Knowledge about how the COVID-19 pandemic can affect aquatic wildlife is still extremely limited, and no effect of SARS-CoV-2 or its structural constituents on invertebrate models has been reported so far. Thus, we investigated the presence of the 2019-new coronavirus in different urban wastewater samples and, later, evaluated the behavioral and biochemical effects of the exposure of Culex quinquefasciatus larvae to two SARS-CoV-2 spike protein peptides (PSPD-2002 and PSPD-2003) synthesized in our laboratory. Initially, our results show the contamination of wastewater by the new coronavirus, via RT-qPCR on the viral N1 gene. On the other hand, our study shows that short-term exposure (48 h) to a low concentration (40 μg/L) of the synthesized peptides induced changes in the locomotor and the olfactory-driven behavior of the C. quinquefascitus larvae, which were associated with increased production of ROS and AChE activity (cholinesterase effect). To our knowledge, this is the first study that reports the indirect effects of the COVID-19 pandemic on the larval phase of a freshwater invertebrate species. The results raise concerns at the ecological level where the observed biological effects may lead to drastic consequences.

The increasing demand for fresh water has forced many countries to use reclaimed wastewater for agricultural purposes. This water contains pharmaceuticals and personal care products (PPCPs) that remain biologically active following passage through wastewater treatment plants. Run-off from farms and contaminated water from treatment facilities exposes aquatic ecosystems to PPCPs. This study examined the effects of PPCPs on a lower trophic organism. Culex quinquefasciatus larvae were reared in water contaminated with environmentally relevant concentrations of common PPCPs. Acetaminophen alone and a mixture of contaminants were found to increase developmental time of larvae. Susceptibility to Bti increased in larvae exposed to antibiotics, acetaminophen, or a mixture of PPCPs. Antibiotics, hormones, and the mixture altered the mosquito bacterial microbiome. Overall, the results indicate that at environmentally relevant concentrations, PPCPs in reclaimed water can have biologically important effects on an ecologically and medically important lower trophic level insect.

Medicinal and aromatic plants represent an outstanding source of green active ingredients for a broad range of real-world applications. In the present study, we investigated the insecticidal potential of the essential oils obtained from three medicinal and aromatic plants of economic importance in Algeria, Artemisia campestris, Pulicaria arabica, and Saccocalyx satureioides. Gas chromatography coupled with mass spectrometry (GC-MS) was used to study the essential oil chemical compositions. The three essential oils were tested against a mosquito vectoring filariasis and arboviruses, i.e., Culex quinquefasciatus, a fly pest acting also as pathogens vector, Musca domestica, and an agricultural moth pest, i.e., Spodoptera littoralis, using WHO and topical application methods, respectively. The essential oil from A. campestris, containing β-pinene (15.2%), α-pinene (11.2%), myrcene (10.3%), germacrene D (9.0%) (Z)-β-ocimene (8.1%) and γ-curcumene (6.4%), showed remarkable toxicity against C. quinquefasciatus (LC₅₀ of 45.8 mg L⁻¹) and moderate effects (LD₅₀ of 99.8 μg adult⁻¹) against M. domestica. Those from P. arabica and S. satureioides, containing epi-α-cadinol (23.9%), δ-cadinene (21.1%), α-cadinol (19.8%) and germacrene D-4-ol (8.4%), and thymol (25.6%), α-terpineol (24.6%), borneol (17.4%) and p-cymene (11.4%), respectively, were more active on S. littoralis showing LD₅₀ values of 68.9 and 61.2 μg larva⁻¹, respectively. Based on our results, the essential oil from A. campestris may be further considered a candidate ingredient for developing botanical larvicides.

The essential oil (EO) of Cupressus sempervirens was obtained by hydrodistillation and analysed using gas chromatography–flame ionization detection (GC–FID) and gas chromatography–mass spectrometry (GC–MS). Two monoterpenes, α-pinene (49.1%) and δ-3-carene (21.4%), and one sesquiterpene hydrocarbon, α-cedrol (5.1%), were isolated as the EO major terpenes. An oil-in-water nanoemulsion (particle size 71.2 nm) was produced from the EO through a low-energy method. The EO, its nanoemulsion and its main constituents showed mosquitocidal and biochemical effects against Culex quinquefasciatus Say, the common vector of lymphatic filariasis parasites. All treatments showed dose-dependent bioactivity, and adults were more susceptible to the EO products than the larvae. The nanoemulsion showed superior activity, followed by the crude EO and α-cedrol. At 40 μg/ml, the nanoemulsion caused 100% larval mortality, while the EO and α-cedrol required twice this concentration to achieve the same larval mortality. The LC₅₀ values were 8.4, 16.1, 15.1, 30.7 and 53.4 μg/ml at 24 h after exposure for the nanoemulsion, crude oil, α-cedrol, δ-3-carene and α-pinene, respectively. For adults, 20.0 μl/l nanoemulsion caused 100% mortality, while twice this concentration of the EO was required to achieve the same effect. The LC₅₀’s against adults ranged between 6.2 and 40.4 μl/l. EO products prominently repelled mosquitoes at concentrations between 0.75 and 6.0 μl/cm². The EO products caused remarkable inhibition of Cx. quinquefasciatus acetylcholinesterase activity but were safer towards the non-target aquatic species Gambusia affinis. These results recommend the use of C. sempervirens EO, its nanoemulsion and main terpenes as natural tools to control Cx. quinquefasciatus.

Populations of Culex quinquefasciatus Say, 1823 (Diptera: Culicidae) have shown resistance to insecticides of the carbamate and organophosphate classes. The objective of this study was to assess the susceptibility of C. quinquefasciatus larvae to essential oils from leaves of Eugenia uniflora L., Melaleuca armillaris (Sol. ex Gaertn.) Sm., and Schinus molle L and C. quinquefasciatus larvae’s biochemical responses after their exposure to these leaves. The essential oils were chemically analyzed by GC and GC/MS. First, the lethal concentration for 50% (LC₅₀) values was estimated using different concentrations of essential oils and probit analysis. The larvae were exposed for 1 h at the LC₅₀ estimated for each essential oil. The susceptibility of the larvae to essential oils was evaluated using the following biochemical parameters: concentrations of total protein and reduced glutathione; levels of production of hydrogen peroxide and lipid peroxidation; and the activity of the enzyme acetylcholinesterase (AChE). The main chemical constituents in E. uniflora were E-β-ocimene, curzerene, germacrene B, and germacrone; in M. armillaris were 1,8-cineole and terpinolene; and in S. molle were sabinene, myrcene, and sylvestrene. The essential oils had LC₅₀ values between 31.52 and 60.08 mg/L, all of which were considered effective. All of them also promoted changes in biochemical parameters when compared to the control treatment. The essential oils of S. molle and E. uniflora inhibited the activity of the AChE enzyme, and the essential oil of M. armillaris increased it. All essential oils had larvicidal activity against C. quinquefasciatus, but the essential oil of E. uniflora was the most efficient. Thus, the findings of the present study suggest that the essential oil of E. uniflora can be considered promising for the development of botanical larvicides.

Slaughterhouse waste water (SHW) is discarded as unused disposals into the environment every day. The objective of the present study is to explore the possibility on the utilization of SHW for the preparation of culture media to produce mosquitocidal bacterium (Bacillus thuringiensis subsp. israelensis). This would help to overcome the problem of dumping SHW in to environment. A judicial combination of SHW with a mineral salt (MnCl2) was made to produce an enhanced level of bacterial production when compared with other culture media including conventional medium (Luria Bertani, LB). A complete degradation of SHW by the bacteria was observed. The biomass yield, bacterial growth, toxin production, and larvicidal activity against mosquito vectors were satisfactory. Cell mass yield of 4.55 gm l−1 (dry wt) and larvicidal activity of 0.006 mg ml−1 and 0.026 mg ml−1 at LC50 and LC90 levels were observed, respectively, against the filarial vector of Culex quinquefasciatus with bacteria grown in SHW + MnCl2. The B. thuringiensis subsp. israelensis also controlled the larvae in the field significantly for three weeks (>90% mortality) and the effect was comparable with LB. Cost-analysis for production of B. thuringiensis subsp. israelensis showed that it is more economical. Thus, this study suggested the dual benefit of efficient production of mosquitocidal toxin and management of slaughterhouse wastewater.

Echinophora spinosa (Apiaceae) is a psammophilous species growing along Mediterranean coastal sand dunes. This species secretes essential oils made up of monoterpenoids, phenylpropanoids, and polyacetylenes, which may represent effective, eco-friendly and safe active ingredients for the preparation of green insecticides. Based on this, in the present study, we selected a French accession of E. spinosa growing on the coastal dunes of Corsica, extracting the essential oils from aerial parts and roots by hydrodistillation. The essential oils were analyzed by GC-MS for chemical composition and screened for their insecticidal activity on three target insects, Culex quinquefasciatus, Spodoptera littoralis, and Musca domestica. The essential oil from roots was dominated by the phenylpropanoid myristicin and the monoterpene hydrocarbon terpinolene, with a minor contribution of (Z)-falcarinol. The oil from the aerial parts was characterized by monoterpenes, mainly p-cymene, a-phellandrene, and α-pinene. The root essential oil exhibited promising toxicity on the target insects, with special reference to C. quinquefasciatus larvae (LC₅₀ = 15.7 mg L⁻¹), while on M. domestica and S. littoralis, it showed LD₅₀ of 38.3 μg adult⁻¹ and 55.6 μg larva⁻¹, respectively. Based on our findings, E. spinosa can be viewed as a new potential crop to produce insecticidal essential oils in arid salty areas. However, synergistic and antagonistic effects of the major constituents of both oils tested here deserve future research. Further studies on the efficacy of stable formulations of the E. spinosa root essential oil, with special reference to micro- and nanoformulations, are ongoing.

To seek new mosquito control agents while avoiding the environmental impacts and toxicity hazards of conventional pesticides, the essential oil of Dysphania ambrosioides was obtained by hydrodistillation and analysed using GC–FID and GC–MS. The compounds 1-methyl-4-(1-methylethyl)-2,3-dioxabicyclo[2.2.2]oct-5-ene (cis-ascaridole), 1-methyl-4-(1-methylethyl) benzene (р-cymene), and 1-isopropyl-4-methyl-1,3-cyclohexadiene (p-mentha-1,3-diene also known as α-terpinene) were identified as the major components. The EO and the major fractions showed remarkable mosquitocidal activity against third instar larvae and adults of Culex quinquefasciatus Say. The oil and fractions were assayed at 3.125, 6.25, 12.5, 25, and 50 μl/l. Mortality was time- and dose-dependent. At 24 h post-exposure at an assayed concentration of 50 μl/l, the larval and adult mortalities ranged between 80.11–100% and 91.22–100%, respectively. Strong larvicidal and adulticidal activities were recorded in the cases of the crude oil and cis-ascaridole. The LC₅₀ values after 24 h of treatment ranged between 6.2–20.1 μl/l and 5.1–13.9 μl/l against larvae and adults, respectively. The corrected percentage mortalities increased over time with the tested plant oil and the major fractions relative to the control. The time required to achieve 50% mortality (LT₅₀) decreased remarkably with all treatments. The tested EO and major fractions effectively inhibited larval acetylcholinesterase activity with IC₅₀ values ranging from 8.44 to 64.80 mM compared with 2.08 × 10⁻³ mM for the reference standard, methomy. The results indicate the potential of developing natural mosquitocides against C. quinquefasciatus based on the tested EO and its major fractions. Graphical abstract

Aedes aegypti and Culex quinquefasciatus are vectors of diseases that constitute public health problems. The discovery of products capable of inhibiting their development which are less harmful to the environment would have a huge impact on vector control. Here, natural cashew nut shell liquid (CNSL), technical CNSL, anacardic acid, cardanol, and cardol were isolated from Anacardium occidentale and evaluated for larvicidal and pupicidal activity against Ae. aegypti and Cx. quinquefasciatus under laboratory and field conditions. The activities of phenol, resorcinol, salicylic acid, and pentadecane, commercial chemicals similar in structure to nut shell derivatives, were also evaluated. All of the fractions extracted from A. occidentale oil exerted larvicidal effects against both mosquito species (LC₅₀ 5.4–22.6 mg/L), and two of the aforementioned were effective against pupae (LC₅₀ 90.8–109.7 mg/L). Of all the fractions tested, cardol demonstrated the strongest larvicidal and pupicidal effects and presented the most prolonged residual activity against the larvae and pupae of Ae. aegypti and Cx. quinquefasciatus under field conditions. This study suggests that A. occidentale nut shell derivatives are sustainable and promising candidates for the development of novel insecticides to overcome the problem of harmful chemical insecticides.

Mosquitoes are responsible for the transmission of many pathogens and parasites, which cause serious diseases in humans and animals. Currently, botanical products have been suggested as alternative tools in the fight against arthropod vectors. In this study, the essential oil (EO) extracted from Zingiber cernuum was tested as larvicide and oviposition deterrent on six mosquito species of public health relevance, including malaria and Zika virus vectors. The EO showed high toxicity on third instar larvae of Anopheles stephensi (LC₅₀ = 41.34 μg/ml), Aedes aegypti (LC₅₀ = 44.88 μg/ml), Culex quinquefasciatus (LC₅₀ = 48.44 μg/ml), Anopheles subpictus (LC₅₀ = 51.42 μg/ml), Aedes albopictus (LC₅₀ = 55.84 μg/ml), and Culex tritaeniorhynchus (LC₅₀ = 60.20 μg/ml). In addition, low doses of Z. cernuum EO reduced oviposition rates in six mosquito species. The acute toxicity of Z. cernuum EO on four mosquito predators was scarce; LC₅₀ ranged from 3119 to 11,233 μg/ml. Overall, our results revealed that the Z. cernuum EO can be considered for the development of effective and environmental-friendly mosquito larvicides and oviposition deterrents.