Association between exposure to organophosphate pesticides and cytokine levels in a population of flower workers in Mexico.

The ability of organophosphate pesticides to disturb immune function has been demonstrated by in vivo and in vitro studies, but evidence of such effects on humans remains scarce. To assess the association between organophosphate pesticides exposure and cytokine levels in Mexican flower workers, a cross-sectional study was carried out. A questionnaire was provided to 121 male flower workers, and urine and blood samples were collected. Using gas chromatography, urinary concentrations of dialkylphosphate metabolites were determined. The serum cytokine levels, IL-4, IL-5, IL-6, IL-8, and IL-10, were measured using multiplex analysis, and levels of INF-γ and TNF-α by ELISA. We found that a higher dialkylphosphate concentration decreased the pro-inflammatory cytokines INF-γ (ß = -0.63; 95 % CI: -1.22, -0.05), TNF-α (ß= -1.18; 95 % CI: -2.38, 0.02), and IL-6 (ß= -0.59; 95 % CI: -1.29, 0.12), and increased IL-10 (ß=0.56; 95 % CI: 0.02, 1.09), the main anti-inflammatory cytokine, suggesting an imbalance of the immune response in flower workers.

Methylated dialkylphosphate metabolites of the organophosphate pesticide malathion modify actin cytoskeleton arrangement and cell migration via activation of Rho GTPases Rac1 and Cdc42.

The non-cholinergic molecular targets of organophosphate (OP) compounds have recently been investigated to explain their role in the generation of non-neurological diseases, such as immunotoxicity and cancer. Here, we evaluated the effects of malathion and its dialkylphosphate (DAP) metabolites on the cytoskeleton components and organization of RAW264.7 murine macrophages as non-cholinergic targets of OP and DAPs toxicity. All OP compounds affected actin and tubulin polymerization. Malathion, dimethyldithiophosphate (DMDTP) dimethylthiophosphate (DMTP), and dimethylphosphate (DMP) induced elongated morphologies and the formation of pseudopods rich in microtubule structures, and increased filopodia formation and general actin disorganization in RAW264.7 cells and slightly reduced stress fibers in the human fibroblasts GM03440, without significantly disrupting the tubulin or vimentin cytoskeleton. Exposure to DMTP and DMP increased cell migration in the wound healing assay but did not affect phagocytosis, indicating a very specific modification in the organization of the cytoskeleton. The induction of actin cytoskeleton rearrangement and cell migration suggested the activation of cytoskeletal regulators such as small GTPases. We found that DMP slightly reduced Ras homolog family member A activity but increased the activities of Ras-related C3 botulinum toxin substrate 1 (Rac1) and cell division control protein 42 (Cdc42) from 5 min to 2 h of exposure. Chemical inhibition of Rac1 with NSC23766 reduced cell polarization and treatment with DMP enhanced cell migration, but Cdc42 inhibition by ML-141 completely inhibited the effects of DMP. These results suggest that methylated OP compounds, especially DMP, can modify macrophage cytoskeleton function and configuration via activation of Cdc42, which may represent a potential non-cholinergic molecular target for OP compounds.

Pesticides exposure in pregnant Argentinian women: Potential relations with the residence areas and the anthropometric neonate parameters.

Intrauterine environment is the first chemical exposure scenario in life, through transplacental transfer. In this study, the aim was to determine concentrations of organochlorine pesticides (OCPs) and selected current use pesticides in the placentas of pregnant women from Argentina. Socio-demographic information, the mother's lifestyle and neonatal characteristics were also analysed and related to pesticides residue concentrations. Thus, 85 placentas were collected at birth, from an area of intensive fruit production for the international market, in Patagonia Argentina. Concentrations of 23 pesticides including, trifluralin (herbicide), the fungicides chlorothalonil and HCB, and the insecticides chlorpyrifos, HCHs, endosulfans, DDTs, chlordanes, heptachlors, drins and metoxichlor, were determined by GC-ECD and GC-MS. Results were first analysed all together and then grouped by their residential settings, in urban and rural groups. Total mean pesticide concentration was 582.6 ± 1034.4 ng/g lw, where DDTs (325.9 ± 950.3 ng/g lw) and chlorpyrifos (188.4 ± 365.4 ng/g lw) showed a high contribution. Pesticide levels found exceeded those reported in low, middle and high income countries from Europe, Asia and Africa. In general, pesticides concentrations were not associated with neonatal anthropometric parameters. When the results were analysed by residence place, significantly higher concentrations of total pesticides and chlorpyrifos (Mann Whitney test p = 0.0003 and p = 0.032, respectively) were observed in placentas collected from mothers living in rural settings compared to urban areas. Rural pregnant women presented the highest pesticide burden (5.9 µg), where DDTs and chlorpyrifos were the major constituents. These results suggested that all pregnant women are highly exposed to complex pesticide mixtures, including banned OCPs and the widely used chlorpyrifos. Based on the pesticide concentrations found, our results warn of possible health impacts from prenatal exposure through transplacental transfer. This is one of the first reports of both chlorpyrifos and chlorothalonil concentrations in placental tissue, and contributes to the knowledge of current pesticide exposure in Argentina.

Temephos, an organophosphate larvicide for residential use: a review of its toxicity.

Temephos (O,O,O',O'-tetramethyl O,O'-thiodi-p-phenylene bis(phosphorothioate)) is a larvicide belonging to the family of organophosphate pesticides used for the control of different vectors of diseases, such as dengue, Zika, chikungunya, and dracunculiasis. The aim of this review was to discuss the available published information about temephos toxicokinetics and toxicity in mammals. Temephos is quickly absorbed in the gastrointestinal tract, distributed to all organs, and then it accumulates mainly in adipose tissue. It is metabolized by S-oxidation, oxidative desulfuration, and hydrolysis reactions, with the possible participation of cytochrome P450 (CYP). Temephos is mainly eliminated by feces, whereas some of its metabolites are eliminated by urine. The World Health Organization classifies it as class III: slightly dangerous with a NOAEL (no-observed adverse effect level) of 2.3 mg/kg/day for up to 90 days in rats, based on brain acetylcholinesterase (AChE) inhibition. A LOAEL (lowest observable adverse effect level) of 100 mg/kg/day for up to 44 days in rats was proposed based on cholinergic symptoms. However, some studies have shown that temephos causes toxic effects in mammals. The inhibition of the enzyme acetylcholinesterase (AChE) is one of its main demonstrated effects; however, this larvicide has also shown genotoxic effects and some adverse effects on male reproduction and fertility, as well as liver damage, even at low doses. We performed an extensive review through several databases of the literature about temephos toxicokinetics, and we recommend to revisit current assessment of temephos with the new available data.

The synergistic effect of microplastic and malathion exposure on fiddler crab Minuca ecuadoriensis microplastic bioaccumulation and survival.

We assessed the combined effects of polyethylene microplastic (MP) and malathion (MLT) on the survival of the fiddler crab Minuca ecuadoriensis, and MP tissue bioaccumulation in four treatments following 120 h exposure: T1) Control; T2) MLT 50 mg L-1; T3) MP 200 mg L-1; and T4) MLT (50 mg L-1) + MP (200 mg L-1). The highest mortality (80%) was in T4, followed by T2 (28%) and no mortality was in T3. Higher MP bioaccumulation was observed in T4 (572 items g tissue-1) followed by T3 (70 items g tissue-1). Our findings indicate that the synergistic effect of MLT and MP increased M. ecuadoriensis bioaccumulative capacity and decreases survival. Thus, as MP contamination in aquatic environments is ubiquitous, our study raises a warning on the synergistic effects of MP with other environmental contaminants and serves as a baseline for further studies.

Integrating multiple lines of evidence to assess freshwater ecosystem health in a tropical river basin.

Degradation of freshwater ecosystems by uncontrolled human activities is a growing concern in the tropics. In this regard, we aimed at testing an integrative framework based on the IFEQ index to assess freshwater ecosystem health of river basins impacted by intense livestock and agricultural activities, using the Muchacho River Basin (MRB) as a case study. The IFEQ combine multiple lines of evidence such as riverine hydromorphological analysis (LOE 1), physicochemical characterization using ions and pesticides (LOE 2), aquatic macroinvertebrate monitoring (LOE 3), and phytotoxicological essays with L. sativa (LOE 4). Overall, results showed an important reduction in streamflow and an elevated increase in ion concentrations along the MRB caused by deforestation and erosion linked to agricultural and livestock activities. Impacts of the high ion concentrations were evidenced in macroinvertebrate communities as pollution-tolerant families, associated with high conductivity levels, represented 92 % of the total abundance. Pollution produced by organophosphate pesticides (OPPs) was critical in the whole MRB, showing levels that exceeded 270-fold maximum threshold for malathion and 30-fold for parathion, the latter banned in Ecuador. OPPs concentrations were related to low germination percentages of Lactuca sativa in sediment phytotoxicity tests. The IEFQ index ranged from 44.4 to 25.6, indicating that freshwater ecosystem conditions were "bad" at the headwaters of the MRB and "critical" along the lowest reaches. Our results show strong evidence that intense agricultural and livestock activities generated significant impacts on the aquatic ecosystem of the MRB. This integrative approach better explains the cumulative effects of human impacts, and should be replicated in other basins with similar conditions to help decision-makers and concerned inhabitants generate adequate policies and strategies to mitigate the degradation of freshwater ecosystems.

Pilot study of exposure of the male population to organophosphate and pyrethroid pesticides in a region of high agricultural activity (Córdoba, Argentina).

Urinary metabolites of organophosphate (OP) and pyrethroid (PYR) pesticides and biomarkers of effects were studied in a population (n=40) residing in an important agricultural area of the province of Córdoba (Argentina). Detection frequencies (DF) higher than 85% were observed for the metabolites of pirimiphos (2-diethylamino-6-methylpyrimidin-4-ol -DEAMPY-, median 7.5 µg/g creatinine, DF: 100%), parathion (p-nitrophenol, 0.99 µg/g creatinine, 100%), and chlorpyrifos (3,5,6-trichloro-2-pyridinol, 0.25 µg/g creatinine, 85%). The DEAMPY concentrations doubled the levels found in other studies and were negatively associated with Er-AChE activity, suggesting the appearance of health effects already in environmental exposure levels below established acceptable daily intakes (ADIs). 3-Phenoxybenzoic acid, the metabolite of several PYR pesticides, was also found in all samples. This metabolite was also significantly negatively correlated with Er-AChE, indicating effects of pyrethroid pesticides on the acetylcholine system even at concentrations below the ADI.

Chlorpyrifos and Dimethoate in Water and Sediments of Agricultural Drainage Ditches in Northern Sinaloa, Mexico.

In northern Sinaloa state, Mexico, little is known on organophosphate pesticide transport and fate in agricultural drainage systems. Spatial and temporal variation of chlorpyrifos and dimethoate was assessed in two agricultural drainage ditches (Buenaventura and Burrión) and risk for aquatic life was estimated. Analysis was made by high performance liquid chromatography and risk estimates were determined following international reference frameworks. In water, the highest chlorpyrifos concentration in the Buenaventura ditch was 5.49 µg L-1, and 3.43 µg L-1 in the Burrión ditch. Dimethoate was quantified only once in both ditches (0.44 µg L-1 and 0.49 µg L-1). In sediment, chlorpyrifos was quantified only in the Burrión ditch (242 µg kg-1). Chlorpyrifos concentrations surpassed water and sediment quality criteria, representing a hazard for environmental and human health, as both ditches discharge into the Gulf of California and are used for capture of commercial species such as the grey mullet (Mugil cephalus) and cauque prawn (Macrobrachium americanum).

Relationship between internal and external factors and the activity of PON1.

Paraoxonase 1 (PON1) is an A-esterase calcium-dependent enzyme that is associated with high-density lipoprotein (HDL) and capable of hydrolyzing a wide variety of substrates, including organophosphate (OP) pesticides. The PON1 phenotype can be modulated by multiple internal and external factors, thereby affecting the catalytic capacity of the enzyme. The aim of this study was to evaluate factors that could modulate PON1 activity in a sample occupationally exposed to pesticides. A cross-sectional, descriptive, and analytical study was carried out with 240 workers. The participants were stratified according to their level of pesticide exposure as reference, moderate-exposure, and high-exposure groups. PON1 activities (arylesterase/AREase, CMPAase, and ssPONase (salt-stimulated)) were determined by spectrophotometry, and the Q192R and L55MPON1 genotypes by real-time PCR. The most frequent genotypes were heterozygous (QR) and homozygous (LL) for PON1Q192R and PON1L55M polymorphisms, respectively. The internal factors associated with the activity of PON1 were the PON1 genotypes (55 and 192) and biochemical parameters related to the lipid profile, in contrast, various external factors related to diet and harmful habits as well as with exposure to pesticides were associated with the activity of PON1. However, using a multivariate mixed ordinal regression model, we found a significant reduction of ssPONase activity in the high-exposure group compared with the reference group only in haplotypes QQLL and RRLL.

Effects of exposure to malathion on blood glucose concentration: a meta-analysis.

Exposure to malathion (an organophosphate pesticide widely used around the world) has been associated with alterations in blood glucose concentration in animal models. However, the results are inconsistent. The aim of this meta-analysis was to evaluate whether malathion exposure can disturb the concentrations of blood glucose in exposed rats. We performed a literature search of online databases including PubMed, EBSCO, and Google Scholar and reviewed original articles that analyzed the relation between malathion exposure and glucose levels in animal models. The selection of articles was based on inclusion and exclusion criteria. The database search identified thirty-five possible articles, but only eight fulfilled our inclusion criteria, and these studies were included in the meta-analysis. The effect of malathion on blood glucose concentration showed a non-monotonic dose-response curve. In addition, pooled analysis showed that blood glucose concentrations were 3.3-fold higher in exposed rats than in the control group (95% CI, 2-5; Z = 3.9; p < 0.0001) in a random-effect model. This result suggested that alteration of glucose homeostasis is a possible mechanism of toxicity associated with exposure to malathion.

Efficient Nucleophilic Degradation of an Organophosphorus Pesticide "Diazinon" Mediated by Green Solvents and Microwave Heating.

An efficient strategy for the degradation of organophosphate pesticide Diazinon was investigated. In this work, ionic liquids, bio-based solvents, and two conventional organic solvents were used as reaction media. Kinetics studies by means of half-life (t1/2,h) were followed by 31P NMR and the products analyzed by GC-MS, HPLC-MS and NMR techniques. These results have shown that t1/2 values in ionic liquids were the lowest and also they were able to activate two electrophilic centers in Diazinon, whilst degradation in bio-based solvents occurred slowly by only an aromatic pathway. In addition, a study to estimate the influence of green activation techniques was carried out by using Ultrasound irradiation and Microwave heating in combination with greener solvents and two conventional organic solvents. Under Microwave heating, faster degradation than under ultrasound irradiation was found. Finally, considering both families of solvent used here and their behavior under green activation techniques, we propose that the more efficient way for degradation of Diazinon with piperidine is by microwave heating using ionic liquids as solvents.

Evaluación de la exposición a órganofosforados y carbamatos en trabajadores de una comunidad agraria / Exposure to organophosphates and carbamates in workers of an agricultural community

En Venezuela la utilización de organofosforados y carbamatos ha contribuido a generar un escenario propicio para la aparición de intoxicaciones agudas y crónicas a nivel de la población, transformándose esta situación en un problema de Salud Pública. Se analizó la exposición en trabajadores de una comunidad agraria de la Colonia Tovar, Estado Aragua. Estudio descriptivo-correlacional en trabajadores expuestos a plaguicidas. El Grupo Expuesto (GE) 17 trabajadores (58,8% del sexo masculino y 41,2%femenino), edad promedio de 37,06 ± 15,66 años y un Grupo Control (GC) con 13 trabajadores del personal administrativo de la Universidad de Carabobo (53,8% del sexo masculino y 46,2%femenino), edad promedio 39,77 ± 13,23 años. Se realizó una entrevista y se determinaron niveles de colinesterasa sérica y parámetros hematimétricos y bioquímicos. En el GE un valor promedio de la actividad de la colinesterasa de 6,7465 ± 1,0314 U/L y para el GC de 8,6546 ± 1,6014 U/L. Las medias de la actividad de la colinesterasa sérica se encontraron dentro de los parámetros considerados normales (4,970-13,977U/L) para ambos grupos. Se evidencia la existencia de diferencia estadísticamente significativa (p< 0,05) entre las medias para el biomarcador de efecto en estudio en el GE. La manipulación inadecuada de los plaguicidas y los síntomas asociados a manifestaciones clínicas debido a la exposición. Un 41,2% de los empleados utilizan medidas de protección personal. Los trabajadores expuestos a organofosforados y carbamatos presentaron biomarcadores de exposición y de efecto dentro de los rangos normales, con presencia de síntomas que pudieran relacionarse con dicha exposición.

In Venezuela, the use of organophosphates and carbamates has contributed to generate a suitable scenario for the appearance of acute and chronic poisoning in the population, turning this situation into a Public Health problem. The exposure in an agrarian worker community from La Colonia Tovar - Aragua State was analyzed. This is a descriptive-correlational study in workers exposed to pesticides. The exposed group, (EG) 17 workers (58.8% male and 41.2% female), mean age 37.06 ± 15.66 years old, and a control group (CG) with 13 administrative staff from University of Carabobo (53.8% male and 46.2% female), mean age 39.77 ± 13.23 years old. An interview was conducted and serum cholinesterase levels were determined; and even, hematimetric and biochemical parameters. In the EG an average value of the cholinesterase activity was 6,7465 ± 1,0314 U/L, and for the CG 8,6546 ± 1,6014 U/L. The means of serum cholinesterase activity were normal (4,970 - 13,977 U/L) for both groups. It is evident the existence of statistically significant difference (p<0.05) between the means for the effect biomarker under study in the EG. The inappropriate handling of pesticides and symptoms associated with clinical manifestations due to exposure. 41.2% of the employees use personal protection measures. Workers exposed to organophosphates and carbamates presented biomarkers of exposure and effect within normal ranges, with the presence of symptoms related to this exposur.

Detection of trace levels of organophosphate pesticides using an electronic tongue based on graphene hybrid nanocomposites.

Organophosphate (OP) compounds impose significant strains on public health, environmental/food safety and homeland security, once they have been widely used as pesticides and insecticides and also display potential to be employed as chemical warfare agents by terrorists. In this context, the development of sensitive and reliable chemical sensors that would allow in-situ measurements of such contaminants is highly pursued. Here we report on a free-enzyme impedimetric electronic tongue (e-tongue) used in the analysis of organophosphate pesticides comprising four sensing units based on graphene hybrid nanocomposites. The nanocomposites were prepared by reduction of graphene oxide in the presence of conducting polymers (PEDOT:PSS and polypyrrole) and gold nanoparticles (AuNPs), which were deposited by drop casting onto gold interdigitated electrodes. Impedance spectroscopy measurements were collected in triplicate for each sample analyzed, and the electrical resistance data were treated by Principal Component Analysis (PCA), revealing that the system was able to discriminate OPs at nanomolar concentrations. In addition, the electronic tongue system could detect OPs in real samples, where relations between the principal components and the variation of pesticides in a mixture were established, proving to be useful to analyze and monitor mixtures of OP pesticides. The materials employed provided sensing units with high specific surface area and high conductivity, yielding the development of a sensor with suitable stability, good reproducibility, and high sensitivity towards pesticide samples, being able to discriminate concentrations as low as 0.1nmolL-1. Our results indicate that the e-tongue system can be used as a rapid, simple and low cost alternative in the analyses of OPs pesticide solutions below the concentration range permitted by legislation of some countries.

Occupational predictors of urinary dialkyl phosphate concentrations in Mexican flower growers.

Background Flower growers have high potential for exposures to pesticides. Occupational factors, such as tasks performed, the production method (organic or conventional), the use of personal protective equipment (PPE), and workplace characteristics influence the intensity of pesticide exposure. Objective To evaluate occupational characteristics affecting urinary concentration of dialkylphosphate (DAP) metabolites of organophosphate pesticides among a group of Mexican floricultural workers. Methods A questionnaire was administered to 117 workers who also provided a first morning urine sample. According to tasks performed and the production methods, pesticide contact was defined as low, medium, or high. PPE use was categorized as acceptable, fairly acceptable, and unacceptable. Urinary concentration of DAP metabolites were determined using gas-liquid chromatography. Association between occupational characteristics and DAP urinary concentrations was assessed by means of linear regression models. Results After adjusting for potential confounders, the workers in the medium and high contact categories had significantly higher DAP concentrations than those in the low contact category (ß: 0.3, CI 95%: 0.1-0.5). Greenhouse workers had greater DAP concentrations than outdoors workers (ß: 0.3, CI 95%: 0.1-0.5). Compared with non-acceptable use of PPE, acceptable use of PPE was associated with lower DAP concentrations (ß: -0.4, CI 95% -0.6 to -0.1). Conclusion Improved safety training is needed for correct PPE usage, especially among flower growers who use conventional pest control methods and who work in a greenhouse environment.

Multiresidue method for simultaneous analysis of aflatoxin M1, avermectins, organophosphate pesticides and milbemycin in milk by ultra-performance liquid chromatography coupled to tandem mass spectrometry.

A method developed for the simultaneous analysis of aflatoxin M1, abamectin, doramectin, eprinomectin, ivermectin, moxidectin, acephate, azinphos-ethyl, azinphos-methyl, diazinon, methamidophos, methidathion, mevinphos, pirimiphos-ethyl and pirimiphos-methyl in whole raw milk, based on the QuEChERS method for extraction and clean-up, with detection and quantification by ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) is described. The method was validated according to parameters of the Analytical Quality Assurance Manual from the Brazilian Ministry of Agriculture and Commission Decision 2002/657/EC, and proved suitable for analysis of these analytes within the proposed working range, with recovery values between 77% and 110%, a standard deviation lower than 20%, limits of detection between 0.05 and 0.99 µg l(-)(1), and limits of quantification between 0.15 and 1.98 µg l(-1). Samples from animals treated with abamectin, doramectin, ivermectin and diazinon were analysed by the validated method. Residues of aflatoxin M1 were also found in field samples at levels below the established maximum residue limit.

Chronic exposure to organophosphate (OP) pesticides and neuropsychological functioning in farm workers: a review.

BACKGROUND: Previous studies have demonstrated that acute poisoning from exposure to organophosphate (OP) pesticides in agricultural workers causes adverse health effects. However, neuropsychological and cognitive effects of chronic occupational exposure to OP pesticides remain controversial. OBJECTIVE: To identify, evaluate, and systematize existing evidence regarding chronic exposure to OP pesticides and neuropsychological effects in farmworkers. METHODS: Using the PubMed search engine, a systematic review process was implemented and replicated according to the PRISMA statement. Eligibility criteria included workers over 18 years of age exposed to OP pesticides as well as assessment of neuropsychological and cognitive functioning. Search terms were in English and Spanish languages and included organophosphate and workers. RESULTS: Of the search results, 33 of 1,256 articles meet eligibility criteria. Twenty-four studies found an association between chronic occupational exposure to OP pesticides and low neuropsychological performance in workers. We classified nine of the studies to have study design limitations. Studies indicated occupational exposure to OP pesticides is linked to difficulties in executive functions, psychomotor speed, verbal, memory, attention, processing speed, visual-spatial functioning, and coordination. Nine studies find no relationship between OP pesticides exposure and neuropsychological performance. CONCLUSIONS: Overall, evidence suggests an association between chronic occupational exposure to OP pesticides and neuropsychological effects. However, there is no consensus about the specific cognitive skills affected.

Alteration of syncytiotrophoblast mitochondria function and endothelial nitric oxide synthase expression in the placenta of rural residents.

The impact of environmental organophosphate (OP) pesticide exposure on respiratory complexes, enzymatic antioxidant defense activities, and oxidative damage markers in the syncytiotrophoblast and cytotrophoblast mitochondria was evaluated. Placental progesterone (PG) levels and endothelial nitric oxide synthase (eNOS) expression were studied. Samples from women non-exposed (control group-CG) and women living in a rural area (rural group-RG) were collected during pesticide spraying season (RG-SS) and non-spraying season (RG-NSS). In RG-SS, the exposure biomarker placental carboxylesterase decreased and syncytiotrophoblast cytochrome c oxidase activity increased, while 4-hydroxynonenal levels decreased. PG levels decreased in RG-SS and in the RG. Nitric oxide synthase expression decreased in RG, RG-SS and RG-NSS. No significant changes in mitochondrial antioxidant enzyme activities were found. These results suggest that the alteration of syncytiotrophoblast mitochondrial complex IV activity and steroidogenic function may be associated to pesticide exposure. Reduction in placental PG and eNOS expression may account for low newborn weight in RG.

B-esterase determination and organophosphate insecticide inhibitory effects in JEG-3 trophoblasts.

The placenta and trophoblasts express several B-esterases. This family includes acethylcholinesterase (AChE), carboxylesterase (CES) and butyrylcholinesterase (BChE), which are important targets of organophosphate insecticide (OP) toxicity. To better understand OP effects on trophoblasts, B-esterase basal activity and kinetic behavior were studied in JEG-3 choriocarcinoma cell cultures. Effects of the OP azinphos-methyl (Am) and chlorpyrifos (Cp) on cellular enzyme activity were also evaluated. JEG-3 cells showed measurable activity levels of AChE and CES, while BChE was undetected. Recorded Km for AChE and CES were 0.33 and 0.26 mM respectively. Native gel electrophoresis and RT-PCR analysis demonstrated CES1 and CES2 isoform expression. Cells exposed for 4 and 24 h to the OP Am or Cp, showed a differential CES and AChE inhibition profiles. Am inhibited CES and AChE at 4 h treatment while Cp showed the highest inhibition profile at 24 h. Interestingly, both insecticides differentially affected CES1 and CES2 activities. Results demonstrated that JEG-3 trophoblasts express AChE, CES1 and CES2. B-esterase enzymes were inhibited by in vitro OP exposure, indicating that JEG-3 cells metabolization capabilities include phase I enzymes, able to bioactivate OP. In addition, since CES enzymes are important for medicinal drug activation/deactivation, OP exposure may interfere with trophoblast CES metabolization, probably being relevant in a co-exposure scenario during pregnancy.

Organophosphate pesticide method development and presence of chlorpyrifos in the feet of nearctic-neotropical migratory songbirds from Canada that over-winter in Central America agricultural areas.

Recent modeling analysis suggests that numerous birds may be at risk of acute poisoning in insecticide-treated fields. Although the majority of avian field studies on pesticides have focused on treated seed, granule, insect or vegetation (oral exposure) ingestion, dermal exposure is an important exposure route when birds come into contact with deposited pesticides on foliage and other surfaces. Some nearctic-neotropical migratory songbirds are likely exposed to pesticides on their non-breeding habitats and include treated crops, plantations or farmlands. In the present study, we developed a method for four environmentally-relevant organophosphate (OP) pesticides (fenthion, fenamiphos, chlorpyrifos and diazinon) in the feet of migratory songbirds (i.e. Common yellowthroat, Gray catbird, Indigo bunting, America redstart, Northern waterthrush, Northern parula, and an additional 12 species of warblers). A total of 190 specimens of the 18 species of songbirds were sampled from available window-killed birds (spring of 2007 and 2011) in downtown Toronto, Canada. The species that were available most likely over-wintered in Mexican/Central American crops such as citrus, coffee and cacao. The feet of the dead birds were sampled and where OP foot exposure likely occurred during over-wintering foraging on pesticide-treated crops. Chlorpyrifos was the only measurable OP (pg mg feet weight(-1)) and in the 2011-collected feet of Black throated blue warbler (0.5), Tennessee warbler (1.0), Northern parula (1.2), Northern waterthrush (0.6), Common yellowthroat (1.0) and the Blue winged warbler (0.9). Dermal contact with OP pesticides during over-wintering in agricultural areas resulted in low levels of chlorpyrifos and long time retention on the feet of a subset of songbirds.

Methamidophos alters sperm function and DNA at different stages of spermatogenesis in mice.

Methamidophos (MET) is a highly toxic organophosphate (OP) pesticide that is widely used in developing countries. MET has male reproductive effects, including decreased fertility. We evaluated MET effects on sperm quality, fertilization and DNA integrity, exploring the sensitivity of different stages of spermatogenesis. Adult male mice received MET (3.75 or 5mg/kg-bw/ip/day/4 days) and were euthanized 1, 28 or 45 days post-treatment (dpt) to evaluate MET's effects on epididymal maturation, meiosis or mitosis, respectively. Spermatozoa were obtained from the cauda epididymis-vas deferens and were evaluated for sperm quality, acrosome reaction (AR; Coomassie staining), mitochondrial membrane potential (by JC-1), DNA damage (comet assay), oxidative damage (malondialdehyde (MDA) production), in vitro fertilization and protein phosphorylation (immunodetection), and erythrocyte acetylcholinesterase (AChE) activity. At 1-dpt, MET inhibited AChE (43-57%) and increased abnormal cells (6%). While at 28- and 45-dpt, sperm motility and viability were significantly reduced with an increasing MET dose, and abnormal morphology increased at 5mg/kg/day/4 days. MDA and mitochondrial activity were not affected at any dose or time. DNA damage (OTM and %DNA) was observed at 5mg/kg/day/4 days in a time-dependent manner, whereas both parameters were altered in cells from mice exposed to 3.75 mg/kg/day/4 days only at 28-dpt. Depending on the time of collection, initial-, spontaneous- and induced-AR were altered at 5mg/kg/day/4 days, and the fertilization capacity also decreased. Sperm phosphorylation (at serine and tyrosine residues) was observed at all time points. Data suggest that meiosis and mitosis are the more sensitive stages of spermatogenesis for MET reproductive toxicity compared to epididymal maturation.