 group was accompanied with significant decrease in target 28S rRNA concentration and was 8.44 ± 0.14 and 1.72 ± 0.36 times lower in comparison with control (p < 0.05) on the 10th and 14th days, respectively. We decided to make single nucleotide substitutions in Diasp20-11 olinscide to understand which nucleotide will play the most important role in insecticidal effect. We created three sequences with single nucleotide transversion substitutions at the 5'-end - Diasp20(5')-11 (A to T), 3'-end - Diasp20(3')-11 (T to A), and in the middle of the sequence - Diasp20(6)-11 (6th nitrogenous base of the sequence; G to C), respectively. As a result, mortality of mixed population of the field experiment decreased and comprised 53.89 ± 7.25% in Diasp20(5')-11 group, 40.68 ± 4.33% in Diasp20(6)-11 group, 35.74 ± 5.51% in Diasp20(3')-11 group, and 3.77 ± 0.94% in the control group on the 14th day. Thus, complementarity of the 3'-end nucleotide to target 28S rRNA was the most important for pronounced insecticidal effect (significance of complementarity of nucleotides for insecticidal effect: 5' nt < 6 nt < 3' nt). As was found in our previous research works, the most important rule to obtain maximum insecticidal effect is complete complementarity to the target rRNA sequence and maximum coverage of target sequence in insect pest populations. However, in this article we also show that the complementarity of 3'-end is a second important factor for insecticidal potential of olinscides. Also in this article we propose 2-step DNA containment mechanism of action of olinscides, recruiting RNase H. The data obtained indicate the selectivity of olinscides and at the same time provide a simple and flexible platform for the creation of effective plant protection products, based on antisense DNA oligonucleotides.

Less is more: Fewer attract-and-kill sites improve the male annihilation technique against Bactrocera dorsalis (Diptera: Tephritidae).

The Male Annihilation Technique (also termed the Male Attraction Technique; "MAT") is often used to eradicate pestiferous tephritid fruit flies, such as Bactrocera dorsalis (Hendel). MAT involves the application of male-specific attractants combined with an insecticide in spots or stations across an area to reduce the male population to such a low level that suppression or eradication is achieved. Currently, implementations of MAT in California and Florida targeting B. dorsalis utilize the male attractant methyl eugenol (ME) accompanied with a toxicant, such as spinosad, mixed into a waxy, inert emulsion STATIC ME (termed here "SPLAT-MAT-ME"). While highly effective against ME-responding species, such applications are expensive owing largely to the high cost of the carrier matrix and labor for application. Until recently the accepted protocol called for the application of approximately 230 SPLAT-MAT-ME spots per km2; however, findings from Hawaii suggest a lower density may be more effective. The present study adopted the methods of that earlier work and estimated kill rates of released B. dorsalis under varying spot densities in areas of California and Florida that have had recent incursions of this invasive species. Specifically, we directly compared trap captures of sterilized marked B. dorsalis males released in different plots under three experimental SPLAT-MAT-ME densities (50, 110, and 230 per km2) in Huntington Beach, CA; Anaheim, CA; and Sarasota-Bradenton, FL. The plots with a density of 110 sites per km2 had a significantly higher recapture proportion than plots with 50 or 230 sites per km2. This result suggests that large amounts of male attractant may reduce the ability of males to locate the source of the odor, thus lowering kill rates and the effectiveness of eradication efforts. Eradication programs would directly benefit from reduced costs and improved eradication effectiveness by reducing the application density of SPLAT-MAT-ME.

The Economic Analyses of the Drosophila suzukii's Invasions: A Mini-review.

The objective of this paper is to bring to the fore the type of economic analyses that have been carried out on the invasion of the Drosophila suzukii Matsumura (Diptera: Drosophilidae) and what has been learned. The analysis is limited to the original research articles published in peer-reviewed journals and book chapters. A total of 20 relevant studies are identified. The studies are analysed based on their main purpose, which is either economic impact assessment or economic evaluation of management programmes. The analysis also considers the key methodological points highlighted in recent reviews of the economic literature on alien invasive species. Over time, the focus of these studies has gradually shifted from raising awareness of the magnitude of the impact, particularly on agricultural production, to supporting the decision-making process concerning effective pest management. Most studies have been conducted from a private perspective, measuring private costs and providing guidance to the industry. However, some papers include, or focus on, the societal costs caused by the invasions of D. suzukii. This review has found few impact studies in the recently invaded areas and no economic evaluation of management programmes. There are not only geographical areas, but also themes that need more attention and analysis in the economic studies on D. suzukii. Assessing the economic effectiveness of integrated pest management programmes in specific settings, included the Latin American countries, is crucial.

Pheromone traps and climate variations influence populations of Sahlbergella singularis (Hemiptera: Miridae) and associated damage of cocoa in Cameroon.

Knowledge of insect pest ecology and biology is important for maximizing crop protection and reducing crop losses. Currently, we lack an efficient control program for the cocoa mirid Sahlbergella singularis Haglund (Hemiptera: Miridae), the principal insect pest of cocoa in West and Central Africa. A 2-yr study was conducted in 11 plantations across Ayos and Konye, two of the largest cocoa growing areas of Cameroon. We evaluated the effects of mirid sex pheromone and climatic variations on mirid population dynamics and their associated cocoa damage. Sex pheromone traps caught 1.5-fold higher mirids in Ayos than in Konye, with more overall counts in 2015 than in 2016. Cocoa pod counts were also significantly higher in 2015 than in 2016 and were negatively correlated with temperature and relative humidity. In both localities, mirid populations and associated cocoa pod damage were suppressed in plantations where sex pheromone traps were used. Damage incidence was positively correlated with mirid counts, confirming that the cocoa pod is the preferential site for mirid feeding and reproduction. As such, damage incidence could be used as proxy for comparative mirid population level due to the mirid's cryptic habit. Of the recorded weather variables, only relative humidity was correlated (negatively) with damage severity. Our data on the relationships between damage caused by mirids and their population dynamics and sex pheromone trap catches suggest that an effective control strategy using mass trapping could be developed for mirid management in cocoa plantations.

Trapping strategy and diel periodicity affect capture rate of Halyomorpha halys (Hemiptera: Pentatomidae) in agroecosystems.

The polyphagous pest, Halyomorpha halys (Stål) (Hemiptera: Pentatomidae), damages fruit in orchards and field crops and is often found within nearby woodlands. Pheromone-baited traps can be used to monitor H. halys. However, the efficiency of trapping H. halys may vary depending on trapping strategy (live vs. dead capture), location (ground or canopy), and diel periodicity of captures. We compared H. halys capture within fruiting hosts for: (i) live and kill traps on the ground vs. traps in the canopy of black cherry (Prunus serotina Ehrh.) (Rosales: Rosaceae), sugarberry (Celtis laevigata Willdenow) (Rosales: Cannabaceae), and pecan (Carya illinoinensis (Wangenh.) K. Koch) (Fagales: Juglandaceae) trees, (ii) ground and canopy-live traps in sassafras (Sassafras albidum (Nutt.) Nees) (Laurales: Lauraceae), and (iii) whether diel periodicity was detected for live capture in sassafras and cotton. More H. halys adults and nymphs were captured in kill traps than in live traps. More nymphs were captured in kill traps in black cherry and sugarberry on the ground than in the canopy. Live adult capture was significantly greater in sassafras and pecan canopies than on the ground. In cotton and sassafras, more live adults were captured from 8 PM-noon, with the fewest captured from noon-6 PM. A better understanding of stink bug activity in the field allows for improved trapping and, possibly, improved timing of treatment applications.

Efficacy of conventional and organic pesticides following ingestion by Delia radicum (Diptera: Anthomyiidae).

Cabbage maggot (CM) (Delia radicum L.) is a devastating pest of Brassicaceae crops throughout the world, including the Willamette Valley in western Oregon, USA. Chemical control methods for this pest are limited, with reduction or elimination of chlorpyrifos tolerances and expensive alternative chemistries; therefore, there is an increasing need for novel chemical control options. Adult feeding, a strategy used with insecticide-treated baits for other fly species, has yet to be tested as an option for a chemical control delivery for cabbage maggot. Treated bait can exploit the feeding behavior of CM and expose them to insecticides in a field setting. In this study, the efficacy of 5 organic and 5 conventional insecticides was compared in laboratory bioassays of treated bait stations in Aurora, Oregon, USA. The mortality of adult female cabbage maggot flies was assessed over time following ingestion of insecticides. Among organic insecticides tested, spinosad was highly effective 4 h after exposure, while pyrethrins + azadirachtin was moderately effective following 18 h after exposure. Flies exposed to conventional-use pesticides zeta-cypermethrin and bifenthrin had high mortality 1.75 h after exposure, while spinetoram had moderate efficacy 2 h after exposure. Insecticides identified with high or moderate efficacy may have the potential for use in baits or lure formulations that could be used to augment the control of cabbage maggots in field settings.

A method for sampling parasitized Drosophila suzukii (Diptera: Drosophilidae) puparia from soil.

Methods to measure the diversity and biological control impact of parasitoids for the control of spotted-wing drosophila, Drosophila suzukii (Matsumura) (Diptera: Drosophilidae) are being developed in support of biological control programs around the world. Existing methods to determine parasitism levels and parasitoid species composition focus on sampling D. suzukii within fresh and rotting fruit. However, many D. suzukii pupate in the soil or in dropped fruit, where additional parasitism could occur and where their parasitoids are thought to overwinter. Here we introduce a method for extracting parasitized D. suzukii puparia from the soil through a sieve and flotation system, allowing for effective collection of puparia, from which parasitoids can then be reared. Although the method considerably underestimates the absolute number of puparia in soil samples, it nonetheless yields a high number of puparia relative to sampling effort and provides a robust estimate of the relative abundance of puparia among samples. Using this method, we confirmed that at least 5 species of parasitoids, including some that have rarely been detected in past studies, overwinter in their immature stages inside D. suzukii puparia in south coastal British Columbia, Canada. The ability to sample puparia from the soil will lead to a more comprehensive view of both D. suzukii and parasitoid abundance throughout the season, help confirm parasitoid establishment following intentional releases, and provide a way to measure the diversity of parasitoid species and potential interactions among parasitoids (e.g., hyper- or klepto-parasitism) that may often occur on the soil surface.

Comparative analysis of the Potter Tower and a new Track Sprayer for the application of residual sprays in the laboratory.

BACKGROUND: Efforts to evaluate the residual efficacy of new indoor residual spraying (IRS) formulations have identified limitations with the industry standard laboratory sprayer, the Potter Spray Tower (PT). Calibrating the PT can be time-consuming, and the dosing of surfaces may not be as accurate or uniform as previously assumed. METHODS: To address these limitations, the Micron Horizontal Track Sprayer with Spray Cabinet (TS) was developed to provide higher efficiency, ease of operation and deposition uniformity equal to or better than the PT. A series of studies were performed using a fluorescent tracer and three IRS formulations (Actellic® 300CS, K-Othrine WG250 and Suspend PolyZone) sprayed onto surfaces using either the PT or the TS. RESULTS: Deposition volumes could be accurately calibrated for both spray systems. However, the uniformity of spray deposits was higher for the TS compared to the PT. Less than 12% of the volume sprayed using the PT reaches the target surface, with the remaining 88% unaccounted for, presumably vented out of the fume hood or coating the internal surfaces of the tower. In contrast, the TS deposits most of the spray on the floor of the spray chamber, with the rest contained therein. The total sprayed surface area in one run of the TS is 1.2 m2, and the operational zone for spray target placement is 0.7 m2, meaning that 58% of the applied volume deposits onto the targets. The TS can treat multiple surfaces (18 standard 15 × 15 cm tiles) in a single application, whereas the PT treats one surface at a time and a maximum area of around 0.0225 m2. An assessment of the time taken to perform spraying, including the setup, calibration and cleaning, showed that the cost of application using the TS was around 25-35 × less per tile sprayed. Standard operating procedures (SOPs) for calibration and use of both the Potter Tower and Track Sprayer have been developed. CONCLUSIONS: Overall, the TS represents a significant improvement over the PT in terms of the efficiency and accuracy of IRS formulation applications onto test substrates and offers a useful additional tool for researchers and manufacturers wanting to screen new active ingredients or evaluate the efficacy of IRS or other sprayable formulations for insect control.

Diffuse Associations Between Drosophila suzukii and Filamentous Fungal Microbes May Alter Caneberry Disease Dynamics.

Interactions between microorganisms and frugivorous insects can modulate fruit rot disease epidemiology. Insect feeding and/or oviposition wounds may create opportunities for fungal infection. Passive and active dispersal of fungal inoculums by adult insects also increases disease incidence. In fall-bearing raspberries and blackberries, such vectoring interactions could increase crop damage from the invasive pestiferous vinegar fly Drosophila suzukii (spotted-wing drosophila). Periods of peak D. suzukii activity are known to overlap with several species of primary fruit rot pathogen, particularly Botrytis cinerea and Cladosporium cladosporioides, and previous work indicates that larvae co-occur with and feed on various filamentous fungi at low rates. To further our understanding of the epidemiological consequences that may emerge from these associations, we surveyed the filamentous fungal community associated with adult D. suzukii, isolating and molecularly identifying fungi externally and internally (indicating feeding) from field-collected adults over 3 years. We isolated and identified 37 unique genera of fungi in total, including known raspberry pathogens. Most fungi were detected infrequently, and flies acquired and carried fungi externally at higher richness, frequency, and density relative to internally. In a worst-case scenario laboratory vectoring assay, D. suzukii adults were able to transfer B. cinerea and C. cladosporioides to sterile media at 0, 24, 48, and 72 h after exposure to sporulating cultures in Petri dishes. These results collectively suggest an adventitious vectoring association between D. suzukii and fruit rot fungi that has the potential to alter caneberry disease dynamics.

Using electric fields to control insects: current applications and future directions.

Chemical-based interventions are mostly used to control insects that are harmful to human health and agriculture or that simply cause a nuisance. An overreliance on these insecticides however raises concerns for the environment, human health, and the development of resistance, not only in the target species. As such, there is a critical need for the development of novel nonchemical technologies to control insects. Electrocution traps using UV light as an attractant are one classical nonchemical approach to insect control but lack the specificity necessary to target only pest insects and to avoid harmless or beneficial species. Here we review the fundamental physics behind electric fields (EFs) and place them in context with electromagnetic fields more broadly. We then focus on how novel uses of strong EFs, some of which are being piloted in the field and laboratory, have the potential to repel, capture, or kill (electrocute) insects without the negative side effects of other classical approaches. As EF-insect science remains in its infancy, we provide recommendations for future areas of research in EF-insect science.

Trichopria anastrephae as a Biological Control Agent of Drosophila suzukii in Strawberries.

In South America, the resident pupal parasitoid Trichopria anastrephae Costa Lima (Hymenoptera: Diapriidae) is a potential biological control agent of the pest Drosophila suzukii Matsumura (Diptera: Drosophilidae). In the present study, we (1) examined the behavior of T. anastrephae towards different host (D. suzukii) and host-substrate (strawberry) cues in choice and non-choice bioassays in laboratory, and (2) examined the density-dependent parasitism of T. anastrephae in D. suzukii-infested strawberries in a greenhouse. When given a choice, female parasitoids walked longer over chambers with fruits infested with eggs, larvae, or pupae of D. suzukii, when compared to healthy uninfested strawberries, and over overripe fruits when compared to unripe or ripe fruits. In the greenhouse assay, we observed an increase in parasitism and a decrease in the number of D. suzukii emerging per fruit with an increase in the number of parasitoids released. Our results allow a better understanding of the behavior and parasitism of T. anastrephae in D. suzukii-infested strawberries and provide useful data for potential biological control programs using this parasitoid.

Neotropical Flora's Contribution to the Development of Biorational Products for Drosophila suzukii Control.

Essential oils (EOs) produced by aromatic plants belonging to different families, such as Asteraceae, Lamiaceae, Lauraceae, Myrtaceae, and Piperaceae, are generally suggested as potential sources of new molecules with insecticidal activity. The EOs are constituted bioactive molecules that may have to control Drosophila suzukii (Matsumura), a serious economic invasive pest of small fruits worldwide. Currently, the control strategy against D. suzukii depends especially on treatment with synthetic insecticides. Due to impacts to human health and the environment, efforts have been made to seek efficient insecticides in chemical pest control. Thus, sixty-five oils extracted from plants were selected to find new alternative types of insecticides active against D. suzukii. The monoterpenes, such as limonene, α-pinene, 1,8-cineole, linalool, menthol, geranial, and neral, were the most representative, which stand out for their insecticidal efficiency. The OEs demonstrated to be used in the management of D. suzukii, thus being an effective strategy to control this pest, ensuring crop protection and agricultural sustainability. Therefore, the substitution by natural products or eco-friendly pesticides instead of synthetic pesticides represents a notable option to mitigate harmful effects on human health and the environment.

Ants evade harmful food by active abandonment.

Invasive ants, such as the Argentine ant, pose a severe economic and ecological threat. Despite advancements in baiting techniques, effectively managing established ant populations remains a daunting challenge, often ending in failure. Ant colonies employ behavioural immunity against pathogens, raising the question of whether ants can collectively respond to toxic baits. This study investigates whether ant colonies actively abandon palatable but harmful food sources. We provided two sucrose feeders, each generating a new foraging trail, with one transitioning to offering toxic food. Six hours later, ant activity on that path decreases, while activity on the non-toxic food and the trunk trail remains unaffected, excluding factors like population decline or satiation as reasons for the activity decline. Laboratory experiments confirmed that ants remained alive six hours after ingesting toxic food. Ant presence remains low on the toxic food path for days, gradually decreasing along the nearest section of the trunk trail. This abandonment behaviour minimises the entry of harmful food into the nest, acting as a protective social mechanism. The evasion of toxic bait-treated areas likely contributes considerably to control failures. Understanding the behavioural response to toxic baits is essential for developing effective strategies to combat invasive ant species.

Volatiles from nutritional fungal symbiont influence the attraction of Anisandrus maiche (Coleoptera: Curculionidae) to ethanol-baited traps.

Anisandrus maiche Stark (Coleoptera: Curculionidae: Scolytinae) is a non-native ambrosia beetle from central Asia that has been spreading throughout the eastern United States since 2005. Preferred hosts of A. maiche are not well characterized within its currently invaded range, but it is established in managed and natural forests throughout Indiana. Current monitoring and detection efforts for this beetle rely on ethanol-baited traps, but fungal volatiles may alter the attraction of A. maiche to ethanol. In this study, we conducted trapping experiments in Indiana to determine the extent to which a suite of common fungal alcohols influences the response of A. maiche to ethanol-baited traps. We then evaluated isoamyl and isobutyl alcohol as potential attractants for A. maiche and their ability to enhance attraction to ethanol. Lastly, we used SPME-GC-MS to identify volatiles from Ambrosiella cleistominuta (Mayers & Harr.), the fungal symbiont of A. maiche, grown for 7 and 14 days on malt extract agar. Benzyl alcohol, isobutyl alcohol, hexanol, methyl phenylacetate, phenethyl alcohol, and piperitone reduced the attraction of A. maiche to ethanol-baited traps in the field. Moreover, adding methyl benzoate and isoamyl alcohol individually to ethanol-baited traps did not further increase A. maiche capture. When paired with ethanol, isoamyl alcohol repelled beetles in the early flight period but did not significantly increase trap capture during the fall flight. These results represent a first step in understanding the role of fungal volatiles in the colonization behavior of A. maiche and may ultimately inform management strategies for this species.

Color, pattern, and background contrast affect attraction of Euxesta eluta and Chaetopsis massyla (Diptera: Ulidiidae).

Euxesta eluta Loew and Chaetopsis massyla Walker (Diptera: Ulidiidae) are primary pests of sweet corn in Florida. Attraction of adult flies to various visual stimuli was evaluated in the laboratory to provide insight into the potential development of enhanced trapping strategies. In assays evaluating different colored sticky traps, more E. eluta were collected on light blue, mid blue, lime green, and orange yellow traps, whereas attraction of C. massyla was greater to lime green and fluorescent green traps. In a comparison of yellow 3-dimensional shapes, more E. eluta were collected on yellow cylinders than on spheres or cubes; however, more C. massyla were collected on cylinders than on cubes or spheres. When colored traps were placed against a white background, more E. eluta were collected on lime green compared with yellow, blue, and orange traps; however, when placed against a black background, attraction to the lime green trap was reduced. Against the white background attraction of C. massyla, was strongest to yellow, then lime green and orange traps, followed by blue, but with a black background, differences between traps collections were reduced. The addition of ultraviolet (UV) reflectance to yellow increased the attraction of E. eluta, but C. massyla were more attracted to yellows without UV reflection or fluorescence. Black patterns on yellow traps affected attraction, with E. eluta more attracted to wide stripes, a large square, or many small squares, whereas C. massyla was more strongly attracted to stripes. Utilization of these visual attributes could be useful in improving surveillance for these species.

The exposure risk of heavy metals to insect pests and their impact on pests occurrence and cross-tolerance to insecticides: A review.

Heavy metal (HM) pollution is a severe global environmental issue. HMs in the environment can transfer along the food chain, which aggravates their ecotoxicological effect and exposes the insects to heavy metal stress. In addition to their growth-toxic effects, HMs have been reported as abiotic environmental factors that influence the implementation of integrated pest management strategies, including microbial control, enemy insect control, and chemical control. This will bring new challenges to pest control and further highlight the ecotoxicological impact of HM pollution. In this review, the relationship between HM pollution and insecticide tolerance in pests was analyzed. Our focus is on the risks of HM exposure to pests, pests tolerance to insecticides under HM exposure, and the mechanisms underlying the effect of HM exposure on pests tolerance to insecticides. We infer that HM exposure, as an initial stressor, induces cross-tolerance in pests to subsequent insecticide stress. Additionally, the priming effect of HM exposure on enzymes associated with insecticide metabolism underlies cross-tolerance formation. This is a new interdisciplinary field between pollution ecology and pest control, with an important guidance value for optimizing pest control strategies in HM polluted areas.

A fast and reliable larval sampling method for improving the monitoring of fruit flies in soft and stone fruits.

The spotted-wing drosophila, Drosophila suzukii (Matsumura) (Diptera: Drosophilidae), threatens both the soft-skinned and stone fruit industry in Asia, Europe, and America. Integrated pest management requires monitoring for infestation rates in real time. Although baited traps for adult D. suzukii are widely used for field monitoring, trap captures are weakly correlated to larval infestation rates. Thus, monitoring for larvae instead of adult flies represents the most reliable monitoring technique. Current methods for larval monitoring (e.g., sugar or salt floatation) are time-consuming and labor-intensive. In this study, we develop a new "sleeve method" for detecting larvae in strawberries through the inspection of individual fruits crushed within transparent plastic sleeves. Samples can be optionally frozen until further processing. Based on count data from non-expert observers, the estimation of larval infestation with the sleeve method is fast, precise, and highly repeatable within and among observers. Mean processing time is half the time compared to previous methods (33-80 s per sample depending on infestation levels). As the accuracy of the sleeve method decreases with infestation levels, we suggest ways to improve its accuracy by incubating fruits for 48 h and calibrating data using fruits with a known number of larvae. The method could also be used in other fruits, as it is easier to use, faster, and requires less equipment than previous monitoring methods. Finally, the method represents a promising tool for growers or researchers to effectively monitor and manage D. suzukii and other insect pests of soft and stone fruits.

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%.