Exposure to neonicotinoid pesticides induces physiological disorders and affects color performance and foraging behavior in goldfish.

We investigated the effects of neonicotinoid pesticides (NEOs) on the spontaneous swimming and foraging behavior, as well as the morphological and physiological changes of goldfish. Most fish reared in thiamethoxam (THM)-sprayed rice fields showed the scales easily peeled off, and increased ascites. Some individuals showed decreased bio-defense activity and low plasma Ca2+. Similar changes were found in the exposure test to THM (1.0 and 20.0 µg/L) and dinotefuran (1.2 and 23.5 µg/L). Next, the effects of a low concentration of THM (1.0 µg/L) on the spontaneous swimming and foraging behavior of fish were examined. Fish exposed to THM for 1 week became restless and had increased the swimming performance, especially under natural light, white LED lighting and blue LED lighting. Goldfish exposed to THM had also increased intake of shiny white beads under green LED illumination. These results indicate that the exposure to NEO, even for a short period and at low levels, not only suppressed bio-defense activities and metabolic abnormalities, but also stress response, the swimming and foraging behavior of the fish are likely to be significantly suffered.

Toxicokinetics and bioavailability of indoxacarb enantiomers and their new metabolites in rats.

Indoxacarb is a chiral insecticide that consists of two enantiomers, S-(+)-indoxacarb and R-(-)-indoxacarb, of which only S-(+)-indoxacarb has insecticidal activity. Previous enantioselective toxicology studies of indoxacarb focused mostly on simple environmental model organisms. The lack of a toxicology evaluation of indoxacarb conducted in a mammalian system could mean that the extent of the potential health risk posed by the insecticide to humans is not adequately known. In this study, we reported on a new pair of enantiomers, S-IN-RM294 and R-IN-RM294, derived from the metabolic breakdown of S-(+)-indoxacarb and R-(-)-indoxacarb, respectively, in rats. The toxicokinetics of S-(+)-indoxacarb, R-(-)-indoxacarb, S-IN-RM294, and R-IN-RM294 in rats were evaluated to provide a more comprehensive risk assessment of these molecules. The bioavailability and excretion rates of both S-(+)-indoxacarb and R-(-)-indoxacarb were relatively low, which may be due to their faster metabolism and accumulation in the tissues. In addition, there were significant differences in the metabolism and distribution between the two indoxacarb enantiomers and their metabolites in vivo. S-(+)-Indoxacarb was found to be more easily metabolized in the blood compared with R-(-)-indoxacarb, as shown by the differences in pharmacokinetic parameters between oral and intravenous administration. Analysis of their tissue distribution showed that S-(+)-indoxacarb was less likely to accumulate in most tissues. The results obtained for the two metabolites were consistent with those of the two parent compounds. S-IN-RM294 was more readily cleared from the blood and less likely to accumulate in the tissues compared with R-IN-RM294. Therefore, whether from the perspective of insecticidal activity or from the perspective of mammalian and environmental friendliness, the application of optically pure S-(+)-indoxacarb in agriculture may be a more efficient and safer strategy.

Toxic effects of commercial grade indoxacarb and endosulfan on Gammarus kischineffensis (Schellenberg, 1937) (Crustacea: Amphipoda).

This study was designed to investigate the toxic effects of two frequently used commercial insecticides containing endosulfan and indoxacarb on a freshwater amphipod Gammarus kischineffensis. In this context, the 24, 48, 72 and 96 h LC50 values of these pesticides were determined for G. kischineffensis. Then the histopathological effects of these pesticides on the gill tissues of this species were evaluated. At the end of the study, the 96 h LC50 values of commercial-grade endosulfan and indoxacarb for G. kischineffensis were determined as 1.861 µg L-1 and 20.212 mg L-1, respectively. Histopathologically, the most common histopathological alterations in individuals exposed to sublethal concentrations of commercial-grade endosulfan and indoxacarb were pillar cell hypertrophy resulting in atrophy of the hemocoelic space and hemocytic infiltration. Considering these results, it can be said that commercial-grade endosulfan is extremely and indoxacarb is slightly toxic to G. kischineffensis.

Sublethal effect and detoxifying metabolism of metaflumizone and indoxacarb on the fall armyworm, Spodoptera frugiperda.

The fall armyworm (FAW), Spodoptera frugiperda (J.E. Smith) (Lepidoptera, Noctuidae), is a highly polyphagous invasive pest that damages various crops. Pesticide control is the most common and effective strategy to control FAW. In this study, we evaluated the toxicity of metaflumizone and indoxacarb against third-instar FAW larvae using the insecticide-incorporated artificial diet method under laboratory conditions. Both metaflumizone and indoxacarb exhibited substantial toxicity against FAW, with LC50 values of 2.43 and 14.66 mg/L at 72 h, respectively. The sublethal effects of metaflumizone and indoxacarb on parental and F1 generation FAW were investigated by exposing third-instar larvae to LC10 and LC30 concentrations of these insecticides. Sublethal exposure to these two insecticides significantly shortened adult longevity, extended pupal developmental times and led to reduced pupal weight, pupation rates, and adult fecundity in the treated parental generation and F1 generation at LC10 or LC30 concentrations, in comparison to the control group. The larval developmental times were shortened in the parental generation but prolonged in the F1 generation, after being treated with sublethal concentrations of metaflumizone. Furthermore, larvae exposed to LC10 or LC30 concentrations of indoxacarb exhibited elevated activity levels of cytochrome P450 monooxygenase and glutathione S-transferase, which coincides with the observed synergistic effect of piperonyl butoxide and diethyl maleate. In conclusion, the high toxicity and negative impact of metaflumizone and indoxacarb on FAW provided significant implications for the rational utilization of insecticides against this pest.

Does commercial indoxacarb pose ecotoxicological consequences? Employing a multi-marker approach in the model species Theba pisana.

Indoxacarb is one of the most extensively used oxadiazine insecticides worldwide, but it may exert detrimental effects on ecosystems, population dynamics, and health. Due to the lack of knowledge on the ecotoxicity of indoxacarb, it is still challenging to assess whether this insecticide poses an ecotoxicological impact on terrestrial environments. Therefore, our study aims to provide novel data on the toxic effects of 28-day dietary exposure to commercial grade indoxacarb at two environmentally relevant concentrations, 0.02 µg/mL and tenfold (0.2 µg/mL) on the model species, Theba pisana. Their effects were studied using a multiple biomarker approach by evaluating physiological, biochemical, and histopathological responses. After 28 days of treatment, indoxacarb at both concentrations significantly reduced the food intake and growth of the treated snails. Also, it caused decreases in lipid peroxidation (LPO)  levels after 7 and 14 days of exposure, whereas an opposite effect occurred after 21 and 28 days. All treated snails were found to exhibit a lower content of glutathione (GSH) after all times of exposure. Moreover, catalase (CAT), glutathione-S-transferase (GST), and glutathione peroxidase (GPx) activities, as well as protein content (PC), were elevated in the treated snails after all time intervals. Post exposure to both realistic indoxacarb concentrations, changes in acetylcholinesterase (AChE) activity between a decrease and an increase were observed. Furthermore, indoxacarb caused histo-architectural changes in the hepatopancreas of T. pisana. Our results demonstrate that, at environmentally relevant concentrations, indoxacarb poses negative consequences for T. pisana, indicating its ecotoxicological impacts.

Microbial degradation mechanism and pathway of the insecticide thiamethoxam by isolated Bacillus Cereus from activated sludge.

The high water solubility and ecotoxicity of thiamethoxam (TMX) is a potential hazard to ecosystems and human health. Here, a strain of Bacillus cereus with high TMX degradation activity was isolated from the sediment of the A2O process in the wastewater treatment plant and was able to utilize TMX as its sole carbon source. Under different environmental conditions, the degradation efficiency of TMX by Bacillus cereus-S1 (strain S1) ranged from 41.0% to 68.9% after 216 h. The optimum degradation conditions were DO = 3.5 mg/L and pH 9.0. The addition of an appropriate carbon-to-nitrogen ratio could accelerate the degradation of TMX. A plausible biodegradation pathway has been proposed based on the identified metabolites and their corresponding degradation pathways. TMX can be directly converted into Clothianidin (CLO), TMX-dm-hydroxyl and TMX-Urea by a series of reactions such as demethylation, oxadiazine ring cleavage and C=N substitution by hydroxy group. The main products were TMX-dm-hydroxyl and TMX-Urea, the amount of CLO production is relatively small. This study aims to provide a new approach for efficient degradation of TMX; furthermore, strain S1 is a promising biological source for in situ remediation of TMX contamination.

Low concentration of indoxacarb interferes with the growth and development of silkworm by damaging the structure of midgut cells.

As an important economic insect, Bombyx mori plays an essential role in the development of the agricultural economy. Indoxacarb, a novel sodium channel blocker insecticide, has been widely used for the control of various pests in agriculture and forestry, and its environmental pollution caused by flight control operations has seriously affected the safe production of sericulture in recent years. However, the lethal toxicity and adverse effects of indoxacarb on silkworm remain largely unknown. In this study, the toxicity of indoxacarb on the 5th instar larvae of silkworm was determined, with an LC50 (72 h) of 2.07 mg/L. Short-term exposure (24 h) to a low concentration of indoxacarb (1/2 LC50) showed significantly reduced body weight and survival rate of silkworm larvae. In addition, indoxacarb also led to decreased cocoon weight and cocoon shell weight, but had no significant effects on pupation, adult eclosion, and oviposition. Histopathological and ultrastructural analysis indicated that indoxacarb could severely damage the structure of the midgut epithelial cells, and lead to physiological impairment of the midgut. A total of 3883 differentially expressed genes (DEGs) were identified by midgut transcriptome sequencing and functionally annotated using GO and KEGG. Furthermore, the transcription level and enzyme activity of the detoxification related genes were determined, and our results suggested that esterases (ESTs) might play a major role in metabolism of indoxacarb in the midgut of B. mori. Future studies to examine the detoxification or biotransformation function of candidate genes will greatly enhance our understanding of indoxacarb metabolism in B. mori. The results of this study provide a theoretical basis for elucidating the mechanism of toxic effects of indoxacarb on silkworm by interfering with the normal physiological functions of the midgut.

Comparative toxicity of two neonicotinoid insecticides at environmentally relevant concentrations to telecoprid dung beetles.

Dung beetles (Coleoptera: Scarabaeinae) frequently traverse agricultural matrices in search of ephemeral dung resources and spend extended periods of time burrowing in soil. Neonicotinoids are among the most heavily applied and widely detected insecticides used in conventional agriculture with formulated products designed for row crop and livestock pest suppression. Here, we determined the comparative toxicity of two neonicotinoids (imidacloprid and thiamethoxam) on dung beetles, Canthon spp., under two exposure profiles: direct topical application (acute) and sustained contact with treated-soil (chronic). Imidacloprid was significantly more toxic than thiamethoxam under each exposure scenario. Topical application LD50 values (95% CI) for imidacloprid and thiamethoxam were 19.1 (14.5-25.3) and 378.9 (200.3-716.5) ng/beetle, respectively. After the 10-day soil exposure, the measured percent mortality in the 3 and 9 µg/kg nominal imidacloprid treatments was 35 ± 7% and 39 ± 6%, respectively. Observed mortality in the 9 µg/kg imidacloprid treatment was significantly greater than the control (p = 0.04); however, the 3 µg/kg imidacloprid dose response may be biologically relevant (p = 0.07). Thiamethoxam treatments had similar mortality as the controls (p > 0.8). Environmentally relevant concentrations of imidacloprid measured in airborne particulate matter and non-target soils pose a potential risk to coprophagous scarabs.

Assessment of Global DNA Methylation in SH-SY5Y Cells Exposed to the Neonicotinoid Insecticides Imidacloprid and Thiamethoxam.

Neonicotinoid insecticides, known for their selectivity and low mammalian toxicity, have been widely used in recent years as alternatives to organophosphate insecticides. Although neonicotinoids are generally considered to be safe, data show that they can cause harmful effects on human and environmental health. Due to the lack of information on their mechanism of toxicity, the effects of imidacloprid and thiamethoxam on DNA methylation as the most used marker for epigenetic effects were investigated in human neuroblastoma (SH-SY5Y) cells. The cells were exposed to imidacloprid and thiamethoxam in concentrations of 100, 200, and 500 µM for 24 hours, then global DNA methylation and expression of genes involved in global DNA methylation (DNMT1, DNMT3a and DNMT3b) were investigated. Global DNA methylation significantly increased after imidacloprid exposure at 100 µM, and thiamethoxam exposures at 200 µM and 500 µM (>1.5-fold). Imidacloprid significantly decreased the expression of DNMT1 and DNMT3a, whereas thiamethoxam did not cause any significant changes in the expression of DNMT genes. Our findings suggested that alteration in global DNA methylation may be involved in the toxic mechanisms of imidacloprid and thiametoxam.

Differential effects of thiamethoxam and clothianidin exposure on their tissue distribution and chronic toxicity in mice.

The frequent application of second-generation neonicotinoids thiamethoxam (TMX) and clothianidin (CLO) has led to a high detectable rate in environment samples and poses threats to nontarget organisms and human beings, however, the information on the influences of long-term exposure at low doses was limited. In this study, the tissue distribution of TMX and CLO in mice at acceptable daily intake (ADI) level and 5 × ADI was determined and the health effects were assessed. TMX and CLO were detected in the liver, serum, lung, heart and kidney in the TMX exposure groups, which indicated that TMX degraded to CLO in mice. Residue levels of TMX in tissues increased with the increasing of doses. The concentrations of CLO in different tissues in the CLO exposure groups were in the order Ckidney > Clung > Cheart > Cliver. Measurement of biochemical indicators, combined with metabolomic analysis of liver, kidney, and cecal contents, examination of changes in the gut microbiota, and histopathological assessment indicated that both TMX and CLO affected energy absorption and lipid metabolism in mice and destroyed tissue structures. Furthermore, we found that CLO had a stronger effect on metabolism in mice, despite its lower acute toxicity. These results have prompted us to consider the chronic toxicity and potential hazards of chemicals in future risk assessments.

In vitro/in silico prediction of drug induced steatosis in relation to oral doses and blood concentrations by the Nile Red assay.

The accumulation of lipid droplets in hepatocytes is a key feature of drug-induced liver injury (DILI) and can be induced by a subset of hepatotoxic compounds. In the present study, we optimized and evaluated an in vitro technique based on the fluorescent dye Nile Red, further named Nile Red assay to quantify lipid droplets induced by the exposure to chemicals. The Nile Red assay and a cytotoxicity test (CTB assay) were then performed on cells exposed concentration-dependently to 60 different compounds. Of these, 31 were known to induce hepatotoxicity in humans, and 13 were reported to also cause steatosis. In order to compare in vivo relevant blood concentrations, pharmacokinetic models were established for all compounds to simulate the maximal blood concentrations (Cmax) at therapeutic doses. The results showed that several hepatotoxic compounds induced an increase in lipid droplets at sub-cytotoxic concentrations. To compare how well (1) the cytotoxicity test alone, (2) the Nile Red assay alone, and (3) the combination of the cytotoxicity test and the Nile Red assay (based on the lower EC10 of both assays) allow the differentiation between hepatotoxic and non-hepatotoxic compounds, a previously established performance metric, the Toxicity Separation Index (TSI) was calculated. In addition, the Toxicity Estimation Index (TEI) was calculated to determine how well blood concentrations that cause an increased DILI risk can be estimated for hepatotoxic compounds. Our findings indicate that the combination of both assays improved the TSI and TEI compared to each assay alone. In conclusion, the study demonstrates that inclusion of the Nile Red assay into in vitro test batteries may improve the prediction of DILI compounds.

Enantioselectivity of indoxacarb during the growing, processing, and brewing of tea: Degradation, metabolites, and toxicities.

Chiral pesticides are unique hazardous materials. Here, we systematically studied the potentially harmful products of enantioselective indoxacarb degradation throughout tea growth, processing, and brewing and tested their toxicity to tea geometrid larvae and honeybees. The half-lives of S-indoxacarb and R-indoxacarb during tea growth were 2.6 d and 3.3 d, respectively. There was a trend toward the production of S-indoxacarb from R-indoxacarb. The degradation products IN-JT333, IN-MK638, IN-MF014, and IN-KG433 were also characterized in tea growth and processing and detected. IN-JT333, previously known as a direct insecticidal compound produced by the enzymatic transformation of indoxacarb in insects, was first found in plant samples. The fixation and rolling of green tea and the rolling of black tea were the most important steps that affected indoxacarb and its degradation products. The leaching rates of R-indoxacarb and S-indoxacarb were slightly higher in green tea than in black tea. The maximum leaching rates of IN-MK638 and IN-MF014 during the brewing process reached 89.9% and 94.1%, respectively. Contact toxicity tests with honeybees and tea geometrid larvae in the lab showed that the relative toxicities of the compounds could be ranked as follows: S-indoxacarb > indoxacarb (3S + 1R) ≫ R-indoxacarb. TEST toxicity predictions showed that relative toxicities were ranked IN-KG433 > indoxacarb > IN-JT333 > IN-MK638 > IN-MF014. The toxicity of the degradation product IN-KG433 is higher than that of indoxacarb itself, and its maximum leaching rate is as high as 88.2%. It therefore transfers readily from processed tea to the tea infusion during the brewing process. These findings indicate the need to pay attention to the risk of metabolites and enantiomeric differences and provide new, comprehensive insight into the risk factors for indoxacarb in tea and are relevant to the study of other chiral pesticides.

Insecticide resistance in Australian Spodoptera frugiperda (J.E. Smith) and development of testing procedures for resistance surveillance.

Spodoptera frugiperda (J.E. Smith) is a highly invasive noctuid pest first reported in northern Australia during early 2020. To document current status of resistance in S. frugiperda in Australia, insecticide toxicity was tested in field populations collected during the first year of establishment, between March 2020 and March 2021. Dose-response was measured by larval bioassay in 11 populations of S. frugiperda and a susceptible laboratory strain of Helicoverpa armigera. Emamectin benzoate was the most efficacious insecticide (LC50 0.023µg/ml) followed by chlorantraniliprole (LC50 0.055µg/ml), spinetoram (LC50 0.098µg/ml), spinosad (LC50 0.526µg/ml), and methoxyfenozide (1.413µg/ml). Indoxacarb was the least toxic selective insecticide on S. frugiperda (LC50 3.789µg/ml). Emamectin benzoate, chlorantraniliprole and methoxyfenozide were 2- to 7-fold less toxic on S. frugiperda compared with H. armigera while spinosyns were equally toxic on both species. Indoxacarb was 28-fold less toxic on S. frugiperda compared with H. armigera. There was decreased sensitivity to Group 1 insecticides and synthetic pyrethroids in S. frugiperda compared with H. armigera: toxicity was reduced up to 11-fold for methomyl, 56 to 199-fold for cyhalothrin, and 44 to 132-fold for alpha cypermethrin. Synergism bioassays with metabolic inhibitors suggest involvement of mixed function oxidase in pyrethroid resistance. Recommended diagnostic doses for emamectin benzoate, chlorantraniliprole, spinetoram, spinosad, methoxyfenozide and indoxacarb are 0.19, 1.0, 0.75, 6, 12 and 48µg/µl, respectively.

Real-Life Impact of Drug Toxicity on Dolutegravir Tolerability: Clinical Practice Data from a Multicenter Italian Cohort.

Dolutegravir (DTG) is currently one of the most used Integrase inhibitors (INI) in antiretroviral therapies (ARV) in both naïve and experienced people living with HIV (PLWHIV). We analyzed a multicenter cohort of PLWHIV, both naïve and experienced, starting an ARV including DTG. We enrolled 3775 PLWHIV: 2763 (73.2%) were males, with a median age of 50 years. During 9890.7 PYFU, we observed 930 discontinuations (9.4 per 100 PYFU). Estimated probabilities of maintaining DTG at three and five years were 75.1% and 67.2%, respectively. Treatment-naïve pts showed a lower probability of maintaining DTG at three and five years compared to treatment-experienced PLWHIV (log-rank p < 0.001). At a multivariate analysis, a longer time of virological suppression (aHR 0.994, p < 0.001) and having experienced a previous virological failure (aHR 0.788, p = 0.016) resulted protective against DTG discontinuation. Most discontinuations (84.0%) happened within the first 12 months of DTG initiation, in particular, 92.2% of discontinuations due to neuropsychiatric toxicity were observed in the first year. Our data confirm the overall good tolerability of DTG in clinical practice, with a low rate of discontinuations. CNS toxicity resulted the main reason for DTG discontinuation, with most related interruptions happening in the first year from DTG introduction.

Is there a risk to honeybees from use of thiamethoxam as a sugar beet seed treatment?

The ban imposed by the European Union on the use of neonicotinoids as sugar beet seed treatments was based on the exposure of bees to residues of neonicotinoids in pollen and nectar of succeeding crops. To address this concern, residues of thiamethoxam (TMX) and clothianidin (CTD) were analyzed in soil collected from fields planted in at least the previous year with thiamethoxam-treated sugar beet seed. This soil monitoring program was conducted at 94 sites across Germany in two separate years. In addition, a succeeding crop study assessed residues in soil, guttation fluid, pollen, and nectar sampled from untreated succeeding crops planted in the season after thiamethoxam seed-treated sugar beet at eight field sites across five countries. The overall mean residues observed in soil monitoring were 8.0 ± 0.5 µg TMX + CTD/kg in the season after the use of treated sugar beet seed. Residue values decreased with increasing time interval between the latest thiamethoxam or clothianidin application before sugar beet drilling and with lower application frequency. Residues were detected in guttation fluid (2.0-37.7 µg TMX/L); however, the risk to pollinators from this route of exposure is likely to be low, based on the reported levels of consumption. Residues of thiamethoxam and clothianidin in pollen and nectar sampled from the succeeding crops were detected at or below the limit of quantification (0.5-1 µg a.i./kg) in 86.7% of pollen and 98.6% of nectar samples and, unlike guttation fluid residues, were not correlated with measured soil residues. Residues in pollen and nectar are lower than reported sublethal adverse effect concentrations in studies with honeybee and bumble bee individuals and colonies fed only thiamethoxam-treated sucrose, and are lower than those reported to result in no effects in honeybees, bumble bees, and solitary bees foraging on seed-treated crops. Integr Environ Assess Manag 2022;18:709-721. © 2021 SYNGENTA. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).

Spatiotemporal elements in a poisoned bait strategy against the long-tailed silverfish (Lepismatidae: Zygentoma).

The long-tailed silverfish Ctenolepisma longicaudatum (Lepismatidae: Zygentoma) is a nuisance problem in buildings and a major concern in museums, libraries and archives where it cause damage to historical and priceless items. We used laboratory bioassays and two field studies of infested buildings to evaluate spatial and temporal elements of a poisoned bait strategy. In both laboratory experiments and field studies, the efficiency of poisoned bait with indoxacarb as the active ingredient was significantly improved by placing many small bait droplets evenly distributed along all edges of the treated area compared to more clustered distributions. Extended duration of bait presence and removal of competing food sources improved the control effect significantly in the laboratory bioassays. Bait-treated populations also showed a significant decline in the number of eggs deposited and emergence of new nymphs. The study supports poisoned bait as an efficient and low risk approach against the long-tailed silverfish in which extended duration of bait presence, wide distribution of bait droplets in combination with sanitation was crucial for control in the infested premises.

Dolutegravir Inhibition of Matrix Metalloproteinases Affects Mouse Neurodevelopment.

Dolutegravir (DTG) is a first-line antiretroviral drug (ARV) used in combination therapy for the treatment of human immunodeficiency virus type-1 (HIV-1) infection. The drug is effective, safe, and well tolerated. Nonetheless, concerns have recently emerged for its usage in pregnant women or those of child-bearing age. Notably, DTG-based ARV regimens have been linked to birth defects seen as a consequence of periconceptional usages. To this end, uncovering an underlying mechanism for DTG-associated adverse fetal development outcomes has gained clinical and basic research interest. We now report that DTG inhibits matrix metalloproteinases (MMPs) activities that could affect fetal neurodevelopment. DTG is a broad-spectrum MMPs inhibitor and binds to Zn++ at the enzyme's catalytic domain. Studies performed in pregnant mice show that DTG readily reaches the fetal central nervous system during gestation and inhibits MMP activity. Postnatal screenings of brain health in mice pups identified neuroinflammation and neuronal impairment. These abnormalities persist as a consequence of in utero DTG exposure. We conclude that DTG inhibition of MMPs activities during gestation has the potential to affect prenatal and postnatal neurodevelopment.

Enzymatic responses in the head and midgut of Africanized Apis mellifera contaminated with a sublethal concentration of thiamethoxam.

The increasing use of insecticides, promoted by the intensification of agriculture, has raised concerns about their influence on the decline of bee colonies, which play a fundamental role in pollination. Thus, it is fundamental to elucidate the effects of insecticides on bees. This study investigated the damage caused by a sublethal concentration of thiamethoxam - TMX (0.0227 ng/µL of feed) in the head and midgut of Africanized Apis mellifera, by analyzing the enzymatic biomarkers, oxidative stress, and occurrence of lipid peroxidation. The data showed that the insecticide increased acetylcholinesterase activity (AChE) and glutathione-S-transferase (GST), whereas carboxylesterase (CaE3) activity decreased in the heads. Our results indicate that the antioxidant enzymes were less active in the head because only glutathione peroxidase (GPX) showed alterations. In the midgut, there were no alkaline phosphatase (ALP) or superoxide dismutase (SOD) responses and a decrease in the activity of CaE was observed. Otherwise, there was an increase in GPX, and the TBARS (thiobarbituric acid reactive substances) assay also showed differences in the midgut. The TBARS (thiobarbituric acid reactive substances) assay also showed differences in the midgut. The results showed enzymes such as CaE3, GST, AChE, ALP, SOD, and GPX, as well as the TBARS assay, are useful biomarkers on bees. They may be used in combination as a promising tool for characterizing bee exposure to insecticides.

Phenoxazine nucleoside derivatives with a multiple activity against RNA and DNA viruses.

Emerging and re-emerging viruses periodically cause outbreaks and epidemics all over the world, eventually leading to global events such as the current pandemic of the novel SARS-CoV-2 coronavirus infection COVID-19. Therefore, an urgent need for novel antivirals is crystal clear. Here we present the synthesis and evaluation of an antiviral activity of phenoxazine-based nucleoside analogs divided into three groups: (1) 8-alkoxy-substituted, (2) acyclic, and (3) carbocyclic. The antiviral activity was assessed against a structurally and phylogenetically diverse panel of RNA and DNA viruses from 25 species. Four compounds (11a-c, 12c) inhibited 4 DNA/RNA viruses with EC50 ≤ 20 µM. Toxicity of the compounds for the cell lines used for virus cultivation was negligible in most cases. In addition, previously reported and newly synthesized phenoxazine derivatives were evaluated against SARS-CoV-2, and some of them showed promising inhibition of reproduction with EC50 values in low micromolar range, although accompanied by commensurate cytotoxicity.

Transcription factor CncC potentially regulates the expression of multiple detoxification genes that mediate indoxacarb resistance in Spodoptera litura.

The xenobiotic transcription factor cap 'n' collar isoform C (CncC) is considered the central regulator of antioxidant and detoxification genes. Previous research indicated that CncC regulates three-phase enzymes responsible for insecticide resistance. In this study, the SlituCncC gene from Spodoptera litura was obtained and characterized. Quantitative polymerase chain reaction (qPCR) analysis showed that SlituCncC was expressed in all developmental stages and tissues, but was highly expressed in 3rd- and 4th-instar larvae, and in the Malpighian tubule, fat body, and midgut. In addition, SlituCncC was up-regulated and more highly induced with indoxacarb treatment in the indoxacarb-resistant strains compared with the susceptible strain. RNA interference-mediated gene silencing of SlituCncC significantly increased mortality of S. litura when exposed to indoxacarb. Furthermore, comparative transcriptome analysis showed that 842 genes were down-regulated and 127 genes were up-regulated in SlituCncC knockdown S. litura. Further analysis indicated that 18 three-phase enzymes were identified in the down-regulated genes, of which seven were associated with indoxacarb resistance in S. litura. qPCR analysis confirmed that expression of six of these seven genes was consistent with RNA sequencing data. All six detoxification genes were induced by indoxacarb, and the expression patterns were similar to that of SlituCncC. Finally, the CncC-Maf binding site was predicted in all six gene promoters. This study indicates that the transcription factor SlituCncC may regulate multiple detoxification genes that mediate indoxacarb resistance in S. litura.