A single post-exposure oxime RS194B treatment rapidly reactivates acetylcholinesterase and reverses acute symptoms in macaques exposed to diethylphosphorothioate parathion and chlorpyrifos insecticides.

Millions of individuals globally suffer from inadvertent, occupational or self-harm exposures from organophosphate (OP) insecticides, significantly impacting human health. Similar to nerve agents, insecticides are neurotoxins that target and inhibit acetylcholinesterase (AChE) in central and peripheral synapses in the cholinergic nervous system. Post-exposure therapeutic countermeasures generally include administration of atropine with an oxime to reactivate the OP-inhibited AChE. However, animal model studies and recent clinical trials using insecticide-poisoned individuals have shown minimal clinical benefits of the currently approved oximes and their efficacy as antidotes has been debated. Currently used oximes either reactivate poorly, do not readily cross the blood-brain barrier (BBB), or are rapidly cleared from the circulation and must be repeatedly administered. Zwitterionic oximes of unbranched and simplified structure, for example RS194B, have been developed that efficiently cross the BBB resulting in reactivation of OP-inhibited AChE and dramatic reversal of severe clinical symptoms in mice and macaques exposed to OP insecticides or nerve agents. Thus, a single IM injection of RS194B has been shown to rapidly restore blood AChE and butyrylcholinesterase (BChE) activity, reverse cholinergic symptoms, and prevent death in macaques following lethal inhaled sarin and paraoxon exposure. The present macaque studies extend these findings and assess the ability of post-exposure RS194B treatment to counteract oral poisoning by highly toxic diethylphosphorothioate insecticides such as parathion and chlorpyrifos. These OPs require conversion by P450 in the liver of the inactive thions to the active toxic oxon forms, and once again demonstrated RS194B efficacy to reactivate and alleviate clinical symptoms within 60 mins of a single IM administration. Furthermore, when delivered orally, the Tmax of RS194B at 1-2 h was in the same range as those administered IM but were maintained in the circulation for longer periods greatly facilitating the use of RS194B as a non-invasive treatment, especially in isolated rural settings.

Chlorella vulgaris algae ameliorates chlorpyrifos toxicity in Nile tilapia with special reference to antioxidant enzymes and Streptococcus agalactiae infection.

BACKGROUND: Chlorpyrifos (CPF) is a widely used pesticide in the production of plant crops. Despite rapid CPF biodegradation, fish were exposed to wastewater containing detectable residues. Recently, medicinal plants and algae were intensively used in aquaculture to replace antibiotics and ameliorate stress impacts. METHODS AND RESULTS: An indoor experiment was conducted to evaluate the deleterious impacts of CPF pollution on Nile tilapia health and the potential mitigation role of Chlorella vulgaris algae. Firstly, the median lethal concentration LC50 - 72 h of CPF was determined to be 85.8 µg /L in Nile tilapia (35.6 ± 0.5 g body weight) at a water temperature of 27.5 °C. Secondly, fish were exposed to 10% of LC50 - 72 h for six weeks, and tissue samples were collected and examined every two weeks. Also, Nile tilapia were experimentally infected with Streptococcus agalactiae. Exposed fish were immunosuppressed expressed with a decrease in gene expressions of interleukin (IL) 1ß, IL-10, and tumor necrosis factor (TNF)-α. Also, a decline was recorded in glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT) gene expression in the head kidney tissue. A high mortality rate (MR) of 100% was recorded in fish exposed to CPF for six weeks and challenged with S. agalactiae. Fish that received dietary C. vulgaris could restore gene expression cytokines and antioxidants compared to the control. After six weeks of CPF exposure, fish suffered from anemia as red blood cell count (RBCs), hemoglobin (Hb), and packed cell volume (PCV) significantly declined along with downregulation of serum total protein (TP), globulin (GLO), and albumin (ALB). Liver enzymes were significantly upregulated in fish exposed to CPF pollution, alanine aminotransferase (ALT) (42.5, 53.3, and 61.7 IU/L) and aspartate aminotransferase (AST) (30.1, 31.2, and 22.8) after 2, 4, and 6 weeks, respectively. On S. agalactiae challenge, high MR was recorded in Nile tilapia exposed to CPF (G3) 60%, 60%, and 100% in week 2, week 4, and week 6, and C. vulgaris provided a relative protection level (RPL) of 0, 14.29, and 20%, respectively. CONCLUSIONS: It was concluded that CPF pollution induces immunosuppressed status, oxidative stress, and anemic signs in Nile tilapia. In contrast, C. vulgaris at a 50 g/kg fish feed dose could partially ameliorate such withdrawals, restoring normal physiological parameters.

Chlorpyrifos enrichment enhances tolerance of Anabaena sp. PCC 7119 to dimethoate.

Organophosphorus (OP) insecticides are widely used for on-field pest control, constituting about 38% of global pesticide consumption. Insecticide tolerance has been recorded in microorganisms isolated from the contaminated soil. However, the cross-tolerance of laboratory-enriched cultures remains poorly understood. A chlorpyrifos tolerant (T) strain of Anabaena sp. PCC 7119 was developed through continuous enrichment of the wild strain (W). The cross-tolerance of the T strain to the OP insecticide dimethoate was assessed by measuring photosynthetic performance, key enzyme activities and degradation potential. The presence of dimethoate led to a significant reduction in the growth and pigment content of the W strain. In contrast, the T strain demonstrated improved growth and metabolic performance. Chl a and carotenoids were degraded faster than phycobiliproteins in both strains. The T strain exhibited superior photosynthetic performance, metabolic efficiency and photosystem functions, than of W strain, at both the tested dimethoate concentrations (100 and 200 µM). The treated T strain had more or less a normal OJIP fluorescence transient and bioenergetic functions, while the W strain showed a greater fluorescence rise at ≤ 300 µs indicating the inhibition of electron donation to PS II, and at 2 ms due to reduced electron release beyond QA. The T strain had significantly higher levels of esterase and phosphatases, further enhanced by insecticide treatment. Dimethoate degradation efficiency of the T strain was significantly higher than of the W strain. T strain also removed chlorpyrifos more efficiently than W strain at both the tested concentrations. The BCFs of both chlorpyrifos and dimethoate were lower in the T strain compared to the W strain. These findings suggest that the enriched strain exhibits promising results in withstanding dimethoate toxicity and could be explored for its potential as a bioremediating organism for OP degradation.

Exposure to chlorpyrifos and pyrethroid insecticides and symptoms of Attention Deficit Hyperactivity Disorder (ADHD) in preschool children from the Odense Child Cohort.

BACKGROUND: Attention Deficit Hyperactivity Disorder (ADHD) is a common childhood psychiatric disorder with severe and lifelong impact on mental health and socioeconomic achievements. Environmental factors may play a role in the increasing incidens rates. Previous studies on associations between prenatal and childhood exposure to organophosphate and pyrethroid insecticides and ADHD symptoms have yielded mixed findings. OBJECTIVES: To investigate associations between prenatal and childhood exposure to chlorpyrifos and pyrethroids and ADHD symptoms in 5-year-old children from the Odense Child Cohort. METHODS: Spot urine samples from pregnant women in gestational week 28 (n = 614) and offspring at 5 years of age (n = 814) were collected and analyzed for the specific metabolite of chlorpyrifos, TCPY (3,5,6-trichloro-2-pyridinol), as well as the generic pyrethroid metabolite, 3-PBA (3-phenoxybenzoic acid). Offspring ADHD symptoms were assessed at age 5 years using the parent reported "ADHD scale" from the "Child Behavior Checklist 1½-5" (n = 1114). Associations between insecticide exposure variables and an ADHD score ≥90th percentile were analyzed using logistic regression for all children and stratified by sex. RESULTS: Most pregnant women had detectable concentrations of 3-PBA (93%) and TCPY (91%) with median concentrations of 0.20 µg/L and 1.62 µg/L, respectively. In children, 3-PBA and TCPY concentrations were detectable in 88% and 82% of the samples, and the median concentrations were 0.17 and 1.16 µg/L. No statistically significant associations were observed between insecticide metabolites and an ADHD score ≥90th percentile at age 5. CONCLUSION: In this relatively large Danish birth cohort study with mainly low dietary insecticide exposure, we found no statistically significant associations between prenatal or childhood exposure to chlorpyrifos or pyrethroids, and excess ADHD-symptom load, in 5-year-old children. Prospective studies with multiple urine samples across vulnerable windows of neurodevelopment is warranted to improve assessment of safe exposure levels, which is particularly relevant for pyrethroids, since their use is increasing.

Investigating the prevalence of erectile dysfunction among men exposed to organophosphate insecticides.

INTRODUCTION: Erectile dysfunction (ED) poses a significant disease morbidity and contributor to male infertility, where an estimated 20-40% of men are affected annually. While several risk factors have been identified in the etiology of ED (e.g., aging, heart disease, diabetes, and obesity), the complete pathogenesis remains to be elucidated. Over the last few decades, the contribution of environmental exposures to the pathogenesis of ED has gained some attention, though population studies are limited and results are mixed. Among environmental contaminants, organophosphate (OP) insecticides represent one of the largest chemical classes, and chlorpyrifos is the most commonly used OP in the U.S. OP exposure has been implicated in driving biological processes, including inflammation, reactive oxygen species production, and endocrine and metabolism disruption, which have been demonstrated to adversely affect the hypothalamus and testes and may contribute to ED. Currently, studies evaluating the association between OPs and ED within the U.S. general population are sparse. METHODS: Data were leveraged from the National Health and Nutrition Examination Survey (NHANES), which is an annually conducted, population-based cross-sectional study. Urinary levels of 3,5,6-trichloro-2-pyridinol (TCPy), a specific metabolite of the most pervasive OP insecticide chlorpyrifos, were quantified as measures of OP exposure. ED was defined by responses to questionnaire data, where individuals who replied "sometimes able" or "never able" to achieve an erection were classified as ED. Chi-square, analysis of variance (ANOVA), and multivariable, weighted linear and logistic regression analyses were used to compare sociodemographic variables between quartiles of TCPy exposure, identify risk factors for TCPy exposure and ED, and to analyze the relationship between TCPy and ED. RESULTS: A total of 671 adult men were included in final analyses, representing 28,949,379 adults after survey weighting. Approximately 37% of our cohort had ED. Smoking, diabetes, aging, Mexican-American self-identification, and physical inactivity were associated with higher ED prevalence. Analysis of TCPy modeled as a continuous variable revealed nonsignificant associations with ED (OR = 1.02 95% CI [0.95, 1.09]). Stratification of total TCPy into quartiles revealed increased odds of ED among adults in the second and fourth quartiles, using the first quartile as the reference (OR = 2.04 95% CI [1.11, 3.72], OR = 1.51 95% CI [0.58, 3.93], OR = 2.62 95% CI [1.18, 5.79], for quartiles 2, 3, and 4, respectively). CONCLUSIONS: The results of our study suggest a potential role for chlorpyrifos and other OPs the pathogenesis of ED. Future studies are warranted to validate these findings, determine clinical significance, and to investigate potential mechanisms underlying these associations.

Hesperidin counteracts chlorpyrifos-induced neurotoxicity by regulating oxidative stress, inflammation, and apoptosis in rats.

Chlorpyrifos (CPF), considered one of the most potent organophosphates, causes a variety of human disorders including neurotoxicity. The current study was designed to evaluate the efficacy of hesperidin (HSP) in ameliorating CPF-induced neurotoxicity in rats. In the study, rats were treated with HSP (orally, 50 and 100 mg/kg) 30 min after giving CPF (orally, 6.75 mg/kg) for 28 consecutive days. Molecular, biochemical, and histological methods were used to investigate cholinergic enzymes, oxidative stress, inflammation, and apoptosis in the brain tissue. CPF intoxication resulted in inhibition of acetylcholinesterase (AChE) and butrylcholinesterase (BChE) enzymes, reduced antioxidant status [superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione (GSH)], and elevation of malondialdehyde (MDA) levels and carbonic anhydrase (CA) activities. CPF increased histopathological changes and immunohistochemical expressions of 8-OHdG in brain tissue. CPF also increased levels of glial fibrillary acidic protein (GFAP) and nuclear factor kappa B (NF-κB) while decreased levels of nuclear factor erythroid 2-related factor 2 (Nrf-2), heme oxygenase-1 (HO-1) and peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α). Furthermore, CPF increased mRNA transcript levels of caspase-3, Bax, PARP-1, and VEGF, which are associated with apoptosis and endothelial damage in rat brain tissues. HSP treatment was found to protect brain tissue by reducing CPF-induced neurotoxicity. Overall, this study supports that HSP can be used to reduce CPF-induced neurotoxicity.

Effects of cylindrospermopsin, chlorpyrifos and their combination in a SH-SY5Y cell model concerning developmental neurotoxicity.

The cyanotoxin cylindrospermopsin (CYN) has been postulated to cause neurotoxicity, although the studies in this concern are very few. In addition, some studies in vitro indicate its possible effects on development. Furthermore, pesticides can be present in the same environmental samples as cyanotoxins. Therefore, chlorpyrifos (CPF) has been one of the most common pesticides used worldwide. The aim of this report was to study the effects of CYN, isolated and in combination with CPF, in a developmental neurotoxicity in vitro model. The human neuroblastoma SH-SY5Y cell line was exposed during 6 days of differentiation to both toxics to study their effects on cell viability and neurite outgrowth. To further evaluate effects of both toxicants on cholinergic signaling, their agonistic and antagonistic activities on the α7 homomeric nicotinic acetylcholine receptor (nAChR) were studied upon acute exposure. Moreover, a transcriptomic analysis by qPCR was performed after 6 days of CYN-exposure during differentiation. The results showed a concentration-dependent decrease on both cell viability and neurite outgrowth for both toxics isolated, leading to effective concentration 20 (EC20) values of 0.35 µM and 0.097 µM for CYN on cell viability and neurite outgrowth, respectively, and 100 µM and 58 µM for CPF, while the combination demonstrated no significant variations. In addition, 95 µM and 285 µM CPF demonstrated to act as an antagonist to nicotine on the nAChR, although CYN up to 2.4 µM had no effect on the efficacy of these receptors. Additionally, the EC20 for CYN (0.097 µM) on neurite outgrowth downregulated expression of the 5 genes NTNG2 (netrin G2), KCNJ11 (potassium channel), SLC18A3 (vesicular acetylcholine transporter), APOE (apolipoprotein E), and SEMA6B (semaphorin 6B), that are all important for neuronal development. Thus, this study points out the importance of studying the effects of CYN in terms of neurotoxicity and developmental neurotoxicity.

Deciphering the impact of greenhouse pesticides on hepatic metabolism profile: Toxicity experiments on HepG2 cells using chlorpyrifos and emamectin benzoate.

Epidemiological evidence on the health effects of pesticide exposure among greenhouse workers is limited, and the mechanisms are lacking. Building upon our team's previous population study, we selected two pesticides, CPF and EB, with high detection rates, based on the theoretical foundation that the liver serves as a detoxifying organ, we constructed a toxicity model using HepG2 cells to investigate the impact of individual or combined pesticide exposure on the hepatic metabolism profile, attempting to identify targeted biomarkers. Our results showed that CPF and EB could significantly affect the survival rate of HepG2 cells and disrupt their metabolic profile. There were 117 metabolites interfered by CPF exposure, which mainly affected ABC transporter, biosynthesis of amino acids, center carbon metabolism in cancer, fatty acid biosynthesis and other pathways, 95 metabolites interfered by EB exposure, which mainly affected center carbon metabolism in cancer, HIF-1 signaling pathway, valine, leucine and isoleucine biosynthesis, fatty acid biosynthesis and other pathways. The cross analysis and further biological experiments confirmed that CPF and EB pesticide exposure may affect the HIF-1 signaling pathway and valine, leucine and isoleucine biosynthesis in HepG2 cells, providing reliable experimental evidence for the prevention and treatment of liver damage in greenhouse workers.

Biochemical and molecular-level effects of co-exposure to chlorpyrifos and lambda-cyhalothrin on the earthworm (Eisenia fetida).

Farmland soil organisms frequently encounter pesticide mixtures presented in their living environment. However, the underlying toxic mechanisms employed by soil animals to cope with such combined pollution have yet to be explored. This investigation aimed to reveal the changes in cellular and mRNA levels under chlorpyrifos (CPF) and lambda-cyhalothrin (LCT) co-exposures in earthworms (Eisenia fetida). Results exhibited that the combination of CPF and LCT triggered an acute synergistic influence on the animals. Most exposures resulted in significant alterations in the activities of total superoxide dismutase (T-SOD), copper/zinc superoxide dismutase (Cu/Zn-SOD), caspase 3, and carboxylesterase (CarE) compared to the basal level. Moreover, when exposed to chemical mixtures, the transcription levels of four genes [heat shock protein 70 (hsp70), gst, sod, and calreticulin (crt)] also displayed more pronounced changes compared with their individual exposures. These changes in determined parameters indicated the occurrence of oxidative stress, cell death, detoxification dysfunction, and endoplasmic reticulum damage after co-exposure to CPF and LCT in E. fetida. The comprehensive examination of mixture toxicities of CPF and LCT at different endpoints would help to understand the overall toxicity they cause to soil invertebrates. The augmented deleterious effect of these pesticides in a mixture suggested that mixture toxicity assessment was necessary for the safety evaluation and application of pesticide mixtures.

Long-term behavioral and neurochemical paradoxical alterations elicited following intranasal application of a chlorpyrifos formulation in mice.

The intranasal (IN) administration route represents a pathway for xenobiotics to reach the brain. The present study aimed to address the long-term consequences of IN administration of a chlorpyrifos (CPF) commercial formulation (fCPF) in mice. For this purpose, adult male CF-1 mice were intranasally administered with fCPF (10 mg/kg/day) three days a week, for 2 and 4 weeks, respectively. Behavioral and biochemical analyses were conducted 3-7, and 7.5 months after the last IN fCPF administration, respectively. Following a 6-month fCPF-free washout period, fur appearance and body injuries scores improved in the fCPF-treated groups. Notably, spatial learning and memory enhancement was observed 4 and 7 months after the last IN fCPF administration. Changes in oxidative stress markers and the activities of enzymes involved in cholinergic and glutamatergic pathways were observed in different brain areas from fCPF-treated mice, still after 7.5 months from fCPF application. Altogether, these neurochemical disturbances could be responsible for the described behavioral observations.

Interactive transgenerational effects of parental co-exposure to prochloraz and chlorpyrifos: Disruption in multiple biological processes and induction of genotoxicity.

The application of different types of pesticides can result in the coexistence of multiple pesticide residues in our food and the environment. This can have detrimental effects on the health of offspring across generations when parents are exposed to these pesticides. Therefore, it is imperative to understand the long-term effects that can be inherited by future generations when assessing the risks associated with pesticides. To study the genotoxic effects of commonly used pesticides, prochloraz (PRO) and chlorpyrifos (CHL), and assess whether their combined exposures have a different toxic effect, we modeled the transgenerational effects of parental (F0-generation) and/or offspring (F1-generation) exposures on zebrafish embryos in the F1-generation. Following the exposures, we proceeded to assess the impacts of these exposures on a range of biological processes in F1-generation zebrafish. Our results revealed that exposure to PRO and CHL altered multiple biological processes, such as inflammation, apoptosis, oxidative stress, and thyroid hormone synthesis, and detoxification system, providing molecular targets for subsequent studies on toxicity mechanisms. Notably, our study also found that the biological processes of F1-generation zebrafish embryos were altered even though they were not exposed to any pesticide when F0-generation zebrafish were exposed to PRO or CHL, suggesting potential genotoxicity. In conclusion, we provided in-vivo evidence that parental exposure to PRO and/or CHL can induce genotoxicity in the offspring. Moreover, we observed that the toxic effects resulting from the combined exposure were interactive, suggesting a potential synergistic impact on the offspring.

Niosomal hesperidin attenuates the M1/M2-macrophage polarization-based hepatotoxicity followed chlorpyrifos -induced toxicities in mice.

Chlorpyrifos(CPF) is a well-known hepatotoxic agent that has side effects on several organs. On the contrary, hepatic macrophages are crucial in maintaining liver tissue integrity. The main objective of this study was to evaluate the effects and possible mechanisms of niosomal hesperidin (Nio + Hesp), a flavanone glycoside found in citrus fruits, on M1-M2 liver macrophage polarization and inflammatory cells in the brain, liver, and ovarian tissues. Forty C57 mice were divided into CPF(3 mg/kg), Sham(Dimethyl sulfoxide 40 µL/kg), CPF + Hesp(100 mg/kg), and CPF + Nio + Hesp (100 mg/kg) groups. The activity of sera superoxide dismutase (SOD) and malondialdehyde (MDA), brain, liver, and ovary tissues changes, and M1-M2 liver macrophage polarization were evaluated by examining the expression of CD163 and CD68 genes. Hepatic lesions consisting of sporadic foci of coagulation necrosis, inflammatory cell reaction, and regenerative fibrosis were seen following CPF injection, reflected by significant overexpression of CD163 and CD68 genes. In comparison, Nio + Hesp declined the amount of cell apoptosis in the liver and downregulated CD163 and CD68 gene expression. Both Nio + Hesp and Hesp alleviated CPF-induced hepatotoxicity, however, Nio + Hesp was superior to hesperidin in the downregulation of the CD163 and CD68 gene expression. Even though a significant difference between hesperidin and Nio + Hesp was observed in the number of Graafian follicles, corpus luteum, and peri-antral follicles, no substantial difference was observed in primary follicles. The ameliorative effects of Hesp and Nio + Hesp may be at least in part due to their antioxidant and anti-inflammatory properties. These findings showed that both M1- and M2-macrophages contributed to the development of hepatic lesions induced by CPF and provided information about macrophage activation, indicating the importance of analysis of macrophage phenotypes for hepatotoxicity based on M1/M2-polarization which can be downregulated by niosomal nesperidin.

Eucalyptol ameliorates chlorpyrifos-induced necroptosis in grass carp liver cells by down-regulating ROS/NF-κB pathway.

Chlorpyrifos (Diethoxy-sulfanylidene-(3,5,6-trichloropyridin-2-yl) oxy-λ5-phosphane, CPF) was extensively used organophosphorus pesticide, extensively deteriorating public problem with the enrichment in the water bodies. Eucalyptol (1,3,3-Trimethyl-2-oxabicyclo[2.2.2] octane, EUC), a colorless cyclic monoterpene oxide, has shown anti-inflammatory and anti-oxidation properties. To explore the effect of EUC on CPF-induced necroptosis in the grass carp liver cells (L8824 cells), we treated L8824 cells with 60 mM CPF and 5 µM EUC for 24 h. The results showed that CPF exposed lead to excessive accumulation of reactive oxygen species (ROS) and oxidative stress, activating the NF-κB and RIPK1 pathway, increasing the level of cell necroptosis. However, EUC treatment attenuated the toxic effects of CPF treatment on L8824 cells. In summary, the study demonstrated that CPF induced necroptosis and inflammation, and EUC treatment could decrease CPF-caused cell injury.

De novo transcriptome analysis of Protohermes xanthodes Navás (Megaloptera: Corydalidae) reveling the effects of sublethal chlorpyrifos on the expression of cholinergic neuronal genes.

The insect cholinergic neuron system is the target for various pesticides, including organophosphate, carbamate and neonicotinoid pesticides. In this study, we conducted a de novo transcriptome analysis on the aquatic insect Protohermes xanthodes and identified for the first time presenting sixteen genes encoding cholinergic neuronal proteins (PxanChaT, PxanVAChT, PxanmAChR, PxannAChRs, and PxanAChEs), which are candidates for acetylcholine synthesis, transport, reception and degradation in cholinergic synapse. FPKM analysis revealed that these genes are primarily expressed in head and nerve cord of P. xanthodes larvae, and some of them are also abundant in hindgut, malpighian tubules and tracheae. After exposed to different concentrations of sublethal chlorpyrifos (CPF), expression of these cholinergic neuronal genes are generally increased and then decreased with the increase of CPF concentration, except PxannAChRα9 which is increased in both 4.2 and 8.4 µg/L CPF groups. Short-term (96 h) CPF exposure resulted in significant up-regulation of PxanAChE1 expression in P. xanthodes larvae exposed to 0.42 and 4.2 µg/L CPF concentrations, while PxanAChE2 was up-regulated only in 0.42 µg/L CPF group. After long-term (14 d) CPF exposure, PxanAChE1 expression was down-regulated in 0.168 and 0.42 µg/L CPF groups. PxanAChE2 expression was dramatically decreased in all CPF groups. Moreover, acetylcholinesterase (AChE) activity was significantly decreased across all long-term CPF exposure groups. These results suggested that sublethal exposure to CPF can disrupt the expression of cholinergic neuronal genes in P. xanthodes larvae, and implied that long-term sublethal CPF exposure may cause toxic effects on P. xanthodes larvae by inhibiting AChE activity. Furthermore, identification of cholinergic neuronal genes in P. xanthodes provided candidate molecular markers for study the toxic effects of environmental pollutants on the neuron system of an aquatic predatory insect with ecological importance.

Combined pesticides in field doses weaken honey bee (Apis cerana F.) flight ability and analyses of transcriptomics and metabolomics.

Imidacloprid, chlorpyrifos, and glyphosate rank among the most extensively employed pesticides worldwide. The effects of these pesticides and their combined on the flight capability of Apis cerana, and the potential underlying mechanisms remain uncertain. To investigate these effects, we carried out flight mill, transcriptome, and metabolome experiments. Our findings reveal that individual acute oral treatments with pesticides, specifically 20 µL of 10 ng/g imidacloprid (0.2 ng per bee), 30 ng/g chlorpyrifos (0.6 ng per bee), and 60 ng/g glyphosate (1.2 ng per bee), did not impact the flight capability of the bees. However, when bees were exposed to a combination of two or three pesticides, a notable reduction in flight duration and distance was observed. In the transcriptomic and metabolomic analyses, we identified 307 transcripts and 17 metabolites that exhibited differential expression following exposure to combined pesticides, primarily associated with metabolic pathways involved in energy regulation. Our results illuminate the intricate effects and potential hazards posed by combined pesticide exposures on bee behavior. These findings offer valuable insights into the synergistic potential of pesticide combinations and their capacity to impair bee behavior. Understanding these complex interactions is essential for comprehending the broader consequences of pesticide formulations on honey bee populations.

Effects of chlorpyrifos toxicity on brain, pseudobranchial neurosecretory system and swimming performance of a catfish, Heteropneustes fossilis.

In the present study, it was aimed to evaluate the adverse effects of CPF on the histopathology of the optic tectum and cerebellum, pseudobranchial neurosecretory system (PNS), biochemical assays of brain tissue, and locomotory behavior in catfish, Heteropneustes fossilis. The fishes were exposed to an environmentally relevant concentration of 0.09 and 0.192 mg/L of CPF for 7, 15, and 30 d. The CPF toxicity induced degenerative changes with significantly decreased cell size, number, and nucleo-cytoplasmic (N/C) ratio of the PNS; and altered neuro-architectural pattern of optic tectum with degenerative changes in mononuclear and granular cells and necrotic variation in granular and Purkinje cells of the cerebellum. The Acetylcholinesterase (AChE) and Catalase (CAT) activity in the CPF-exposed brain was significantly decreased, whereas Superoxide dismutase (SOD) and Malondialdehyde (MDA) level was significantly increased in comparison with control. In CPF-exposed fishes, the respiratory movements and locomotory behavioral pattern like swimming speed, total distance traveled, time mobile, absolute turn angle, head: distance traveled, maximum speed were significantly decreased, whereas time immobile and time freezing episodes were significantly increased as compared to control fishes. The present study concludes that environmentally relevant concentration of CPF may induce histopathological, biochemical, physiological, and behavioral disturbances in a non-target organism, H. fossilis.

Effect of chlorpyrifos-exposure on the expression levels of CYP genes in Daphnia magna and examination of a possibility that an up-regulated clan 3 CYP, CYP360A8, reacts with pesticides.

Daphnia magna is a test organism used for ecological risk assessments of pesticides, but little is known about the expression levels of cytochrome P450s (CYP)s and their changes after pesticide exposure in the less than 24-h-olds used for ecotoxicity tests. In this study, D. magna juveniles were exposed to 0.2 µg/L of chlorpyrifos under the conditions for acute immobilization test as specified by the OECD test guideline for 24 h, and then the gene expression was compared between the control and chlorpyrifos-exposure groups by RNA-sequencing analysis, with a focus on CYP genes. Among 38 CYP genes expressed in the control group, seven were significantly up-regulated while two were significantly down-regulated in the chlorpyrifos-exposure group. Although the sublethal concentration of chlorpyrifos did not change their expression levels so drastically (0.8 < fold change < 2.6), CY360A8 of D. magna (DmCYP360A8), which had been proposed to be responsible for metabolism of xenobiotics, was abundantly expressed in controls yet up-regulated by chlorpyrifos. Therefore, homology modeling of DmCYP360A8 was performed based on the amino acid sequence, and then molecular docking simulations with the insecticides that were indicated to be metabolized by CYPs in D. magna were conducted. The results indicated that DmCYP360A8 could contribute to the metabolism of diazinon and chlorfenapyr but not chlorpyrifos. These findings suggest that chlorpyrifos is probably detoxified by other CYP(s) including up-regulated and/or constitutively expressed one(s).

Changes in thermal sensitivity of Rhinella arenarum tadpoles (Anura: Bufonidae) exposed to sublethal concentrations of different pesticide fractions (Lorsban® 75WG).

The intensive use of agrochemicals and the rapid increase of global temperatures have modified the thermal conditions of aquatic environments, thus increasing amphibians' vulnerability to global warming and positioning them at great risk. Commercial formulations of chlorpyrifos (COM) are the pesticides most widely used in agricultural activities, with a high toxic potential on amphibians. However, little is known about the separate effects of the active ingredient (CPF) and adjuvants (AD). We studied the thermal sensitivity at different concentrations and pesticide fractions in Rhinella arenarum tadpoles, on thermal tolerance limits (CTmax = Critical thermal maximum and CTmin = Critical thermal minimum), swimming speed (Ss), Optimum temperature (Top), and Thermal breadth 50 (B50). Our results demonstrate that the pesticide active ingredient, the adjuvants, and the commercial formulation of chlorpyrifos differentially impair the thermal sensitivity of R. arenarum tadpoles. The pesticide fractions affected the heat and the cold tolerance (CTmax and CTmin), depending on the concentrations they were exposed to. The locomotor performance (Ss, Top, and B50) of tadpoles also varied among fractions, treatments, and environmental temperatures. In the context of climate change, the outcomes presented are particularly relevant, as mean temperatures are increasing at unprecedented rates, which suggests that tadpoles inhabiting warming and polluted ponds are currently experiencing deleterious conditions. Considering that larval stages of amphibians are the most susceptible to changing environmental conditions and the alarming predictions about environmental temperatures in the future, it is likely that the synergism between high temperatures and pesticide exposure raise the threat of population deletions in the coming years.

Urtica dioica Extract Abrogates Chlorpyrifos-Induced Toxicity in Zebrafish Larvae.

Chlorpyrifos (CPF) is a widely used organophosphate insecticide, though its excessive use causes environmental contamination, raising concerns about its adverse effects on human health. In this regard, Urtica dioica stands out as a promising candidate for counteracting chemical 'contaminant' toxicity thanks to its therapeutic properties. Therefore, our study aimed to investigate the potential of an Urtica dioica ethanolic extract (UDE) to mitigate chlorpyrifos-induced toxicity. Eight compounds in the Urtica dioica ethanolic extract have been identified, most of which present significant potential as antioxidant, anti-inflammatory, and neuroprotective agents. Chlorpyrifos exposure altered hatching rates, increased the incidence of teratogenic effects, and upregulated the expression of brain-derived neurotrophic factor (Bdnf) in zebrafish larvae telencephalon. On the other hand, UDE demonstrated a preventive effect against CPF-induced teratogenicity, which is expressed by a lower morphological deformity rate. Moreover, the UDE showed a rather protective effect, maintaining the physiological condition of the telencephalon. Additionally, CPF altered the locomotor behavior of larvae, which was characterized by irregular swimming and increased activity. This defective behavioral pattern was slightly attenuated by the UDE. Our findings suggest that the UDE possesses significant protective properties against CPF-induced toxicity, probably conferred by its natural antioxidant and anti-inflammatory contents. Still, further research is needed to elucidate the recruited mechanisms and implicated pathways on UDE's protective effects.

Molecular docking analysis of chlorpyrifos at the human α7-nAChR and its potential relationship with neurocytoxicity in SH-SY5Y cells.

The organophosphate insecticide chlorpyrifos (CPF), an acetylcholinesterase inhibitor, has raised serious concerns about human safety. Apart from inducing synaptic acetylcholine accumulation, CPF could also act at nicotinic acetylcholine receptors, like the α7-isoform (α7-nAChR), which could potentially be harmful to developing brains. Our aims were to use molecular docking to assess the binding interactions between CPF and α7-nAChR through, to test the neurocytotoxic and oxidative effects of very low concentrations of CPF on SH-SY5Y cells, and to hypothesize about the potential mediation of α7-nAChR. Docking analysis showed a significant binding affinity of CPH for the E fragment of the α7-nAChR (ΔGibbs: -5.63 to -6.85 Kcal/mol). According to the MTT- and Trypan Blue-based viability assays, commercial CPF showed concentration- and time-dependent neurotoxic effects at a concentration range (2.5-20 µM), ten-folds lower than those reported to have crucial effects for sheer CPF. A rise of the production of radical oxygen species (ROS) was seen at even lower concentrations (1-2.5 µM) of CPF after 24h. Notably, our docking analysis supports the antagonistic actions of CPF on α7-nAChR that were recently published. In conclusion, while α7-nAChR is responsible for neuronal survival and neurodevelopmental processes, its activity may also mediate the neurotoxicity of CPF.