High Larval Mortality Research Articles

Growth Performance of Galaxy Rasbora Danio margaritatus Larvae Maintained at Different Stocking Densities

The issue of low survival rates and high mortality of larvae is commonly encountered during the rearing process. The success of larval rearing is significantly influenced by optimal stocking density, good water quality, and controlled feed management, all of which affect the growth and survival of the larvae. The aim of this applied research is to implement rearing techniques with different stocking densities and assess their effects on the growth and survival of galaxy rasbora (Danio margaritatus) larvae. This study employed a Completely Randomized Design (CRD) with four different stocking density treatments: 1 fish L⁻¹ for treatments KT1 and PT1, 2 fish L⁻¹ for treatment PT2, and 3 fish L⁻¹ for treatment PT3, each with three repetitions. The results showed that different stocking densities had a significant effect on the growth and survival of the larvae. The highest survival rate was observed in treatment PT2, with 96% survival. The control treatment (KT1), which used a semi-intensive system, achieved the highest absolute length growth of 6.87 mm and the highest daily length growth rate of 32.73% day⁻¹. The stocking density of 2 fish L⁻¹ in treatment PT2 demonstrated optimal results in terms of survival and growth.

Potential of native entomopathogenic fungi isolates against the fall armyworm, Spodoptera frugiperda (Lepidoptera: Noctuidae) in Maize in Ethiopia

ABSTRACT The fall armyworm (Spodoptera frugiperda) invasion poses a significant threat to maize production in Africa. Some entomopathogenic fungal strains, particularly those from the genera Beauveria and Metarhizium, have been considered as potential biological control agents for this pest. When these entomopathogenic fungi are present in fields, they can not only infect individual pests but also spread among pest populations, leading to long-term suppression. In this study, 17 entomopathogenic fungi isolates, including 12 Beauveria spp. and 5 Metarhizium spp., were evaluated under laboratory conditions. Pathogenicity assessments revealed significant variation in mortality rates among the isolates. Specifically, Beauveria isolates APPRC-44BC and APPRC-27 demonstrated high larval mortality rates of 88.7% and 83.5%, respectively. The dose–response study showed that APPRC-44BC had the lowest LC50 (2.8 × 105 conidia/mL) and LC90 (2.2 × 107 conidia/mL) values. Subsequently, the three most pathogenic isolates, two from Beauveria spp. (APPRC-44BC and APPRC-27) and one from Metarhizium sp. (APPRC-34GM), were evaluated under field conditions in maize. Treatments with these isolates, particularly APPRC-44BC, significantly reduced leaf damage and increased yield compared to the negative control. These findings underscore the potential of native entomopathogenic fungi isolates as sustainable alternatives for managing S. frugiperda. The present study addresses a critical knowledge gap in Africa and offers an environmentally friendly alternative to chemical pesticides for effective pest management. Further evaluation of these promising isolates in diverse agroecological settings is needed to confirm their effectiveness across different regions.

Immune stimulation by inactivated Vibrio splendidus promotes the development and resistance of oyster Crassostrea gigas larvae

The oyster Crassostrea gigas larvae in embryonic development stage have been suffering high mortality rate during hatching, which have seriously hindered the sustainable development of oyster seedling industry. This study explored the immune protection and developmental effects of priming with inactivated Vibrio splendidus in oyster larvae at middle umbo stage (10 d post fertilization, dpf). The results showed that the immune system of umbo larvae was activated after pre-immune stimulation with inactivated V. splendidus. The expressions of immune recognition receptors (CTL-3, Integrin β-1, TLR4), NF-kB signaling component (IKK), effector molecules (IL17-5, Defh2, HSP70) were significantly up-regulated, and the activities of antioxidant enzymes (superoxide dismutase and catalase), hydrolytic enzyme (lysozyme) also increased significantly. The proteins from the stimulated umbo larvae have obvious characteristics of agglutination and inhibition of V. splendidus growth. When the larvae at late umbo (18 dpf) or pediveliger stage (21 dpf) were challenged by live V. splendidus, much lower death rate was observed in the stimulation group compared to the control group. Simultaneously, the expressions of above immune related genes and the activities of antioxidant enzymes were all rapidly up-regulated in pediveliger larvae of immune stimulated group. Moreover, the significantly increased shell height and shell length as well as accelerated development rate, and higher settlement rate were revealed after the umbo larvae are stimulated by inactivated Vibrio splendidus. In summary, inactivated V. splendidus stimulation in oyster umbo larvae could activate their immune system, enhance their resistance against V. splendidus infection till to pediveliger stage, and promote their following growth and development. All the results provided a theoretical basis for solving poor disease resistance and high mortality of larvae in oyster seedling industry.

Competition is the main factor, compared to non-lethal threat of predation, affecting life-history traits of Aedes aegypti (Diptera, Culicidae) mosquitoes.

Mosquito larvae coexist with different species of predators in small freshwater bodies. These environments can vary widely in terms of the amount of food and larval densities. Our objective was to understand the impacts of the non-lethal threat of predation, combined with competition effects on Aedes aegypti development and adult life cycle. We tested the effects of two types of non-lethal threat of predation, divided into nine combinations of larvae densities and resource availability on Ae. aegypti larvae. We measured their isolated and combined impacts on larval mortality rate, time to reach pupation, adult size, and lifespan. Real non-lethal threat of predation resulted in a higher larval mortality (71%), compromised the sizes of the individuals (-7%) and prolonged the lifespan (19%) compared to control. Simulated treatment, the anti-predation behaviors led to an increase in larval mortality (21%) compared to the control treatment. Accelerating the larva's development time (-14%) to escape from a dangerous environment severely compromised the size of the individuals (-23%) and increased adult life (10%). The scarcity of food resources, high levels of competition, and non-lethal threat of predation as well, are effective stressors for Ae. aegypti. We observed that resources (food and space) were more important than predation risk for the fitness of the survivors. However, these stressors, individually or combined, can result in high larvae mortality, altered larval development and pupation, affecting the emergence of the adults, with small sizes and altered lifespan, ultimately reducing the fitness of the individuals. © 2024 Society of Chemical Industry.

Toxicity of pesticide cocktails in amphibian larvae: understanding the impact of agricultural activity on aquatic ecosystems in the Salado River basin, Argentina

Aquatic communities are increasingly exposed to complex mixtures of contaminants, mainly pesticides due to the impact of agricultural activity. The aim of this study was to evaluate the toxicity of an eight-pesticide cocktail on larvae of the South American common toad, Rinella arenarum. The cocktail represents a realistic mixture of insecticides (cypermethrin, chlorpyrifos and lambda-cyhalothrin), herbicides (glyphosate, glufosinate ammonium, prometryn and metolachlor), and a fungicide (pyraclostrobin) previously found in aquatic organisms (Prochilodus lineatus) from the Salado River Basin, an area with strong agricultural pressure. Computational simulations through the Density Functional Tight-Binding method indicated a strong spontaneous trend toward the formation of the cocktail, suggesting that it may act as a novel xenobiotic entity in the environment. The cocktail effects were evaluated in early-developing and premetamorphic larvae, at feasible concentrations found in real scenarios. The mixture led to high mortality and teratogenicity in early-developing larvae. Premetamorphic larvae showed endocrine disruption, oxidative stress, and impairments in detoxification and hepatic functioning. Neurotoxicity, genotoxicity, cardiotoxicity and high mortality under stress conditions were also observed in exposed larvae. This novel evaluation highlights the ecotoxicological risk for aquatic organisms exposed to complex mixtures and underscores the need to consider cocktail effects in studies regarding ecosystems health.

Larvicidal activity, molecular docking, and molecular dynamics studies of 7-(trifluoromethyl)indolizine derivatives against Anopheles arabiensis.

A novel series of 7-(trifluoromethyl)indolizine derivatives (4a-4n) was synthesized using a 1,3-Dipolar cycloaddition reaction. Structure elucidation of the synthesized compounds was done using various spectroscopic techniques. Compounds were assessed for their larvicidal activity against Anopheles arabiensis. Exposure of Anopheles arabiensis larvae to a series of 7-(trifluoromethyl)indolizine at 4µg/mL for 24 and 48h resulted in moderate to high larval mortality rates. Among them, compounds 4b, 4a, 4g, and 4m exhibited the most promising larvicidal activities, with mortality rates of 94.4%, 93.3%, 80.00%, and 85.6%, respectively, compared to controls, Acetone and Temephos. The structural activity relationship analysis of the evaluated compounds revealed that substitution with halogens or electron-withdrawing groups (CN, F, Cl, Br) at the para position of the benzoyl group is crucial for achieving promising larvicidal activity. Molecular docking studies were carried out involving six potential larvicidal target proteins to predict how the tested compounds might work. Compounds 4a and 4b showed strong binding to the Mosquito Juvenile Hormone-Binding Protein (5V13). Molecular dynamics (MD) simulations confirmed the stability of the protein-ligand complexes over the simulation period, reinforcing the reliability of the docking results. Compounds 4a and 4b also exhibited favourable ADMET profiles, showing high oral bioavailability, good permeability, moderate distribution, low plasma protein binding, sufficient metabolic stability, efficient renal clearance and low toxicity. Given the crucial role of Juvenile Hormone in regulating gene expression and developmental pathways through receptor interactions, compounds 4a and 4b show promise as inhibitors of this protein. Inhibiting this process could hinder larval growth and reproduction, presenting a promising approach for early-stage mosquito larvicidal activity. Therefore, compounds 4a and 4b represent lead candidates for further optimization and the development of new larvicidal agents.

Effects of elevated CO2 and temperature on the performance of a diet specialized neotropical herbivore and its host plant

AbstractLittle is known about the potential responses of ecologically specialized tropical species to atmospheric change and global warming. In 2 years of greenhouse experiments simulating climate change impacts, we quantified the effects of mean ambient temperature, elevated temperature (Te), current ambient CO2 concentration ([CO2]a), and doubled CO2 concentration ([CO2]e) on biomass, growth rate, and foliar chemistry of the morning glory vine, Camonea umbellata. In addition, we measured the impacts of climate change simulations on the performance and survival of the tortoise beetle, Acromis sparsa, which feeds exclusively on C. umbellata. Full‐sib A. sparsa larval broods were divided into cohorts. Each cohort was placed in one of four temperature‐CO2 controlled chambers and fed leaves grown in their respective treatments. Vines growing in [CO2]e more than doubled their biomass and their leaves expanded faster. The [CO2]a and Te treatments interacted to yield the greatest foliar [C]. Vines in [CO2]e and Te had the greatest C:N ratios, the lowest availability of nitrogen, and highest larval mortality. Whereas pupae were smaller and suffered lower survival in Te, pupal mass and survival increased in both the [CO2]e and Te treatments. Overall, the simultaneous elevation of both [CO2] and temperature caused declines in host quality, larval survivorship, and pupal mortality that were not observed when only one climate factor was altered. Based on this first tropical experimental study, we predict that C. umbellata will benefit from elevation of temperature and atmospheric [CO2] by altering its foliar chemistry to the detriment of its diet‐specialized herbivore enemy.Abstract in Spanish is available with online material.

Developmental ecology in embryos of an estuarine pupfish endemic of the Yucatan peninsula: Survival out of water, metabolic depression, and asynchronous hatching.

Theory predicts that drought-resistant embryos with extended incubations are evolutionarily favored in environments with high mortality of larvae but safe for eggs. Here, we experimentally test, under common garden conditions, the effect of three incubation temperatures and media on embryonic developmental length, extended incubation out of the water, survival, metabolic rate, and hatching dynamics in the estuarine pupfish Garmanella pulchra. We also described the morphological changes of embryonic cortical structures related to air exposure. We found that embryos incubated out of water in low and medium temperatures present an extended incubation period beyond their hatching capability with a deep metabolic depression. Also, these embryos exhibited a hatching asynchrony not related to water availability. Embryos incubated at high temperatures did not show extended incubation, with decreased probability of survival out of water. Our morphological observations of the embryonic cortical structures reveal that the perivitelline space and hair-like filaments buffer the deleterious drought effects. Our results reveal that G. pulchra possesses life-history traits typical of two separate phenomena: delay hatching and diapause; supporting a true continuum between them, rather than a dichotomy. The evolution of these traits may respond to aerial exposure during low tides in the estuaries of Yucatán they inhabit.

Zootechnical performance, feed efficiency and larval composition of Zophobas morio (Coleoptera: Tenebrionidae) fed with poultry litter-based diets

Abstract The yellow mealworm, super worm, and house cricket are among the most widely produced insects, with high feed conversion efficiency. However, their nutritional composition and development rate generally vary with environmental conditions. From an economic point of view, rearing conditions such as diet, temperature, and development time; insect performance such as mortality and nutritional value are the most important factors. In order to assess the development, growth, feed conversion efficiency, and chemical composition of Zophobas morio (Coleoptera: Tenebrionidae) larvae fed with diets containing poultry litter, an experiment was conducted. Five diets with varying levels of poultry litter inclusion (0, 25, 50, 75, and 100%) were used to replace the control diet (broiler feed), with five replicates containing 25 larvae per sample unit. Larval growth and development were assessed, and the chemical compositions of both the diet and Z. morio larvae were determined. Significant differences were observed among treatments for development time, survival rate, pupa weight, adult weight of Z. morio subjected to different poultry litter-based diets, as well as in the nutritional index bioassay. The longest development time and the highest larval mortality were observed in the 100% poultry litter-based diets. The highest percentage of crude protein in larval meal (%CP) was obtained with the addition of 25 and 50% poultry litter and was lowest with 100%. The incorporation of poultry litter into the diet of Z. morio has a multifaceted impact on growth and feed conversion efficiency. Elevated levels of poultry litter inclusion led to an extension in development time, yet food conversion efficiency attains optimization with an inclusion rate of up to 50%. Consequently, the decision regarding the proportion of poultry litter in the diet should be carefully weighed, taking into account breeding objectives, efficiency considerations, and cost factors. This ensures the attainment of an optimal balance between larval growth and nutritional efficiency.

Assessing the In Vitro Individual and Combined Effect of Arthrobotrys oligospora and A. musiformis (Orbiliales) Liquid Culture Filtrates against Infective Larvae of the Sheep Blood-Feeding Nematode Haemonchus contortus (Trichostrongylidae).

Arthrobotrys species are nematophagous fungi that secrete extracellular nematocidal products (ECP). The individual and combined effects of ECP from Arthrobotrys oligospora (Ao) and A. musiformis (Am) growth in liquid media against Haemonchus contortus L3 (HcL3) were assessed. The isolation, morphological (MI) and molecular identification (Mol-I), assessment of nematocidal activity (NA) of fungal liquid culture filtrates (LCF) in two liquid media alone and in combination and the myco-compound profile identification (MCP) were performed. The MI suggested that the fungi corresponded to the species Ao and Am. This result was confirmed by PCR analysis followed by sequencing, alignment and a phylogenetic analysis. Likewise, the highest Hc mortalities were 91.4% with individual LCF of Am and 86.2% with those of Ao at the highest concentration (100 mg/mL) in Czapek-Dox Broth. The combination of both LCF resulted in a similarly high larval mortality with no statistical differences in relation to individual activity (p > 0.05). The MCP showed the presence of alkaloids in both fungi. Coumarins, sterols and saponins were found only in Ao. Both fungi produced ECP with a high NA that could be identified and assessed in future studies as potential natural anthelmintic compounds.

Roles of a newly lethal cuticular structural protein, AaCPR100A, and its upstream interaction protein, G12-like, in Aedes aegypti

Mosquitoes form a vital group of vector insects, which can transmit various diseases and filarial worms. The cuticle is a critical structure that protects mosquitoes from adverse environmental conditions and penetration resistance. Thus, cuticle proteins can be used as potential targets for controlling the mosquito population. In the present study, we found that AaCPR100A is a structural protein in the soft cuticle, which has flexibility and elasticity allowing insects to move or fly freely, of Aedes aegypti. RNA interference (RNAi) of AaCPR100A caused high mortality in Aedes aegypti larvae and adults and significantly decreased the egg hatching rate. Transmission electron microscopy (TEM) analysis revealed that the larval microstructure had no recognizable endocuticle in AaCPR100A-deficient mosquitoes. A yeast two-hybrid assay was performed to screen proteins interacting with AaCPR100A. We verified that the G12-like protein had the strongest interaction with AaCPR100A using yeast two-hybrid and GST pull-down assays. Knockdown of G12-like transcription resulted in high mortality in Ae. aegypti larvae, but not in adults. Interestingly, RNAi of G12-like rescued the high mortality of adults caused by decreased AaCPR100A expression. Additionally, adults treated with G12-like dsRNA were found to be sensitive to low temperature, and their eggshell formation and hatching were decreased. Overall, our results demonstrated that G12-like may interacts with AaCPR100A, and both G12-like and AaCPR100A are involved in Ae. aegypti cuticle development and eggshell formation. AaCPR100A and G12-like can thus be considered newly potential targets for controlling the Ae. aegypti mosquito.

Efficacy of entomopathogenic fungi for the management of Trogoderma granarium Everts on wheat grains

Four species of Trichoderma genius (T. harzianum, T. citrinoviride, T. viride and T. asperellum) and Metarhizium anisopliae were assessed for their effects on larval mortality and progeny production of Trogoderma granarium Everts (Coleoptera: Dermestidae). The fungal strains were tested at 2.0 × 106, 2.0 × 107, 2.0 × 108 and 2.0 × 109 spores/kg. The larval mortality of T. granarium was recorded after 7, 14 and 21 days of treatment. The emerged adults (F1) from treated larvae were counted after 30 days of treatment, while F2 and grain weight loss were examined after 80 days of treatment. In general, increasing of spore concentration of all fungal strains enhanced the larval mortality. After 21 of treatment, all fungal strains induced notable larval mortality particularly at the highest concentration (2.0 × 109 spores/kg) except T. viride (19.4%). The highest larval mortality was observed in treatments with M. anisopliae (82.1%) and T. citrinoviride (69.0%) at 2.0 × 109 spores/kg, respectively. Furthermore, all spore concentrations of tested fungi decreased the emerged adults (F1 and F2) with M. anisopliae, T. citrinoviride and T. harzianum being the most potent, particularly at 2.0 × 109 spores/kg. After 80 days, M. anisopliae and T. citrinoviride at 2.0 × 109 spores/kg induced the highest protection of wheat grains against larvae of T. granarium with weight loss of 0.7 and 1.3%, respectively. The outcomes of the present study indicate that the two fungi, M. anisopliae and T. citrinoviride are highly effective seed protectants against T. granarium and could be applied in the integrated pest management programs (IPM) of khapra beetle as effective biological control agents.

Struggling to survive: A comparison of Vanessa cardui larval survivorship on putative host plants

AbstractThe painted lady butterfly (Vanessa cardui) is a generalist herbivore with a global distribution. In North America, over 100 species have been identified as V. cardui larval host plants. This cosmopolitan species is commercially available in all life stages and can be reared on an artificial diet. As a result, V. cardui commonly serves as a model organism for education and outreach. However, gaps in our knowledge remain with regard to V. cardui ecology and larval host plant suitability for supporting complete development of larvae to reproductive adulthood. In a laboratory setting, we tested host plant suitability of five reported host plants commonly found in California sage scrub ecosystems by assessing survival of V. cardui caterpillars in its entirety. We also assessed how commercially sourced larvae survived on artificial diet: both raising the caterpillars exclusively on artificial diet as well as switching diets from artificial to thistle (a commonly reported host plant) and switching from thistle to artificial diet. We found that all commercially sourced caterpillars exhibited both high larval mortality and low pupation rates when reared on host plants, although larvae reared on Malacothamnus fasciculatus and Sphaeralcea ambigua survived longest. Moreover, M. fasciculatus and S. ambigua were the only host plants tested that supported successful pupation. This contrasted with 63% higher survival of wild‐collected larvae. Our findings suggest that commercially obtained V. cardui may struggle to utilize wild host plants and that future investigations into host plant suitability in wild populations are needed. Additionally, future research using commercial larvae should consider the implications for interpreting host plant suitability as larvae may exhibit adaptations to artificial diet or the loss of adaptation to consuming plant material.

Larvicidal Efficiency of Citrus maxima against Aedes aegypti and Aedes albopictus

Mosquitoes transmit serious human diseases, causing millions of deaths every year and developed resistance to chemical insecticides resulting in rebounding vectorial capacity. Plants are the alternative sources of mosquito control agents. In the present study, various concentrations of 10, 20, 30, 40, and 50 mg of Citrus maxima (Brunnich) Merr. were tested against the third instar larvae of Aedes aegypti (Linn) and Aedes albopictus (Skuse). The highest larval mortality was found in the extract of C. maxima against third-instar larvae with P value = 0.0287. This study concluded that the increase in concentration of C. maxima extract increased the mortality of Aedes aegypti and Aedes albopictus.

Early ontogeny of the African bony-tongue (Heterotis niloticus) and the effect of Artemia and rotifer live feeds on larval digestive enzyme activity and performance

High larval and fry mortality has been a major challenge to the commercial culture of the African bony-tongue (Heterotis niloticus). Research indicates inadequate feeding to be a possible cause of the high mortality rates of larvae/fry grown in captivity. An experiment was conducted to first describe the developmental stages ofH. niloticuseggs and larvae until schooling, 6 days after hatching (DAH), at 26°C. The morphological development, survival, growth (weight gain and length), and swimming behavior of the larvae were monitored. A follow-up feeding trial was conducted withH. niloticusfry from 6 to 27 DAH involving four treatments: feeding withArtemianauplii; a combined feed of 50%Artemianauplii and 50% rotifers (w/w); feeding with rotifers; and no feeding. The activities of three digestive enzymes (trypsin, lipase, and amylase) were assessed in fry exposed to the live feed treatments. At hatching, the larvae had a large, vascularized yolk sac filled with yolk platelets, which occupied approximately one-third of the total body length of the abdominal cavity. Yolk platelet reabsorption started at 1 DAH and was complete at 6 DAH. Metamorphosis was completed at 6 DAH, and schooling behavior was observed. For the feeding trial, the fry that receivedArtemianauplii underwent the fastest development until 27 DAH. Similarly, the survival rate was considerably higher in fry fed withArtemianauplii than in those that received a combination ofArtemianauplii and rotifers or only rotifers. Overall, the survival rates for fry fed withArtemianauplii and larvae co-fed were 72.7% ± 0.9% and 41.2% ± 3.3%, respectively. Fry did not survive until 27 DAH in the rotifer-fed and unfed treatment groups. Digestive enzyme activity differed significantly (p< 0.05) between treatments and sampling dates. All digestive enzymes were detectable at the onset of mouth opening (5 DAH) and fluctuated through the 7 and 9 DAH. Lower enzyme activity was observed when the fry were subjected to starvation or the rotifer treatment. Results indicate thatArtemianauplii is the best live feed organism forH. niloticuslarviculture.

Bazı Uçucu Yağların Kimyasal Yapısı ve Laboratuvar Koşullarında Ceratitis capitata ‘nın (Wiedemann) (Diptera: Tephritidae) Larva Canlılığı Üzerindeki İnsekitisidal Etkisi

Aims:The Mediterranean fruit fly, medfly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae) is a serious pest of many fruits and vegetables. This study was conducted to determine antifeeding and insecticidal activities of some essential oils extracted from Pelargonium graveolens (Geraniaceae), geranium, Lavandula intermedia Mill. (Lamiaceae), lavender, Nigella sativa L. (Ranunculaceae) black cumin, and Laurus nobilis L. (Lauraceae), laurel, against second instars of the Medfly on artificial diet.
 Material and Methods:The essential oils were obtained by Clevenger-type water distillation and a laboratory reared medfly colony was used in the study. Each tested essential oils with 6 different doses were distributed over the artificial diet in a Petri dish having 20 larvae. All experiments were performed under laboratory conditions of 23±1°C, 50% RH and 16: 8 (L:D) photoperiods. Probit MsChart was used to estimate LC50 and LC90 values of the tested essential oils. GGE Biplot analyzes were created with the larval mortality based on the different essential oil doses.
 
 Results: As results, the highest larval mortality was determined with addition of laurel and black cumin oils into the diet. The highest mortality was detected in black cumin oil at the lowest concentrations. Based on GGE Biplot analyzes, the essential oil of black cumin has observable larvicidal properties. 
 Discussion: The results were provided fundemental information about insecticidal and antifeeding properties of the medfly in laboratory.
 Conclusions:Further studies needed to integrate sustainable management programmes with natural insecticides against the medfly larvae.

Control of phytopathogen organisms using bioactive compounds contained in orange wax

AbstractOrange wax (OW) is a waxy waste generated in the essential oil purification process (winterization). It has no commercial value. It is discarded or burnt, polluting the environment. The present study demonstrates the possibility of applying it to control phytopathogens. The ethanol‐soluble bioactive compounds of OW were identified and quantified. The effect of orange wax extracts on the growth inhibition of phytopathogenic fungi (Fusarium oxysporum and Colletotrichum gloeosporioides) and Spodoptera frugiperda larvae was evaluated. High‐performance liquid chromatography–mass spectrometry (HPLC‐MS) and gas chromatography–mass spectrometry (GC–MS) quantified 22 bioactive compounds representing 23% (w/w) of OW. Terpineol, limonene, and eremophilene were the most abundant bioactive substances in this material. The distortion of the radial growth of the fungal hyphae was measured in a potato dextrose agar (PDA), and all tested concentrations (6.3, 12.5, 25, 50, and 100 mg/mL) inhibited the radial expansion of C. gloeosporioides. In contrast, only the OW extracts at 50 and 100 mg mL−1 inhibited F. oxysporum. Spray treatments were performed using a Potter spray tower on third‐instar Spodoptera frugiperda larvae. Spraying OW extract at 50 mg mL−1 for 10 s caused the highest larval mortality. However, third‐instar larvae were attracted to consume corn leaves moistened with extract at lower concentrations. Up to 93% of the first‐instar larvae died after eating corn leaves moistened with the extract at 50 mg mL−1. As far as the authors are aware, this is the first scientific report about the use of bioactive compounds of orange wax to control fungi. Spodoptera frugiperda could form the basis for the manufacture of new biopesticides.

The effects of temperature on pathogenicity of entomopathogenic fungi for controlling larval populations of the European grapevine moth (Lobesia botrana) (Lepidoptera: Tortricidae)

The European grapevine moth (Lobesia botrana) is currently the most damaging pest in many viticultural regions of South and Central Europe and is highly responsible for the development of secondary infections by Botrytis cinerea to both table and wine grapes. Entomopathogenic fungi (EPF) are considered as substantial biological control agents of insect pests. However, they are able to infect their hosts only under a narrow spectrum of environmental conditions. In this study, the effects of temperature on pathogenicity of EPF for the control of L. botrana are reported. Among a collection of 33 EPF isolates, six virulent isolates were selected for the experiments. The bioassays took place at controlled conditions, under the temperature regimes of 20, 22.5, 25, 27.5, 30, and 32.5 °C. A conidial concentration of 1 × 108 conidia/ml was prepared for each isolate, and fungi were applied by spraying them on third-instar larvae (L3) of L. botrana. Both factors, fungal isolate and temperature, significantly affected mortality of L. botrana. Most fungal isolates exhibited their highest larval mortality rates seven days post-treatment at the temperatures of 25 and 27.5 °C. Some isolates caused considerably high larval mortality that exceeded 90% under specific temperature conditions. Overall, EPF showed great potential as bio-control agents for controlling larval populations of L. botrana.

In-vitro Study to Determine the Dose of Beauveria bassiana for Management of Spodoptera frugiperda

Pest insects known as fall army worm (Spodoptera frugiperda) can feed on over 80 different types of crops. It has the potential to significantly reduce the yields of major cereal crops including oats (Avena sativa L.). The oat crop is one of the preferred hosts of this pest. This fodder crop is directly used as animal feed so, chemical control of pests is not preferable. One of the alternatives for the management of fall army worm on fodder oats is the use of microbials which have some advantages over the use of chemical insecticides, as they are highly host specific with low environmental contamination. Beauveria bassiana is a fungus and can be used to control this pest. So, the present study was undertaken to standardize the dose of Beauveria bassiana for management of fall army worm. The virulence test was conducted against third instar larvae of S. frugiperda. The larvae were treated with three doses of B. bassiana of 1×1012spore/ml concentration i.e.,1, 2, and 3ml/L, by dipping them for 30 seconds in 10 ml of suspension. In case of control, larvae were treated with distilled water containing a drop of Tween- 80 (0.02%). At 24 hours after treatment, the differences in the mean larval mortality among different doses of B. bassiana were non-significant. However, at 48 and 72 hours after treatment, the differences in the mean larval mortality among different doses of B. bassiana were significant. Highest larval mortality was recorded at 48 and 72 hours after treatment in 3 ml/L (29.39 and 86.67%, respectively).

Survival of Nematode Larvae after Treatment with Eugenol, Isoeugenol, Thymol, and Carvacrol.

Helminthiases inflict annual losses on the meat and dairy livestock industries. The commonest species of ruminant parasites are the nematodes: Strongyloides papillosus and Haemonchus contortus, which lay eggs in the intestine and enter the feces. There, the eggs develop into larvae, which when voided with the feces crawl onto plants. In our experiment, we evaluated the survivability of the noninvasive and invasive (L1-2 and L3, respectively) larvae of S. papillosus, H. contortus (L3), and Muellerius capillaris (L1) in vitro by subjecting each to natural compounds present in the essential oils of many plants. In the experiment, we used aqueous emulsions of eugenol, isoeugenol, thymol, and carvacrol. Administering 1% concentrations of those compounds killed 100% of the nematode larvae following 24 h of exposure. Thymol, eugenol, and isoeugenol at a concentration of 0.1% also caused high larvae mortality (over 96%). Continuous usage of synthetic anthelmintic drugs in veterinary medicine has led to the parasites developing resistance, thus, a search for novel nematicidal drugs is required. Eugenol, isoeugenol, thymol, and carvacrol are promising compounds against nematodes. However, additional research is required regarding peculiarities in their actions toward the bodies of mammals and parasitic nematodes.