Tapping for love: courtship, mating, and behavioral asymmetry in two aphid parasitoids, Aphidius ervi and Aphidius matricariae (Hymenoptera: Braconidae: Aphidinae).

Understanding the biology and ecology of parasitoids can have direct implications for their evaluation as biological control agents, as well as for the development and implementation of mass-rearing techniques. Nonetheless, our current knowledge of the possible influence of lateralized displays (i.e., the asymmetric expression of cognitive functions) on their reproductive behavior is scarce. Herein, we characterized the behavioral elements involved in courtship, and quantified the durations of 2 important aphid parasitoids, Aphidius ervi Haliday and Aphidius matricariae Haliday (Hymenoptera: Braconidae: Aphidiinae). We quantified the main indicators of copulation and examined the occurrence of lateralized traits at population level. Results indicated that A. matricariae exhibited longer durations of wing fanning, antennal tapping, pre-copula and copula phases compared to A. ervi. Postcopulatory behavior was observed only in A. matricariae. Unlike other parasitoid species, the duration of wing fanning, chasing, and antennal tapping did not affect the success of the mating of male A. ervi and A. matricariae. Both species exhibited a right-biased female kicking behavior at the population level during the pre-copula. Our study provides insights into the fundamental biology of aphidiine parasitoids and reports the presence of population-level lateralized mating displays, which can serve as useful benchmarks to evaluate the quality of mass-rearing systems.

Grazing system and Hyalomma marginatum tick infestation in cattle with high prevalence of SFG Rickettsia spp.

Tick-borne diseases (TBDs) represent a significant portion of infectious diseases of global public health interest. In Italy, knowledge about the occurrence of tick-borne pathogens (TBPs) in ticks parasitizing cattle is scarce. In this research, we focused on ticks infesting Maremmana cattle grazing in open pasture and silvopasture systems. After being morphologically identified, ticks were molecularly tested for the presence of pathogens of the genus Rickettsia. Of the 794 ticks detected, 117 were collected, being the majority Hyalomma marginatum (72.6%), followed by other Hyalomma species (23%), Rhipicephalus turanicus (1.7%), Rh. bursa (0.9%), Hy. lusitanicum (0.9%) and Dermatocentor marginatus (0.9%). All ticks were adults, 58.1% males and 41.8% females. The highest tick prevalence was noted in April for silvopasture system cattle (90%), and in May for open pasture ones (85%). TBPs were detected only in Hy. marginatum, and all belong to Rickettsia spp. of zoonotic interest. In particular, 21/40 (52.5%) ticks scored positive for Rickettsia spp. by gltA gene and of these 15/21 (71.4%) also to spotted fever group (SFG) rickettsiae by ompA gene. Of the total positive specimens, 19 were successfully sequenced and scored Rickettsia aeschilimannii (17/19, 89.5%), R. slovaca (1/19, 5%), and R. massiliae (1/19, 5%). This research highlights the potential impact of grazing systems on cattle parasitization by hard ticks. The molecular investigation of TBPs in ticks collected from Maremmana cattle shed light on the presence of pathogenic bacteria of SFG Rickettsia spp., pointing out the potential risk of TBPs transmission between livestock and humans.

Lethal and sublethal effects of carlina oxide on Tetranychus urticae (Acari: Tetranychidae) and Neoseiulus californicus (Acari: Phytoseiidae).

BACKGROUND: Tetranychus urticae Koch, is a polyphagous and damaging pest, presenting several resistant populations worldwide. Among new and more environmentally friendly control tools, botanical pesticides represent a valuable alternative to synthetic ones within integrated pest management strategies. Accordingly, we investigated the lethal and sublethal effects of carlina oxide isolated from Carlina acaulis (Asteraceae) roots on T. urticae and its natural enemy, the predatory mite, Neoseiulus californicus (McGregor). RESULTS: Carlina oxide (98.7% pure compound) was used for acaricidal tests on eggs, nymphs, and adult females of T. urticae (concentrations of 312.5, 625, 1250, 2500 and 5000 µL L-1 ), and eggs and females of N. californicus (1250 and 5000 µL L-1 on eggs and females, respectively). Behavioral two-choice tests were also conducted on phytoseiid females. Carlina oxide toxicity was higher on T. urticae females than nymphs (median lethal dose 1145 and 1825 µL L-1 , respectively), whereas egg mortality and mean hatching time were significantly affected by all tested concentrations. A decreasing daily oviposition rate for T. urticae was recorded with concentrations ranging from 625 to 5000 µL L-1 , whereas negative effects on the population growth rate were recorded only with the three higher concentrations (1250, 2500 and 5000 µL L-1 ). No toxic effect on N. californicus females was found, but a strong repellent activity lasting for 48 h from application was recorded. CONCLUSION: Carlina oxide reduced longevity and fecundity of T. urticae adults, but not of N. californicus. This selective property allows us to propose it as a novel active ingredient of ecofriendly acaricides for T. urticae management. © 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.

Learning algorithms estimate pose and detect motor anomalies in flies exposed to minimal doses of a toxicant.

Pesticide exposure, even at low doses, can have detrimental effects on ecosystems. This study aimed at validating the use of machine learning for recognizing motor anomalies, produced by minimal insecticide exposure on a model insect species. The Mediterranean fruit fly, Ceratitis capitata (Diptera: Tephritidae), was exposed to food contaminated with low concentrations of Carlina acaulis essential oil (EO). A deep learning approach enabled fly pose estimation on video recordings in a custom-built arena. Five machine learning algorithms were trained on handcrafted features, extracted from the predicted pose, to distinguish treated individuals. Random Forest and K-Nearest Neighbor algorithms best performed, with an area under the receiver operating characteristic (ROC) curve of 0.75 and 0.73, respectively. Both algorithms achieved an accuracy of 0.71. Results show the machine learning potential for detecting sublethal effects arising from insecticide exposure on fly motor behavior, which could also affect other organisms and environmental health.

Formal analyses are fundamental for the definition of honey, a product representing specific territories and their changes: the case of North Tyrrhenian dunes (Italy).

Honey is a variegate matrix depending significantly on the floral origin, and it could become an important agri-food product to valorise specific territories. Being so diverse, different analytical techniques are necessary for its description. Herein we characterized the honey produced in one of the Italian sand dunes systems hosting beekeeping activities. In terms of floristic origin, phytochemical characterization, and sensory and colour analysis, honey collected in 2021 and 2022 was comparable. Honey was polyfloral, with several pollens from dune habitat plants classified as minor. The presence of the allochthonous Amorpha fruticosa L. and the ruderal Rubus fruticosus L. pollens in the category of the secondary pollens testifies the alteration of the park vegetation. The phytochemical profile was rich in polyphenols. Other interesting compounds were coumarine derivatives, likely attributable to resin-laden plants as rockroses, long chain hydroxyacids typical of royal jelly and nicotinic acid and its analogues (2-hydroxynicotinic acid and 2-hydroxyquinoline). The above-mentioned honey showed interesting features and was a good representation of the vegetation of this area. Our study pointed out the importance of relying on multiple analytical techniques for the characterization of honey and the advisability of a technical support toward beekeepers to correctly describe and valorise their product.

Do changes in Lactuca sativa metabolic performance, induced by mycorrhizal symbionts and leaf UV-B irradiation, play a role towards tolerance to a polyphagous insect pest?

The increased ultraviolet radiation (UV) due to the altered stratospheric ozone leads to multiple plant physiological and biochemical adaptations, likely affecting their interaction with other organisms, such as pests and pathogens. Arbuscular mycorrhizal fungi (AMF) and UV-B treatment can be used as eco-friendly techniques to protect crops from pests by activating plant mechanisms of resistance. In this study, we investigated plant (Lactuca sativa) response to UV-B exposure and Funneliformis mosseae (IMA1) inoculation as well as the role of a major insect pest, Spodoptera littoralis. Lettuce plants exposed to UV-B were heavier and taller than non-irradiated ones. A considerable enrichment in phenolic, flavonoid, anthocyanin, and carotenoid contents and antioxidant capacity, along with redder and more homogenous leaf color, were also observed in UV-B-treated but not in AMF-inoculated plants. Biometric and biochemical data did not differ between AMF and non-AMF plants. AMF-inoculated plants showed hyphae, arbuscules, vesicles, and spores in their roots. AMF colonization levels were not affected by UV-B irradiation. No changes in S. littoralis-feeding behavior towards treated and untreated plants were observed, suggesting the ability of this generalist herbivore to overcome the plant chemical defenses boosted by UV-B exposure. The results of this multi-factorial study shed light on how polyphagous insect pests can cope with multiple plant physiological and biochemical adaptations following biotic and abiotic preconditioning.

Lethal and behavioural effects of a green insecticide against an invasive polyphagous fruit fly pest and its safety to mammals.

Plant essential oil-based insecticides, with special reference to those that may be obtained from largely available biomasses, represent a valuable tool for Integrated Pest Management. However, the sublethal effects and the potential effects on aggressive insect traits of these green insecticides are understudied. Herein, the lethal and sub-lethal effects of the carlina oxide, constituting more than 97% of the whole Carlina acaulis (Asteraceae) root essential oil (EO), were determined against an invasive polyphagous tephritid pest, Ceratitis capitata (medfly). The carlina oxide was formulated in a mucilaginous solution containing carboxymethylcellulose sodium salt, sucrose, and hydrolysed proteins, showing high ingestion toxicity on medfly adults. The behavioural effects of carlina oxide at LC10 and LC30 were evaluated on the medfly aggressive traits, which are crucial for securing reproductive success in both sexes. Insecticide exposure affected the directionality of aggressive actions, but not the aggression escalation intensity and duration. The EO safety to mammals was investigated by studying its acute toxicity on the stomach, liver, and kidney of rats after oral administration. Only the highest dose (1000 mg/kg) of the EO caused modest neurological signs and moderate effects on the stomach, liver, and kidney. The other doses, which are closer to the practical use of the EO when formulated in protein baits, did not cause side effects. Overall, C. acaulis-based products are effective and safe to non-target mammals, deserving further consideration for eco-friendly pesticide formulations.

Leaf UV-B Irradiation and Mycorrhizal Symbionts Affect Lettuce VOC Emissions and Defence Mechanisms, but Not Aphid Feeding Preferences.

Arbuscular mycorrhizal fungi (AMF) and ultraviolet-B radiation (UV-B) play important roles in plant-insect interactions by altering plant physiology and histology. We hypothesized that UV-B-induced oxidative stress was mitigated by AMF symbiosis. In this study, we conducted a multifactorial experiment to explore lettuce plant response to AMF inoculation and UV-B exposure (0.4 W m-2; 16 h d-1; 2 weeks), either together or individually, as well as the interaction with the polyphagous insect pest Myzus persicae (Sulzer). Lettuce plants subjected to UV-B radiation showed an increase in callose and oxidative stress indicators, as well as a decrease in stomatal density. Mycorrhizal colonization cancelled out the effect of UV-B on stomatal density, while the symbiosis was not affected by UV-B treatment. The plant volatile emission was significantly altered by UV-B treatment. Specifically, the non-terpene 1-undecene abundance (+M/+UVB: 48.0 ± 7.78%; -M/+UVB: 56.6 ± 14.90%) was increased, whereas the content of the non-terpene aldehydes decanal (+M/+UVB: 8.50 ± 3.90%; -M/+UVB: 8.0 ± 4.87%) and undecanal (+M/+UVB: 2.1 ± 0.65%; -M/+UVB: 1.20 ± 1.18%) and the sesquiterpene hydrocarbons (+M/+UVB: 18.0 ± 9.62 %; -M/+UVB: 19.2 ± 5.90%) was decreased. Mycorrhization, on the other hand, had no significant effect on the plant volatilome, regardless of UV-B treatment. Aphid population was unaffected by any of the treatments, implying a neutral plant response. Overall, this study provides new insights about the interactions among plants, UV-B, and AMF, outlining their limited impact on a polyphagous insect pest.

Diatomaceous Earth for Arthropod Pest Control: Back to the Future.

Nowadays, we are tackling various issues related to the overuse of synthetic insecticides. Growing concerns about biodiversity, animal and human welfare, and food security are pushing agriculture toward a more sustainable approach, and research is moving in this direction, looking for environmentally friendly alternatives to be adopted in Integrated Pest Management (IPM) protocols. In this regard, inert dusts, especially diatomaceous earths (DEs), hold a significant promise to prevent and control a wide range of arthropod pests. DEs are a type of naturally occurring soft siliceous sedimentary rock, consisting of the fossilized exoskeleton of unicellular algae, which are called diatoms. Mainly adopted for the control of stored product pests, DEs have found also their use against some household insects living in a dry environment, such as bed bugs, or insects of agricultural interest. In this article, we reported a comprehensive review of the use of DEs against different arthropod pest taxa, such as Acarina, Blattodea, Coleoptera, Diptera, Hemiptera, Hymenoptera, Ixodida, Lepidoptera, when applied either alone or in combination with other techniques. The mechanisms of action of DEs, their real-world applications, and challenges related to their adoption in IPM programs are critically reported.

Bioactivity of Carlina acaulis Essential Oil and Its Main Component towards the Olive Fruit Fly, Bactrocera oleae: Ingestion Toxicity, Electrophysiological and Behavioral Insights.

Among botanical insecticides based on essential oils (EOs) or their main components, Carlina acaulis EO and the aromatic polyacetylene carlina oxide, constituting more than 90% of its EO, were recently proven to be effective against the larvae and adults of some insect vectors and pests. In this study, the toxicity of C. acaulis EO and carlina oxide were tested on Bactrocera oleae adults using a protein bait formulation. The LC50 values of the C. acaulis EO and carlina oxide were 706 ppm and 1052 ppm, respectively. Electroantennographic (EAG) tests on B. oleae adults showed that both carlina EO and oxide elicited EAG dose-dependent responses in male and female antennae. The responses to the EO were significantly higher than those to carlina oxide, indicating that other compounds, despite their lower concentrations, can play a relevant role. Moreover, Y-tube assays carried out to assess the potential attractiveness or repellency of carlina oxide LC90 to B. oleae adults showed that it was unattractive to both males and females of B. oleae, and the time spent by both sexes in either the control or the treatment arm did not differ significantly. Overall, this study points out the potential use of C. acaulis EO and carlina oxide for the development of green and effective "lure-and-kill" tools.

Toxics or Lures? Biological and Behavioral Effects of Plant Essential Oils on Tephritidae Fruit Flies.

The family Tephritidae (Diptera) includes species that are highly invasive and harmful to crops. Due to globalization, international trade, and human displacement, their spread is continuously increasing. Unfortunately, the control of tephritid flies is still closely linked to the use of synthetic insecticides, which are responsible for detrimental effects on the environment and human health. Recently, research is looking for alternative and more eco-friendly tools to be adopted in Integrated Pest Management (IPM) programs. In this regard, essential oils (EOs) and their main compounds represent a promising alternative to chemical insecticides. EOs are made up of phytoconstituents formed from the secondary metabolism of many plants and can act as attractants or toxics, depending on the dose. Because of this unique characteristic, EOs and their main constituents are promising tools that can be used both in Sterile Insect Technique (SIT) programs and in the "lure and kill" technique, exploiting the attractiveness of the product in the former case and its toxicity in the latter. In this article, current knowledge on the biological and behavioral effects of EOs and their main constituents on tephritid fruit flies is reviewed, mainly focusing on species belonging to the Anastrepha, Bactrocera, Ceratitis, and Zeugodacus genera. The mechanisms of action of EOs, their real-world applications, and challenges related to their use in IPM are critically discussed.

Old Parasitoids for New Mealybugs: Host Location Behavior and Parasitization Efficacy of Anagyrus vladimiri on Pseudococcus comstocki.

The Comstock mealybug, Pseudococcus comstocki (Hemiptera: Pseudococcidae) is a primary pest of orchards in the North and Northwest of China. This pest appeared recently in Europe, including Italy, where it is infesting mainly vineyards as well as apple and pear orchards. The present study investigated the efficacy of Anagyrus vladimiri, a known biological control agent (BCA) of Planococcus ficus, on P. comstocki to evaluate a potential use for the management of this new pest. No-choice tests were conducted to quantify the parasitoid behavior against P. ficus and P. comstocki. The parasitoid successfully parasitized both species (parasitization rate: 51% and 67% on P. comstocki and P. ficus, respectively). The A.vladimiri developmental time (19.67 ± 1.12 vs. 19.70 ± 1.07 days), sex ratio (1.16 ± 1.12 vs. 1.58 ± 1.07) and hind tibia length of the progeny showed no differences when P. comstocki and P. ficus, respectively, were exploited as hosts. Two-choice tests, conducted by providing the parasitoid with a mixed population of P. ficus and P. comstocki, showed no host preference for either of the two mealybug species (23 vs. 27 first choices on P. comstocki and P. ficus, respectively). The parasitization rate (61.5% and 64.5% in P. comstocki and P. ficus, respectively) did not differ between the two hosts. Overall, our study adds basic knowledge on parasitoid behavior and host preferences and confirms the use of this economically important encyrtid species as an effective BCA against the invasive Comstock mealybug.

Developing a Highly Stable Carlina acaulis Essential Oil Nanoemulsion for Managing Lobesia botrana.

The growing interest in the development of green pest management strategies is leading to the exploitation of essential oils (EOs) as promising botanical pesticides. In this respect, nanotechnology could efficiently support the use of EOs through their encapsulation into stable nanoformulations, such as nanoemulsions (NEs), to improve their stability and efficacy. This technology assures the improvement of the chemical stability, hydrophilicity, and environmental persistence of EOs, giving an added value for the fabrication of natural insecticides effective against a wide spectrum of insect vectors and pests of public and agronomical importance. Carlina acaulis (Asteraceae) root EO has been recently proposed as a promising ingredient of a new generation of botanical insecticides. In the present study, a highly stable C. acaulis-based NE was developed. Interestingly, such a nanosystem was able to encapsulate 6% (w/w) of C. acaulis EO, showing a mean diameter of around 140 nm and a SOR (surfactant-to-oil ratio) of 0.6. Its stability was evaluated in a storage period of six months and corroborated by an accelerated stability study. Therefore, the C. acaulis EO and C. acaulis-based NE were evaluated for their toxicity against 1st instar larvae of the European grapevine moth (EGVM), Lobesia botrana (Denis & Schiffermüller, 1775) (Lepidoptera: Tortricidae), a major vineyard pest. The chemical composition of C. acaulis EO was investigated by gas chromatography-mass spectrometry (GC-MS) revealing carlina oxide, a polyacetylene, as the main constituent. In toxicity assays, both the C. acaulis EO and the C. acaulis-based NE were highly toxic to L. botrana larvae, with LC50 values of 7.299 and 9.044 µL/mL for C. acaulis EO and NE, respectively. The C. acaulis-based NE represents a promising option to develop highly stable botanical insecticides for pest management. To date, this study represents the first evidence about the insecticidal toxicity of EOs and EO-based NEs against this major grapevine pest.

Trapping Brown Marmorated Stink Bugs: "The Nazgȗl" Lure and Kill Nets.

Improvements to current brown marmorated stink bug (BMSB), Halyomorpha halys, surveillance and killing systems are needed to improve detection sensitivity and to reduce pesticide use. Detection of BMSB in New Zealand with traps is reliant on sticky panels with aggregation pheromone, which are low cost but inefficient compared with beating foliage. Trapping for BMSB adults and nymphs was conducted daily with lethal traps consisting of an aggregation pheromone-baited-coat hanger covered with dark-colored long-lasting insecticide-treated mesh, we termed "The Nazgȗl", based on its sinister appearance. A deep tray lined with white plastic was attached centrally at the base for collecting the dead BMSB. The lethal traps killed and caught up to 3.5-fold more nymphs and adult BMSB than identically-baited sticky panels in the 3 weeks of deployment, and provided a snapshot of phenology by instar. We expect that lure-and-kill stations could contribute to the suppression of a delimited population and could be included as part of a semiochemical-based eradication program. Attracting and killing females and nymphs, thus removing future offspring, could contribute to population suppression during an eradication.

The Competitive Mating of Irradiated Brown Marmorated Stink Bugs, Halyomorpha halys, for the Sterile Insect Technique.

The sterility of eggs and nymphs from gamma-irradiated male Halyomorpha halys was investigated to determine the potential for the sterile insect technique (SIT). Males irradiated at 0, 16, 24 and 32 Gy were placed with untreated virgin females, and egg sterility was determined, showing 54.3% at 16 Gy. The percentage of sterility from irradiation was 26 percent lower than previous results from the USA and the variance was very high. Competitive overflooding ratio trials between irradiated virgin males and fertile virgin males at a 5:1 ratio resulted in the expected egg sterility, indicating competitive performance by irradiated males. By July and August, older, irradiated overwintered males were significantly less competitive than similar, non-irradiated males. There is a need to revisit the irradiation delivery method to achieve proper precision around the paternal dose required for an expected >80% egg sterility and subsequent ~99% endpoint sterility estimated at adult emergence in the F1 phase. These results suggest that the mating competitiveness and competency of males after irradiation at 16 Gy is not limiting to the sterile insect technique for suppression. A wild harvest of overwintering males using the aggregation pheromone, followed by irradiation and male release, might replace rearing. Mass-collected, sterilized bugs could be transported from an area of high H. halys density and shipped for release to enable suppression or eradication elsewhere. This concept is under development but further work is needed now to understand the difference in results between the US and Italian irradiators and increase the reliability of dosimetry.

Green Micro- and Nanoemulsions for Managing Parasites, Vectors and Pests.

The management of parasites, insect pests and vectors requests development of novel, effective and eco-friendly tools. The development of resistance towards many drugs and pesticides pushed scientists to look for novel bioactive compounds endowed with multiple modes of action, and with no risk to human health and environment. Several natural products are used as alternative/complementary approaches to manage parasites, insect pests and vectors due to their high efficacy and often limited non-target toxicity. Their encapsulation into nanosystems helps overcome some hurdles related to their physicochemical properties, for instance limited stability and handling, enhancing the overall efficacy. Among different nanosystems, micro- and nanoemulsions are easy-to-use systems in terms of preparation and industrial scale-up. Different reports support their efficacy against parasites of medical importance, including Leishmania, Plasmodium and Trypanosoma as well as agricultural and stored product insect pests and vectors of human diseases, such as Aedes and Culex mosquitoes. Overall, micro- and nanoemulsions are valid options for developing promising eco-friendly tools in pest and vector management, pending proper field validation. Future research on the improvement of technical aspects as well as chronic toxicity experiments on non-target species is needed.