Μg Larva Research Articles

New liposidomycin congeners produced by Streptomyces sp. TMPU-20A065, anti-Mycobacterium avium complex agents with therapeutic efficacy in a silkworm infection model

Three new liposidomycin congeners (1, 2, and 4), together with 14 known liposidomycins (3 and 5–17), were isolated from the culture broth of Streptomyces sp. TMPU-20A065 as anti-Mycobacterium avium complex agents. The structures of liposidomycins were elucidated by spectroscopic analyses, including NMR and MS. Compounds 1, 2, and 4 belong to type-I liposidomycin-containing sulfate groups and methylglutaric acid, each with a different acyl side chain in the structure. Compounds 1–17 exhibited in vitro anti-M. avium and M. intracellulare activities with MIC values ranging between 2.0 and 64 μg ml−1. Furthermore, 1–17 exerted potent therapeutic effects in an in vivo-mimic silkworm infection model with ED50 values ranging between 0.12 and 3.7 μg larva−1 g−1.

The Essential Oil from the Resurrection Plant Myrothamnus moschatus Is Effective against Arthropods of Agricultural and Medical Interest.

This work aimed to evaluate the chemical composition, insecticidal and acaricidal potential of the essential oil (EO) obtained from the resurrection plant Myrothamnus moschatus (Baill.) Baill. (Myrothamnaceae) from Madagascar. The EO bioactivity was evaluated against selected arthropod pests and vectors of agricultural and public health relevance. The most abundant volatile compounds were trans-pinocarveol (37.7 ± 4.2%) and pinocarvone (20.8 ± 3.1%), similar to the EO of the chemotype collected from the same region. Lethal concentrations (LC50) or doses (LD50) from acute toxicity tests were estimated for Musca domestica (L.) adults at 22.7 µg adult-1, for Spodoptera littoralis (Boisduval) larvae at 35.6 µg larva-1, for Culex quinquefasciatus (Say) at 43.6 µg mL-1, for adults of Metopolophium dirhodum (Walker) at 2.4 mL L-1, and for adults of Tetranychus urticae (Koch) at 1.2 mL L-1. The good insecticidal and acaricidal activities determined in this work may open a new perspective on the use of this plant as a source of botanical insecticide ingredients. The exploitation of this species could also be important for the African economy, helping local farmers cultivating this plant.

Essential Oils from Cameroonian Aromatic Plants as Effective Insecticides against Mosquitoes, Houseflies, and Moths.

Recently, spices have attracted the attention of scientists and agrochemical companies for their potential as insecticidal and acaricidal agents, and even as repellents to replace synthetic compounds that are labeled with detrimental impacts on environment and human and animal health. In this framework, the aim of this study was to evaluate the insecticidal potential of the essential oils (EOs) obtained from three Cameroonian aromatic plants, namely Monodora myristica (Gaertn.) Dunal, Xylopia aethiopica (Dunal) A. Rich., and Aframomum citratum (J. Pereira) K. Schum. They were produced by hydrodistillation, with yields of 3.84, 4.89, and 0.85%, respectively. The chemical composition was evaluated by GC-MS analysis. The EOs and their major constituents (i.e., geraniol, sabinene, α-pinene, p-cymene, α-phellandrene, and β-pinene) were tested against the polyphagous moth pest, i.e., Spodoptera littoralis (Boisd.), the common housefly, Musca domestica L., and the filariasis and arbovirus mosquito vector, Culex quinquefasciatus Say. Our results showed that M. myristica and X. aethiopica EOs were the most effective against M. domestica adults, being effective on both males (22.1 µg adult−1) and females (LD50: 29.1 µg adult−1). The M. myristica EO and geraniol showed the highest toxicity on S. littoralis, with LD50(90) values of 29.3 (123.5) and 25.3 (83.2) µg larva−1, respectively. Last, the EOs from M. myristica and X. aethiopica, as well as the major constituents p-cymene and α-phellandrene, were the most toxic against C. quinquefasciatus larvae. The selected EOs may potentially lead to the production of cheap and effective botanical insecticides for African smallholders, although the development of effective formulations, a safety evaluation, and an in-depth study of their efficacy on different insect species are needed.

Insecticidal Activity of Four Essential Oils Extracted from Chilean Patagonian Plants as Potential Organic Pesticides.

Patagonia is a geographical area characterized by a wide plant biodiversity. Several native plant species are traditionally used in medicine by the local population and demonstrated to be sources of biologically active compounds. Due to the massive need for green and sustainable pesticides, this study was conducted to evaluate the insecticidal activity of essential oils (EOs) from understudied plants growing in this propitious area. Ciprés (Pilgerodendron uviferum), tepa (Laureliopsis philippiana), canelo (Drimys winteri), and paramela (Adesmia boronioides) EOs were extracted through steam distillation, and their compositions were analyzed through GC–MS analysis. EO contact toxicity against Musca domestica L., Spodoptera littoralis (Boisd.), and Culex quinquefasciatus Say was then evaluated. As a general trend, EOs performed better on housefly males over females. Ciprés EO showed the highest insecticidal efficacy. The LD50(90) values were 68.6 (183.7) and 11.3 (75.1) µg adult−1 on housefly females and males, respectively. All EOs were effective against S. littoralis larvae; LD50 values were 33.2–66.7 µg larva−1, and tepa EO was the most effective in terms of LD90 (i.e., <100 µg larva−1). Canelo, tepa, and paramela EOs were highly effective on C. quinquefasciatus larvae, with LC50 values < 100 µL L−1. Again, tepa EO achieved LD90 < 100 µL L−1. This EO was characterized by safrole (43.1%), linalool (27.9%), and methyl eugenol (6.9%) as major constituents. Overall, Patagonian native plant EOs can represent a valid resource for local stakeholders, to develop effective insecticides for pest and vector management, pending a proper focus on their formulation and nontarget effects.

Chemical Composition and Broad-Spectrum Insecticidal Activity of the Flower Essential Oil from an Ancient Sicilian Food Plant, Ridolfia segetum

Several species of the family Apiaceae are aromatic herbs that produce essential oils usable on an industrial scale for pharmaceutical, cosmetic, and food purposes. In particular, some essential oils, such as green insecticides for example, may replace synthetic insecticides, keeping most of their efficacy and avoiding environmental pollution or human poisoning. In the present study, we explored the insecticidal potential of Ridolfia segetum (L.) Moris essential oil (EO) against three different pests: Culex quinquefasciatus Say, Musca domestica L., and Spodoptera littoralis (Boisduval). For this purpose, the EO was obtained by hydrodistillation of flowers and its composition was achieved by gas chromatography/flame ionization detection (GC/FID) and gas chromatography/mass spectrometry (GC/MS). This EO was rich in α-phellandrene (49.3%), β-phellandrene (9.2%), terpinolene (20.7%), and piperitenone oxide (5.9%). Concerning the mosquitocidal efficacy, the EO showed noteworthy toxicity against C. quinquefasciatus 3rd instar larvae, with a LC50 = 27.1 µL L−1 and LC90 = 42.5 µL L−1. Regarding M. domestica, a different toxicity of the R. segetum EO was found on male and female flies, calculating LD50 values of 10.5 and 50.8 µg adult−1, respectively. The EO was also toxic to S. littoralis 3rd instar larvae, achieving LD50 and LD90 values of 37.9 and 99.6 µg larva−1, respectively. Overall, this flower EO, extracted from a traditional Sicilian food plant, merits further investigation for the development of green insecticide formulations to be used in real world conditions, pending a careful assessment of non-target toxicity on beneficial organisms.

Phenolic monoterpene-rich essential oils from Apiaceae and Lamiaceae species: insecticidal activity and safety evaluation on non-target earthworms

The genera Oliveria (Apiaceae), Thymus and Satureja (Lamiaceae) encompass important medicinal and aromatic plants of Iran, which contains noteworthy amounts of essential oils (EOs) that are worthy of exploitation on an industrial level, including pesticide formulation development. In this study, the efficacy of five EOs obtained from O. decumbens, T. daenensis, S. sahendica, S. khuzistanica and S. rechingeri , was evaluated against three insects of economic relevance, the mosquito Culex quiquefasciatus , the housefly Musca domestica, and the moth Spodoptera littoralis . Potential non-target effects of these EOs were assessed on earthworms, Eisenia fetida . The chemical composition of the five EOs was determined by gas chromatography (GC-FID) and gas chromatography-mass spectrometry (GC-MS). EOs from O. decumbens, T. daenensis, S. khuzistanica and S. rechingeri were characterized by oxygenated monoterpenes (70.3, 91.6, 89.7 and 89.4%, respectively), whereas the EO from S. sahendica was dominated by monoterpene hydrocarbons (63.3%) followed by oxygenated monoterpenes (35.3%). In all cases, the biogenetically related thymol (0.3–36.7%), carvacrol (1.3–89.6%), p -cymene (2.1–13.7%) and γ-terpinene (1.5–41.3%) were found as the characteristic volatile compounds of the five EOs, with high quantitative variation according to the species considered. The five EOs exhibited high toxicity on the three target insects, with LC 50 /LD 50 values in the range of 16.2–29.3 µg mL -1 , 18.1–48.1 µg adult -1 and 7.4–23.1 µg larva -1 on C. quinquefasciatus, M. domestica and S. littoralis , respectively. Toxicity of the five EOs on non-target soil invertebrates was minimal, leading to no or very low (5.0–7.5%) mortality on E. fetida when tested at the concentration of 200 mg kg -1 , at variance with the positive control α-cypermetrin, which caused 100% mortality at 0.1 mg kg -1 of soil. Overall, our study discloses new opportunities to farmers growing these Iranian endemic plants, shedding light on the potential of these EOs to formulate effective and eco-friendly insecticides.

Exploring essential oils of Slovak medicinal plants for insecticidal activity: The case of Thymus alternans and Teucrium montanum subsp. jailae

Thymus alternans and Teucrium montanum subsp. jailae are medicinal and aromatic plants, typical of Slovakian flora, producing bioactive essential oils. In the present study, we evaluated the insecticidal potential of the essential oils, obtained by hydrodistillation from the plant aerial parts and analysed by GC-MS, as insecticidal agents. For the purpose, they were assayed against three insect species acting as agricultural pests or vectors of medical relevance, such as the common housefly, Musca domestica L., the lymphatic filariasis vector, Culex quinquefasciatus and the Egyptian cotton leafworm Spodoptera littoralis; α-cypermethrin was tested as positive control. The two essential oils exhibited a different chemical profile, with monoterpenes and sesquiterpenes being the main fractions in the essential oils from Th. alternans and T. montanum subsp. jailae, respectively. Insecticidal tests showed that the T. montanum essential oil was effective against S. littoralis (LD50(90) = 56.7 (170.0) μg larva−1) and Cx. quinquefasciatus larvae (LC50(90) = 180.5 (268.7) mg L−1), whereas T. alternans essential oil displayed good toxicity against M. domestica adults (LD50(90) = 103.7 (223.9) μg adult−1). Overall, our results add useful knowledge about the potential of Slovakian flora as a source of botanicals for the eco-friendly management of insect pests and vectors.

Ascaridole-rich essential oil from marsh rosemary (Ledum palustre) growing in Poland exerts insecticidal activity on mosquitoes, moths and flies without serious effects on non-target organisms and human cells

Marsh rosemary (Ledum palustre, Ericaceae) has been widely used in the traditional medicine of various regions worldwide, and as insect repellent. Little is known on its essential oil insecticidal potential. This study explored the insecticidal effects of the essential oil obtained from L. palustre growing in Poland on selected insect pests and vectors. GC-MS analysis evidenced an uncommon chemotype characterized by ascaridole (35.3% as sum of cis-ascaridole and isoascaridole) and p-cymene (25.5%). The essential oil was effective against Culex quinquefasciatus, Spodoptera littoralis and Musca domestica, showing LC50/LD50 of 66.6 mg L−1, 117.2 μg larva−1 and 61.4 μg adult−1, respectively. It was not toxic to non-target Eisenia fetida earthworms and moderately toxic to Daphnia magna microcrustaceans, over the positive control α-cypermethrin. The essential oil cytotoxicity on human keratinocytes and fibroblasts showed high IC50 values (71.3 and 84.4 μg mL−1, respectively). Comet assay data highlighted no DNA damages. Based on our findings, this essential oil, characterized by the ascaridole/p-cymene chemotype, could be a candidate for the formulation of botanical insecticides; large-scale production of green insecticides by this rare species may be assured by ex situ cultivation and biotechnological techniques.

Origanum syriacum subsp. syriacum: From an ingredient of Lebanese ‘manoushe’ to a source of effective and eco-friendly botanical insecticides

Origanum syriacum subsp. syriacum, also known as ‘Za’tar’ is an aromatic shrub native to Lebanon and cultivated in other Middle East countries. The plant leaves enjoy a high reputation as a traditional remedy against cardiovascular, respiratory and infectious diseases. In addition, they are a famous component of the Lebanese pizza (“manoushe”). Starting from its safety for humans, here O. syriacum subsp. syriacum was selected to assess the insecticidal efficacy of its leaf essential oil (EO) and its major constituent carvacrol against two key agricultural pests, namely the noctuid Spodoptera littoralis and the aphid Myzus persicae, as well as on the fly pest Musca domestica. Furthermore, the Za’tar EO potential impact on beneficial organisms such as the aphid predator Harmonia axyridis and the earthworm Eisenia fetida, which is used in the vermicomposting process, was assessed. GC–MS analysis highlighted the phenolic monoterpene carvacrol as the predominant component (83%) of Za’tar EO. Toxicity of O. syriacum subsp. syriacum EO was noteworthy, showing LC50/LD50 of 103.3 μg larva−1, 2.1 mg L−1 and 58.7 μg adult−1 on S. littoralis, M. persicae and M. domestica, respectively, which were partly consistent with those of its major component carvacrol (38.3 μg larva-1, 1.6 mL L−1 and 59.3 μg adult-1, respectively). When tested up to 3.8 mL L−1 and 200 mg kg−1 on H. axyridis and E. fetida, this EO was not toxic, at variance with α-cypermethrin, which caused 100% mortality at 1 mL L−1 and 25 mg kg−1, respectively. Taken together, these results promote carvacrol-rich Za’tar EO as a promising reservoir of green insecticides to be used for managing insect pests and vectors of economic relevance.

Insecticidal activity of the essential oil and polar extracts from Ocimum gratissimum grown in Ivory Coast: Efficacy on insect pests and vectors and impact on non-target species

Ocimum gratissimum L. (white wild basil) is an aromatic herb cultivated in Western Africa for culinary and pest control purposes. The current research evaluated the insecticidal activity of white wild basil essential oil, ethanolic and water extracts against insect pests and vectors, i.e., the tobacco cutworm Spodoptera littoralis, the housefly Musca domestica, and the filariasis vector Culex quinquefasciatus. Furthermore, the toxicity of the essential oil and polar extracts against the non-target earthworm Eisenia fetida was assessed. The chemical profiles of the essential oil and polar extracts were obtained by GC–MS and HPLC-DAD analyses. Acute toxicity experiments were conducted on larvae of C. quinquefasciatus and S. littoralis and adults of M. domestica and E. fetida, to determine the LC50 and LC90 values of the oil and polar extracts. Chronic toxicity was evaluated on S. littoralis feeding on tomato discs treated with essential oil and polar extracts. The essential oil was dominated by thymol (50.0%) and p-cymene (16.8%), whereas ethanolic and aqueous extracts were characterized by carvacrol (13%) and thymol (11%), and shikimic acid (3%) and rosmarinic acid (2%), respectively. The essential oil was significantly more active on target insects than extracts, showing LC50/LD50 of 39.6 mg L−1 on C. quinquefasciatus, 72.2 μg adult−1 on M. domestica and 30.2 μg larva−1 on S. littoralis. Furthermore, the essential oil and ethanolic extract at sublethal doses (10 and 70 μg cm-2, respectively) affected the survival of S. littoralis larvae from the third day on. White wild basil oil LD50,90 at day 5 were 2.8 and 12.3 μg cm-2. Finally, the essential oil and polar extracts were not toxic to E. fetida over the positive control α-cypermethrin. Overall, our study showed that the essential oil of white wild basil is a potential candidate as a functional ingredient in insecticidal formulations to manage agricultural moth pests and insect vectors of public importance.

Evaluation of two invasive plant invaders in Europe (Solidago canadensis and Solidago gigantea) as possible sources of botanical insecticides

Solidago gigantea and Solidago canadensis (Asteraceae) are two invasive weeds native to North America and introduced in Europe and Asia, where they are spreading quickly threatening the stability of local secondary ecosystems. These two plant invaders may represent an ideal bioresource to be exploited for production of green pesticides. Therefore, herein we evaluated the efficacy of the essential oils (EOs) obtained from their different parts, i.e. leaves, inflorescences and roots, against Culex quinquefasciatus, Spodoptera littoralis and Musca domestica. The essential oil composition was investigated by gas chromatographic–mass spectrometry (GC–MS) analysis. S. canadensis leaf EO was the most toxic to C. quinquefasciatus, with a LC50 of 89.3 μl L−1. The two most effective oils against M. domestica adults were S. canadensis leaf and flower EOs, with LD50 values of 206.9 and 207.1 μg adult−1, respectively. Three EOs highly toxic to S. littoralis were also identified, namely S. gigantea leaf EO, S. canadensis leaf EO and S. gigantea flower EO, with LD50 values of 84.5, 98.9 and 107.4 μg larva−1, respectively. Since the S. canadensis leaf EO was the only green product effective against all the tested insect pests, we selected it for non-target toxicity assays on Eisenia fetida earthworms, along with the leaf EO from S. gigantea. Both the S. canadensis and S. gigantea leaf EOs did not led to mortality of E. fetida adult earthworms, at variance with the positive control α-cypermethrin, allowing us to propose them for pest control purposes in IPM and organic farming.

Not just popular spices! Essential oils from Cuminum cyminum and Pimpinella anisum are toxic to insect pests and vectors without affecting non-target invertebrates

Each year the flavor and fragrance industry produces more than hundred thousand tons of essential oils. An important portion of this production might be directed to the fabrication of green pesticides relying on the well-documented efficacy of many essential oils against insect pests as well as vectors, besides their safety for human health and the environment. In this regard, two popular spices of economic importance, such as cumin (Cuminum cyminum) and anise (Pimpinella anisum), which are not only cheap, readily available, generally recognized as safe (GRAS), but also produces larger amounts of essential oils, are gaining interest for botanical insecticide development. Herein, we evaluated the efficacy of the essential oils obtained from seeds of these two spices on two agricultural pests, i.e., the peach-potato aphid Myzus persicae and the tobacco cutworm Spodoptera littoralis, and on two insect vectors, i.e., the common housefly Musca domestica and the lymphatic filariasis and Zika virus vector Culex quinquefasciatus. Furthermore, their safety on beneficial organisms such as the earthworm Eisenia fetida and the aphid predator Harmonia axyridis was assessed. The two essential oils, characterized by γ-terpinen-7-al (35.3%), cumin aldehyde (21.8%), and α-terpinen-7-al (15.4%), and (E)-anethole (93.0%), respectively, showed noteworthy effects against all pest targets, with anise more effective on larvae of C. quinquefasciatus (LC50 = 25.4 μl L−1) and S. littoralis (LD50 = 57.3 μg larva−1), whereas cumin was more active on adults of M. persicae (LC50 = 3.2 ml L−1) and M. domestica (LD50 = 31.8 μg adult1). Moreover, when compared at sub-lethal concentrations with the commercial insecticide α-cypermethrin, they were definitely devoid of toxic effects on E. fetida and H. axyridis. These results represent a milestone to manufacture and commercialize green formulations to be used in crop protection and to combat insect vectors of public importance.

The essential oil from industrial hemp (Cannabis sativa L.) by-products as an effective tool for insect pest management in organic crops

The inflorescences of industrial hemp (Cannabis sativa L.) represent a consistent by-product that is underutilized. Moving from the concept that this plant part has evolved as a natural weapon against phytophagous insects, secreting important secondary metabolites such as cannabinoids and volatile terpenes, herein we assayed the potential of its essential oil as a botanical insecticide. For the purpose, the essential oil was obtained by fresh inflorescences of hemp (monoecious cv. Felina 32) by steam-distillation and analysed by gas chromatography (GC-FID) and gas chromatography-mass spectrometry (GC–MS). The oil was tested against the filariasis vector Culex quinquefasciatus, the peach-potato aphid Myzus persicae, the housefly Musca domestica and the tobacco cutworm Spodoptera littoralis. To prove its harmlessness on non-target invertebrates, it was tested on the multicolored Asian lady beetle, Harmonia axyridis, and Eisenia fetida earthworms, and compared with α-cypermethrin as the positive control. The essential oil composition was dominated by monoterpene and sesquiterpene hydrocarbons, with (E)-caryophyllene (45.4%), myrcene (25.0%) and α-pinene (17.9%) as the most abundant compounds. Results from insecticidal tests showed that the essential oil from inflorescences of industrial hemp cv Felina 32 was highly toxic to M. persicae aphids (LC50 of 3.5 mL L−1) and M. domestica flies (43.3 μg adult−1), while toxicity was moderate towards S. littoralis larvae (152.3 μg larva−1), and scarce against C. quinquefasciatus larvae (LC50 of 252.5 mL L−1) and adults (LC50 > 500 μg cm−2). Contrary to α-cypermethrin, the hemp cv Felina 32 essential oil was not toxic to non-target invertebrate species, including 3rd instar larvae and adults of H. axyridis ladybugs and adults of E. fetida earthworms. Taken together our results shed light on the possible utilization of the crop residue of industrial hemp as a source of environmental-friendly botanical insecticides to be used in Integrated Pest Management and organic agriculture, particularly to manage aphid and housefly populations.

Resistance Monitoring of Emamectin Benzoate 5SG in Cotton Bollworms in Tamil Nadu

The median lethal dose (LD50) of emamectin benzoate 5SG when monitored against cotton pink bollworm Pectinophora gossypiella, it was found to be 0.4282 mg larva−1 for F1 generation and 0.1802 mg larva−1 for F9 generation. The LD95 was found to decrease from 3.3760 (F1) to 1.1564 mg larva−1 (F9). The susceptibility index to emamectin benzoate based on LD50 and LD95 was 2.37 and 3.03, respectively. With regard to number of generations required for 10-fold decrease in LD50, it was found to be 27. Considering the F9 populations as the most susceptible, the tentative discriminating dose was fixed as 1.1mg−1. Assessing the level of insecticide resistance as one time survey in summer cotton revealed that the % resistance of H. armigera in five districts ranged from 3.33 to 10.00. The highest resistance (10%) was observed at Coimbatore district. The survivability of fruit and shoot borer Earias vitella varied from 6.66 to 15.00% against emamectin benzoate, with Erode population being highly susceptible. The % survival of pink bollworm varied from 3.33 to 6.66% at 1.1μg larva−1.

Coronatin-2 from the entomopathogenic fungus Conidiobolus coronatus kills Galleria mellonella larvae and incapacitates hemocytes.

Coronatin-2, a 14.5 kDa protein, was isolated from culture filtrates of the entomopathogenic fungus Conidiobolus coronatus (Costantin) Batko (Entomophthoramycota: Entomophthorales). After LC-MS/MS (liquid chromatography tandem mass spectrometry) analysis of the tryptic peptide digest of coronatin-2 and a mass spectra database search no orthologs of this protein could be found in fungi. The highest homology was observed to the partial translation elongation factor 1a from Sphaerosporium equinum (protein sequence coverage, 21%), with only one peptide sequence, suggesting that coronatin-2 is a novel fungal protein that has not yet been described. In contrast to coronatin-1, an insecticidal 36 kDa protein, which shows both elastolytic and chitinolytic activity, coronatin-2 showed no enzymatic activity. Addition of coronatin-2 into cultures of hemocytes taken from larvae of Galleria mellonella Linnaeus (Lepidoptera: Pyralidae), resulted in progressive disintegration of nets formed by granulocytes and plasmatocytes due to rapid degranulation of granulocytes, extensive vacuolization of plasmatocytes accompanied by cytoplasm expulsion, and cell disintegration. Spherulocytes remained intact, while oenocytes rapidly disintegrated. Coronatin-2 produced 80% mortality when injected into G. mellonella at 5 µg larva-1. Further study is warranted to determine the relevance of the acute toxicity of coronatin-2 and its effects on hemocytes in vitro to virulence of C. coronatus against its hosts.

Effect of sublethal α-cypermethrin exposure on main macromolecules concentration, energy content, and malondialdehyde concentration in free-feeding Danio rerio larvae.

α-Cypermethrin (Cyp) is a synthetic insecticide used to control pests in agricultural crops and to protect human health against noxious insects; this toxic can reach aquatic systems through ground infiltration or by runoff and could affect the aquatic biota. The present study was aimed at evaluating the acute toxicity of Cyp on zebrafish (Danio rerio) exogenous feeding larvae of 10 and 20days post-fertilization (dpf), and of sublethal concentrations on only 10-dpf larvae. Proteins, lipids, carbohydrates, glycogen concentration, and total energy contents, as well as malondialdehyde (MDA) quantification, through thiobarbituric acid reactive substances, as a lipid peroxidation biomarker, were assessed in free-feeding larvae exposed to sublethal Cyp concentrations. The LC50 for 10-dpf larvae was 1.94µgL(-1), and these were more sensitive than 20-dpf larvae (3.56µgL(-1)). The amount of protein, carbohydrates, and glycogen were not significantly affected (p>0.05), but sublethal Cyp concentrations exposure caused decrement in lipids from 9.05 to 3.74µg larva(-1), as well as a reduction in MDA and in the total energy content, which affected significantly the development of this fish. Although Cyp is considered an insecticide of reduced residual effect in the environment, the present study revealed that relatively low Cyp concentrations produced significant toxic effects on exogenous feeding fish larvae, a situation that could contribute to increase deaths during this already critical developmental stage in which high mortality is observed frequently.

Effects of solvent on the toxicity of dimethoate in a honey bee in vitro larval study

There is increasing interest in the toxicity of pesticides to honey bee larvae. The solubility of pesticides in the artificial diet used to dose larvae is a key issue. There has been no full evaluation of the use of solvents to dissolve pesticides in the larval toxicity test system. Data generated for the toxicity of dimethoate to larvae using 5 and 10% acetone solvent in the diet were similar and close to the toxicity data for adults (LD50 0.1-0.3 µg bee(-1)). With only 1.5% acetone as a vehicle, the toxicity (LD50 0.6-1.14 µg larva(-1)) was lower and closer to published validation data of 1.5-3.1 µg larva(-1) (with some reported data as high as 8.8 µg larva(-1)) generated using dimethoate dissolved in water. This suggests that the solubility of the pesticide in the diet is a key factor in determining its toxicity. Consideration of the solubility of the toxic reference and test items when designing studies is important. The use of higher rates of acetone in the present study more closely represents the true exposure of larvae to pesticides that are not highly soluble in diet. Published 2012. This article is a U.S. Government work and is in the public domain in the USA.

Evaluation of β-(1 → 3, 1 → 6)-glucans and High-M alginate used as immunostimulatory dietary supplement during first feeding and weaning of Atlantic cod ( Gadus morhua L.)

Stimulation of the non-specific defence enhances the disease resistance and growth, and has good potentials as a measure for increased microbial control in juvenile production of marine fish and shellfish. So far, the most commonly used immunostimulants are β-(1 → 3, 1 → 6)-glucans, and in this study the stimulatory potential of a β-(1 → 3, 1 → 6)-glucan of marine origin, the storage polysaccharide from the marine diatom Chaetoceros mülleri, was examined. The glucan (chrysolaminaran) was extracted from cultures of C. mülleri, and used as a dietary supplement in two first feeding experiments with larvae of Atlantic cod Gadus morhua L. In one experiment the microalgal glucan was compared to the commercial yeast-glucan product MacroGard®, and in the other to an alginate with a high content of mannuronic acid (High-M alginate) isolated from Durvillaea antarctica. The stimulants were given via rotifers, and weaning to formulated feed was initiated at day 17 or 18 after hatching. The survival ± SEM at day 27 after hatching was 24.5 ± 2.0%, 14.8 ± 4.5% and 13.1 ± 1.4% for the groups fed C. mülleri-glucan, yeast glucan and for the control, respectively, in the first experiment. The group fed C. mülleri-glucan group had higher survival compared to the control ( P < 0.05) group, whereas the yeast glucan had no positive effect on the survival ( p > 0.05). The dry weights of the groups at day 27 were low, with 203.2 ± 52.2, 165.2 ± 43.4 and 198.5 ± 58.1 μg per larva for the C. mülleri-glucan, yeast glucan and control groups, respectively. In the second experiment the survival in the period of feeding formulated feed (days 18–30) were 44.6 ± 4.3%, 44.7 ± 1.3%, and 33.8 ± 4.1% survival for the C. mülleri-glucan, High-M alginate and control group, respectively. The cod larvae fed C. mülleri-glucan reached an average weight of 531.6 ± 17.2 μg at day 30, which was significantly higher ( p < 0.05) than the control group that had an average of 473.6 ± 3.5 μg. The larvae fed High-M alginate had an average weight of 470.3 ± 31.6 μg per larva at day 30, and not significantly different from the control ( p > 0.05). The early weaning to formulated diet had detrimental effect on the growth of the larvae. In both experiments the C. mülleri-glucan group was the only group showing a positive growth rate in the period of weaning to dry feed. The microbial conditions in larval gut and water were monitored with respect to total colony forming units on Marine agar, and Vibrio- and Pseudomonas-like species on selective agars (TCBS and marine Pseudomonas Agar with CFC-supplement). The larvae were rapidly colonised after hatching, but no or weak effects of the stimulants were observed on the colonisation rates or the composition. The total CFU varied from 10 1 to 10 2 CFU per μg larva after initiation of the first feeding. The percentages of Pseudomonas-like bacteria increased throughout the period, whereas the levels of Vibrio-like bacteria were low and stable. The chrysolaminaran from the diatom C. mülleri was shown to be a promising candidate for use as an immunostimulatory feed additive, and which should be further explored.

Survival, growth and feeding efficiency of Litopenaeus vannamei protozoea larvae fed different rations of the diatom Chaetoceros muelleri

The survival, development and growth of Litopenaeus vannamei protozoea (PZ) larvae were evaluated in three experiments during which they were fed five different rations of the diatom Chaetoceros muelleri, from emerging PZ I larvae until the day of change to mysis I. The total amount of food and energy supplied in 5 days to each larva ranged from 85.5 to 223.2 μg larva − 1 (ash-free dry weight) and between 2.25 and 5.88 J larva − 1 . After 2 days, the development index (DI) showed that some of the larvae fed the highest ration had reached the stage of PZ III. At the end of the experiment the mean DI ranged from 2.9 ± 0.2 to 3.2 ± 0.3 and the differences among treatments were not significant. The mean percentages of survival ranged from 35.8% to 51.7% and were not statistically different, but total length was progressively higher with increasing ration. The final values were 1.91 ± 0.54, 2.10 ± 0.53; 2.34 ± 0.36; 2.45 ± 0.37 and 2.55 ± 0.42 mm and were all statistically different with the exception of treatment 4, that was intermediate between treatments 3 and 5. However, the highest final organic weight (26.0 ± 1.7 μg larva − 1 ) was treatment 4, which was different from the values obtained with the two lower rations. Treatments 3 and 5 had intermediate values. The percentages of the food ingested did not vary with the rations supplied or with the age of the larvae and ranged from 74% to 86%. Food ingestion ( F i) was a function of the food supplied ( F s), according to the equation F i = 0.848 F s − 4.025 ( R 2 = 0.992; P < 0.001). Using the amount of food supplied and ingested with each feeding regime and the respective organic weight gains, it was calculated that the percentages of F s and F i used for body growth were 14.4% and 17.9% with the lowest ration and 6.9% and 8.3% with the highest, whereas the three intermediate rations gave similar efficiencies (9.8% to 10.2% of F s and 11.8% to 12.5% of F i). However, the weight and energy gains obtained with the highest food utilization efficiency were lower than those obtained with the higher rations, indicating that cautious overfeeding is a more convenient alternative for L. vannamei PZ culture.

Feeding rates of gilthead seabream ( Sparus aurata), larvae on microcapsules

Food consumption rates of gilthead seabream larvae from first feeding to 20 days of age were studied by feeding larvae on two types of microcapsule (hard- and soft-walled) having diameters ranging from 25 to 400 μm. Experiments were carried out in 15-litre beakers using larvae cultured with standard techniques and fed on rotifers and Artemia sp. nauplii. Ingestion rate ranged from 3 to 35 particles · larva −1 · h −1. but, except for some increase during the first days of feeding, no consistent trend was observed in relation to larval weight. However, when values were converted to volume or dry weight units, food consumption rate increased progressively with increasing larval weight. On a volume basis, the rate of this increase was different for the two microcapsule types, being generally higher with soft microcapsules, probably due to its better accessibility and to their easier progress through the digestive tract. However, on a dry weight basis the rates of food consumption were similar with both types of microcapsule because of the higher density of hard-walled microcapsules. Ingestion rates increased from 0.5–3 μg · larva −1 · h −1 at first feeding up to 18–25 μg · larva −1 μg · h −1 in older larvae. The daily specific ration increased from 0.5 to 2.5 μg food · μg larva −1 · d −1 at first feeding to a maximum of 3–5 μg food · μg larva −1 · d −1 at a larval dry weight of 70–110 μg. Above this larval weight, the specific ration tended to remain constant. The results show that gilthead seabream larvae can be transferred directly from live to microencapsulated food without appreciable variation in the incidence of feeding or in the rate of food consumption from first feeding onwards.