Monitoring of DNA structural changes after incorporation of the phenylpyrazole insecticide fipronil.

The use of insecticides presents a risk to the environment because they can accumulate in the water, soil, air, and organisms, endangering human and animal health. It is therefore essential to investigate the effects of different groups of insecticides on individual biomacromolecules such as DNA. We studied fipronil, which belongs to the group of phenylpyrazole insecticides. The interaction of fipronil with calf thymus DNA was investigated using spectroscopic methods (absorption and fluorescence spectroscopy) complemented with infrared spectroscopy and viscosity measurement. Fluorescence emission spectroscopy showed the formation of a fipronil/DNA complex with a combined static and dynamic type of quenching. The binding constant was 4.15 × 103 L/mol. Viscosity changes were recorded to confirm/disconfirm the intercalation mode of interaction. A slight change in DNA viscosity in the presence of fipronil was observed. The phenylpyrazole insecticide does not cause significant conformational changes in DNA structure or increase of its chain length. We hypothesize that fipronil is incorporated into the minor groove of the DNA macromolecule via hydrogen interactions as indicated by FT-IR and CD measurements.

Fipronil disturbs the antigen-specific immune responses and GABAergic gene expression in the ovalbumin-immunized BALB/c mice.

BACKGROUND: Fipronil (FPN) is a broad-spectrum pesticide and commonly known as low toxicity to vertebrates. However, increasing evidence suggests that exposure to FPN might induce unexpected adverse effects in the liver, reproductive, and nervous systems. Until now, the influence of FPN on immune responses, especially T-cell responses has not been well examined. Our study is designed to investigate the immunotoxicity of FPN in ovalbumin (OVA)-sensitized mice. The mice were administered with FPN by oral gavage and immunized with OVA. Primary splenocytes were prepared to examine the viability and functionality of antigen-specific T cells ex vivo. The expression of T cell cytokines, upstream transcription factors, and GABAergic signaling genes was detected by qPCR. RESULTS: Intragastric administration of FPN (1-10 mg/kg) for 11 doses did not show any significant clinical symptoms. The viability of antigen-stimulated splenocytes, the production of IL-2, IL-4, and IFN-γ by OVA-specific T cells, and the serum levels of OVA-specific IgG1 and IgG2a were significantly increased in FPN-treated groups. The expression of the GABAergic signaling genes was notably altered by FPN. The GAD67 gene was significantly decreased, while the GABAR ß2 and GABAR δ were increased. CONCLUSION: FPN disturbed antigen-specific immune responses by affecting GABAergic genes in vivo. We propose that the immunotoxic effects of FPN may enhance antigen-specific immunity by dysregulation of the negative regulation of GABAergic signaling on T cell immunity.

Functionalized pyrazole: Synthesis, Insecticide Evaluation & Molecular Docking Studies of Some New Pyrazole Derivatives against the Cotton leafworm, Spodoptera littoralis.

5-(Cyanomethyl)-3-((5,5-dimethyl-3-oxocyclohex-1-en-1-yl)amino)-1H-pyrazole-4-carbonitrile (3) is used as a key for the synthesis of arylidenes 5a-fvia its reaction with some aldehydes 4a-f. 5-[(5,5-Dimethyl-3-oxocyclohex-1-en-1-yl)amino]-3-(2-imino-2H-chromen-3-yl)-1H-pyrazole-4-carbonitrile (7) was synthesized via the reaction of compound (3) with 2-hydroxybenzaldehyde in EtOH/piperidine. The target compounds were tested against cotton leafworm larvae in their second and fourth instar. The available data demonstrated that the LC50 values for commercial phenylpyrazole were 3.37 mg/L and 4.55 mg/L for the most affected synthesized compound, 5b. The chemical structure of compound 5b has two cyano moieties, a pyrazole ring and a chlorophenyl, which may be increasing it efficiency. Evaluation of the latent effects of the examined synthesized compounds on various biological parameters, including adult longevity, pupal weight, proportion of normal, deformed pupae, adult emergency, fecundity, and egg hatchability, was done in an additional effort to slightly improve insecticidal compounds. Twelve synthesized compounds were subjected to a molecular docking analysis against glutamate-activated chloride channels. Twelve artificial compounds with the PDB ID of 4COF were subjected to a molecular docking study against the gamma-aminobutyric acid receptor (GABA).

Metabolism of an insecticide fipronil by soil fungus Cunninghamella elegans ATCC36112.

In this study, the metabolic pathway of the phenylpyrazole insecticide fipronil in Cunninghamella elegans (C. elegans) was investigated. Approximately 92% of fipronil was removed within 5 days, and seven metabolites were accumulated simultaneously. The structures of the metabolites were completely or tentatively identified by GC-MS and 1H, 13C NMR. To determine the oxidative enzymes involved in metabolism, piperonyl butoxide (PB) and methimazole (MZ) were used, and the kinetic responses of fipronil and its metabolites were determined. PB strongly inhibited fipronil metabolism, while MZ weakly inhibited its metabolism. The results suggest that cytochrome P450 (CYP) and flavin-dependent monooxygenase (FMO) may participate in fipronil metabolism. Integrated metabolic pathways can be inferred from the control and inhibitor experiments. Several novel products from the fungal transformation of fipronil were identified, and similarities between C. elegans transformation and mammalian metabolism of fipronil were compared. Therefore, these results will help to gain insight into the fungal degradation of fipronil and potential applications in fipronil bioremediation. At present, microbial degradation of fipronil is the most promising approach and maintains environmental sustainability. In addition, the ability of C. elegans to mimic mammalian metabolism will assist in illustrating the metabolic fate of fipronil in mammalian hepatocytes and assess its toxicity and potential adverse effects.

Thyroid-Disrupting Effects of Exposure to Fipronil and Its Metabolites from Drinking Water Based on Human Thyroid Follicular Epithelial Nthy-ori 3-1 Cell Lines.

Fipronil is a broad-spectrum insecticide used for plants and poultry. Owing to its widespread use, fipronil and its metabolites (fipronil sulfone, fipronil desulfinyl, and fipronil sulfide), termed FPM, can be frequently detected in drinking water and food. Fipronil can affect the thyroid function of animals, but the effects of FPM on the human thyroid remain unclear. We employed human thyroid follicular epithelial Nthy-ori 3-1 cells to examine combined cytotoxic responses, thyroid-related functional proteins including the sodium-iodide symporter (NIS), thyroid peroxidase (TPO), deiodinases I-III (DIO I-III), and the nuclear factor erythroid-derived factor 2-related factor 2 (NRF2) pathway induced by FPM of 1-1000-fold concentrations detected in school drinking water collected from a heavily contaminated area of the Huai River Basin. Thyroid-disrupting effects of FPM were evaluated by examining biomarkers of oxidative stress and thyroid function and tetraiodothyronine (T4) levels secreted by Nthy-ori 3-1 cells after FPM treatment. FPM activated the expression of NRF2, HO-1 (heme oxygenase 1), TPO, DIO I, and DIO II but inhibited NIS expression and increased the T4 level of thyrocytes, indicating that FPM can disrupt the function of human thyrocytes through oxidative pathways. Given the adverse impact of low FPM concentrations on human thyrocytes, supportive evidence from rodent studies, and the critical importance of thyroid hormones on development, the effects of FPM on the neurodevelopment and growth of children warrant priority attention.

Synthesis of Greenish-Yellow Fluorescent Copper Nanocluster for the Selective and Sensitive Detection of Fipronil Pesticide in Vegetables and Grain Samples.

In this paper, a new synthetic route is introduced for the synthesis of high-luminescent greenish-yellow fluorescent copper nanoclusters (PVP@A. senna-Cu NCs) using Avaram senna (A. senna) and polyvinylpyrrolidone (PVP) as templates. A. senna plant extract mainly contains variety of phytochemicals including glycosides, sugars, saponins, phenols, and terpenoids that show good pharmacological activities such as anti-inflammatory, antioxidant, and antidiabetic. PVP is a stable and biocompatible polymer that is used as a stabilizing agent for the synthesis of PVP@A. senna-Cu NCs. The size, surface functionality, and element composition of the fabricated Cu NCs were confirmed by various analytical techniques. The as-prepared greenish-yellow fluorescent Cu NCs exhibit significant selectivity towards fipronil, thereby favoring to assay fipronil pesticide with good linearity in the range of 3.0-30 µM with a detection limit of 65.19 nM. More importantly, PVP@A. senna-Cu NCs are successfully applied to assay fipronil in vegetable and grain samples.

Novel fast pesticides extraction (FaPEx) strategy coupled with UHPLC-MS/MS for rapid monitoring of emerging pollutant fipronil and its metabolite in food and environmental samples.

In this work, we demonstrated a new, environmental-friendly and effective sample preparation strategy named 'in-syringe-assisted fast pesticides extraction (FaPEx)' technique coupled with LC-MS/MS for the rapid identification and monitoring of emerging pollutant fipronil and its metabolite fipronil sulfone in chicken egg and environmental soil samples. FaPEx strategy comprising of two simple steps. Firstly, the sample was placed in the syringe and extracted using low-volume acetonitrile with NaCl and anhydrous MgSO4 salts. Secondly, the extractant was passed through in-syringe-based solid-phase extraction (SPE) kit containing cleanup sorbents and salt combinations (C18, primary secondary amine, and anhydrous MgSO4) for the cleanup process. Then, the obtained clean extractant was injected into LC-MS/MS for the quantification of target analytes. Various important parameters influencing the FaPEx performances, such as solvent type, salt type, salt amount, sorbent type, and amount, were examined and optimized. The method validation results showed excellent linearity with high correlation coefficients were ≥ 0.99. The estimated LODs were between 0.05-0.07 µg/kg, and LOQs ranged between 0.1-0.25 µg/kg for target analytes in both egg and soil sample matrices, and precision values were ≤7.90%. The developed method was applied to commercial chicken egg samples and environmental soil samples analysis. Spiked recoveries ranged between 88.75-110.91% for egg samples with RSDs ≤7.42% and 82.47-107.46% for soil samples with RSDs <7.37%. These results proved that the developed sample preparation method is a simple, fast, green, low-cost, and efficient method for the analysis of fipronil and its metabolites in food and environmental samples. Thus, this method can be applied as an alternative analytical methodology in routine and standard food and environmental testing laboratories.

Effect of melatonin and luzindole antagonist on fipronil toxicity, detoxification and antioxidant enzyme system in different tissues of Helicoverpa armigera (Lepidoptera: Noctuidae).

Studies have investigating the detoxification and antioxidant enzymes with melatonin under pesticide stress in many vertebrates, whereas no reports produced in invertebrates. In this study possible role of melatonin and luzindole effect on fipronil toxicity and the detoxification, antioxidant enzymes in H. armigera has been reported. Result showed high toxicity of fipronil treatment (LC50 4.24 ppm), followed by increased LC50 value with melatonin pretreatment (6.44 ppm). Whereas decreased toxicity was observed with melatonin and luzindole combination (3.72 ppm). The detoxification enzymes AChE, esterase and P450 were increased in larval head and whole body with exogenous melatonin level compared to control 1-1.5 µmol/mg of protein. The antioxidant levels of CAT, SOD and GST in whole body and head tissue had been increased by melatonin and fipronil combination 1.1-1.4 unit/mg of protein followed by GPx and GR in larval head (1-1.2 µmol/mg of protein). Mean while the luzindole antagonist inhibits CAT, SOD, GST and GR oxidative enzyme level (1-1.5 fold) in most of the tissue compared to melatonin and fipronil treatment (p < 0.01). Hence this study concludes that the melatonin pretreatment can reduce the fipronil toxicity by enhanced detoxification and antioxidant enzyme system in H. armigera.

Large scale systemic control short-circuits pathogen transmission by interrupting the sand rat (Psammomys obesus)-to-sand fly (Phlebotomus papatasi) Leishmania major transmission cycle.

Systemic control uses the vertebrate hosts of zoonotic pathogens as "Trojan horses," killing blood-feeding female vectors and short-circuiting host-to-vector pathogen transmission. Previous studies focused only on the effect of systemic control on vector abundance at small spatial scales. None were conducted at a spatial scale relevant for vector control and none on the effect of systemic control on pathogen transmission rates. We tested the application of systemic control, using Fipronil-impregnated rodent baits, in reducing Leishmania major (Kinetoplastida: Trypanosomatidae; Yakimoff & Schokhor, 1914) infection levels within the vector, Phlebotomus papatasi (Diptera: Psychodidae; Scopoli, 1786) population, at the town-scale. We provided Fipronil-impregnated food-baits to all Psammomys obesus (Mammalia:Muridae; Cretzschmar, 1828), the main L. major reservoir, burrows along the southern perimeter of the town of Yeruham, Israel, and compared sand fly abundance and infection levels with a non-treated control area. We found a significant and substantial treatment effect on L. major infection levels in the female sand fly population. Sand fly abundance was not affected. Our results demonstrate, for the first time, the potential of systemic control in reducing pathogen transmission rates at a large, epidemiologically relevant, spatial scale.

The effect of different decontamination processes on the residues of fipronil and its metabolites in chili fruits (Capsicum annuum L.).

Fipronil is a broad-spectrum phenyl pyrazole insecticide that has a high degree of environmental toxicity. Commonly available chilies in the market are treated with fipronil insecticides. Demand for insecticide-free chili has thus been increasing globally. This needs various sustainable and economical methods to remove insecticides from chilies. The present study examined the effectiveness of several cleaning methods to remove pesticide residues in chili fruits. A supervised field trial was conducted in randomized block design at Rajasthan Agricultural Research Institute, Durgapura, Jaipur, India. Chili samples were subjected to seven different household methods. The samples were extracted using the quick, easy, cheap, effective, rugged, and safe (QuEChERS) method. The residues were analyzed using a gas chromatograph-electron capture detector and confirmed by GC-MS. Of the seven methods, the acetic acid treatment removes the maximum residue effect of fipronil and its metabolites (desulfinyl [MB046513]), sulfide (MB045950), and sulfone (MB046136) on chili fruits. By contrast, the tap water treatment was the least effective. The Food Safety and Standards Authority of India (FSSAI) have set the maximum residue limit value of 0.001 mg kg-1 for fipronil on green chili.

Exploring RNA methylation as a promising biomarker for assessing sublethal effects of fipronil on honeybees (Apis mellifera L.).

Honeybees play a crucial role as pollinators for crops and are regarded as sensitive bioindicators of environmental health. The widespread use of pesticides poses a severe threat to honeybee survival. However, there is limited information available on the specific risks associated with fipronil exposure in honeybees, particularly concerning the impact on RNA methylation throughout their lifespan. This study aimed to evaluate the effects of sublethal concentrations of fipronil on RNA m6A and m5C methylations, along with the associated genes in honeybee larvae and newly emerged adults. LC-MS/MS analysis revealed a notable hypomethylation of m5C in larvae, while hypermethylation of m6A was observed in the adult brain. Significant changes in the expression of genes such as AmWTAP, AmYTHDF, AmALKBH4, AmALKBH6, AmALKBH8, AmNSUN5, AmNOP2, AmTET1, and AmYBX1 were observed in the adult brain, whereas alterations in the expression of AmNSUN2, AmMETTL14, AmALKBH1, AmALKBH4, AmALKBH6 AmALYREF, AmTET1, and AmYBX1 were observed in the larvae. Notably, the expression of AmALKBH1 was not detected in any fipronil-treated larvae, suggesting its potential as an early risk indicator for honeybee larvae in future assessments. This pioneering study provides insights into the effects of fipronil on RNA methylations in honeybees and explores the possibility of employing RNA methylation as a tool for assessing pesticide risks in this important pollinator species. These findings offer new perspectives on honeybee protection and the development of toxicity evaluation systems for pesticides.

Biochemical alterations in cotton leafworm, Spodoptera littoralis (Boisd.) related to emamectin benzoate and fipronil compared to their joint action.

Cotton leafworm, Spodoptera littoralis (Boisduval), is one of the major destructive pests of ornamental, industrial, and vegetable crops. The efficacy of technical emamectin benzoate (EMB) and fipronil (FPR) was assessed against the 4th larval instar using leaf-dip bioassay method. EMB was more efficient than FPR based on 96 h LC50 values of 0.004 and 0.023 µg/ml, respectively. Joint toxic action of the dual exposure in sequence with time interval 24 h and in mix were evaluated at LC10:LC10, LC25:LC25 and LC50:LC50 after 96 h posttreatment, as well. Their impacts on detoxification enzymes, esterases (ESTs); alkaline phosphatase (ALP); and glutathione S-transferase (GST) as well as acetylcholine esterase (AChE) were also determined. The sequential exposure of EMB after FPR (S1) produced antagonism, potentiation, and potentiation effects, respectively while sequential exposure of FPR after EMB (S2) interacted as addition, potentiation, and potentiation respectively. The rest of binary mixtures (Mix) revealed antagonistic effect regardless of concentration. Orthogonal contrast analysis showed that the highest elevations of AChE, α-EST, ß- EST and ALP enzymes were obtained from Mix at LC50:LC50 (181.6%, 288.4, 229.2 and 460.9%, respectively), LC25:LC25 (131.5%, 252.8, 205.60 and 252.0, respectively) and LC10:LC10 (106.6%, 215.6%, 201.8% and 170.0%, respectively). Differently, the greatest elevation of GST activity (157.7%) resulted from S1 at LC50:LC50, while it was significantly lower at LC25:LC25 and LC10:LC10 as well as Mix and S2 at all concentrations than corresponding concentrations of FPR. These findings shed some light on the role of GST in FPR toxicity and clarified the risk of these dual exposures in elevating detoxification enzymes dangerously compared to their individual insecticides. These dual exposures should be carefully handled. Although rotational exposure at low concentrations may enhance performance and mitigate resistance risk, rotational exposure at high concentrations and Mix may indirectly contribute to the evolution of cross-resistance to other insecticides.

Micro-matrix cartridge extraction followed by online micro-solid phase extraction based on polystyrene@hydroxypropyl-ß-cyclodextrin nanofibers for selective determination of fipronil and its metabolites in soil.

Micro-matrix cartridge extraction coupled on-line to micro-solid phase extraction-high performance liquid chromatography-mass spectrometry (µ-MCE-online-µ-SPE-HPLC-MS) is presented. Micro-matrix cartridge extraction (µ-MCE) was applied to highly efficient desorption of adsorbed pesticides from contaminated soil with favorable extraction efficiency (100%). Novel polystyrene@hydroxypropyl-ß-cyclodextrin (PS@HPCD) electrospun nanofibers with 3D network structure were prepared to selectively capture fipronil and its metabolites. High selectivity was obtained with adsorption efficiency ≥ 86.64% via complexation, hydrophobic affinity, and π-π interactions. PS@HPCD nanofibers exhibited remarkable advantages such as excellent enrichment factors (24-55), superior permeability, and long service life (> 65 times). Under the optimum conditions, wide linear range (0.1-1000 ng g-1), low detection limits (0.0032-0.0067 ng g-1), high recoveries (84-124.5%), favorable repeatability (RSD ≤ 10.4%, n = 5), and reproducibility (RSD ≤ 7.2%, n = 3) were acquired for fipronil and three metabolites. The developed method was applied to the pesticide determination in actual soils and the ISO-certified soil with satisfactory recoveries (96.5%). The method developed provides a green, efficient, and miniaturized method for the determination of trace pesticide residues in soil.

Chiral resolution of the insecticide fipronil enantiomers and the simultaneous determination of its major transformation products by high-performance liquid chromatography interfaced with mass spectrometry.

A high-performance liquid chromatography-mass spectrometry (HPLC-MS) method was developed using a chiral column based on amylose tris(3-chloro-5-methylphenylcarbamate) for analysis of fipronil (a popular insecticidal nerve agent) and the related transformation products. The optimized method reached the goal of the simultaneous and complete separation of the multiple fiproles in a single run, including the chiral separation of fipronil enantiomers, fipronil metabolites, and photoproducts. The efficacy of such a method was demonstrated by its application in analyzing a series of fipronil samples exposed to sunlight conditions. In general terms, our study provided experimental approaches and an efficient analytical tool for monitoring the environmental fate of fipronil as well as its multitransformation products upon its applications either in agricultural or any other areas.

Determination of fipronil and its metabolites in environmental water samples by meltblown nonwoven fabric-based solid-phase extraction combined with gas chromatography-electron capture detection.

In this study, a new method for the determination of fipronil and its three metabolites in environmental water samples was developed based on meltblown nonwoven fabric solid-phase extraction combined with gas chromatography-electron capture detection. As the core material of medical masks, meltblown nonwoven fabric is made of polypropylene superfine fibers which are randomly distributed and bonded together with a relatively large specific surface area and good permeability. Polypropylene as a high molecular hydrocarbon-based polymer has the characteristics of good hydrophobicity and lipophilicity, which can be applied for the separation and enrichment of hydrophobic substances in food, environment, and biological samples. The meltblown nonwoven fabric is soft and can fill the solid-phase extraction cartridge tightly. This aspect also makes it suitable to be used as an ideal solid-phase extraction sorbent. A series of parameters influencing the extraction efficiency were investigated, and under the optimized conditions, fipronil and its three metabolites had a good linear relationship in the range of 0.2-100 µg/L with a correlation coefficient R2 of more than 0.999. The recoveries at three spiked concentrations were in the range of 99.2-107.3% with the relative standard deviations less than 9.8% (intra-day) and 8.1% (inter-day). The limit of detection for the four target analytes was in the range of 0.02-0.06 µg/L. Finally, this method was successfully applied in the analysis of fipronil and its three metabolites in various types of environmental water samples.

Silencing of CYP6AS160 in Solenopsis invicta Buren by RNA interference enhances worker susceptibility to fipronil.

Cytochrome P450 monooxygenases play a key role in pest resistance to insecticides by detoxification. Four new P450 genes, CYP6AS160, CYP6AS161, CYP4AB73 and CYP4G232 were identified from Solenopsis invicta. CYP6AS160 was highly expressed in the abdomen and its expression could be induced significantly with exposure to fipronil, whereas CYP4AB73 was not highly expressed in the abdomen and its expression could not be significantly induced following exposure to fipronil. Expression levels of CYP6AS160 and CYP4AB73 in workers were significantly higher than that in queens. RNA interference-mediated gene silencing by feeding on double-stranded RNA (dsRNA) found that the levels of this transcript decreased (by maximum to 64.6%) when they fed on CYP6AS160-specific dsRNA. Workers fed dsCYP6AS160 had significantly higher mortality after 24 h of exposure to fipronil compared to controls. Workers fed dsCYP6AS160 had reduced total P450 activity of microsomal preparations toward model substrates p-nitroanisole. However, the knockdown of a non-overexpressed P450 gene, CYP4AB73 did not lead to an increase of mortality or a decrease of total P450 activity. The knockdown effects of CYP6AS160 on worker susceptibility to fipronil, combined with our other findings, indicate that CYP6AS160 is responsible for detoxification of fipronil. Feeding insects dsRNA may be a general strategy to trigger RNA interference and may find applications in entomological research and in the control of insect pests in the field.

Tissue-specific accumulation, transformation, and depuration of fipronil in adult crucian carp (Carassius auratus).

Accumulation and biotransformation of pesticides in fish tissues are essential to assess their toxicity and associated human exposure risk. The mechanisms on time-dependent and tissue-specific accumulation and transformation of fipronil in adult fish are limited. An experiment consisting of 25-d uptake of fipronil at two levels (10 and 50 µg/L) and 25-d depuration in adult crucian carp (Carassius auratus) was conducted. Fipronil concentration at 25-d exposure was tissue-specific with the order of liver > kidney > blood > muscle. The uptake rate constant of fipronil in the liver (low exposure group: 2.38 ± 0.27 L/kg/d; high exposure group: 1.10 ± 0.11 L/kg/d) was significantly higher than that in other tissues (p < 0.05), and the lowest in muscle (low exposure group: 0.10 ± 0.01 L/kg/d; high exposure group: 0.16 ± 0.11 L/kg/d). The bioconcentration factors of fipronil in different tissues were 1.04-12.7 L/kg wet weight and 177-4268 L/kg lipid. The tissue-blood distribution coefficients of the liver and kidney were lower than 1 based on lipid normalized concentration but higher than 1 based on wet weight concentration, suggesting fipronil was dispersed into other tissues mainly via blood in the lipid-combination pattern. Fipronil sulfone had 1.2-32 times higher concentration and longer depuration time than fipronil, implying fipronil sulfone was more retender in fish bodies. The estimated daily intake of fipronil via fish muscle consumption at 25-d exposure was 8.5-101 and 27-320 ng/kg bw/d for adults and children, respectively. Overall, the human health risk of fipronil and its metabolites with consumption of the polluted fish cannot be negligible.

Sensitivity of the stripe-faced dunnart, Sminthopsis macroura (Gould 1845), to the insecticide, fipronil; implications for pesticide risk assessments in Australia.

A lack of toxicity data quantifying responses of Australian native mammals to agricultural pesticides prompted an investigation into the sensitivity of the stripe-faced dunnart, Sminthopsis macroura (Gould 1845) to the insecticide, fipronil (5-amino-3-cyano-1-(2,6-dichloro-4-trifluoromethylphenyl)-4-trifluoromethylsulfinyl pyrazole, CAS No. 120068-37-3). Using the Up-And-Down method for determining acute oral toxicity in mammals (OECD) median lethal dose estimates of 990 mg kg-1 (95% confidence interval (CI) = 580.7-4770.0 mg kg-1) and 270.4 mg kg-1 (95% CI = 0.0->20,000.0 mg kg-1) were resolved for male and female S. macroura, respectively. The difference between median lethal dose estimates for males and females may have been influenced by the older ages of two female dunnarts. Consequently, further modelling of female responses to fipronil doses used the following assumptions: (a) death at 2000 mg kg-1, (b) survival at 500 mg kg-1 and (c) a differential response (both survival and death) at 990 mg kg-1. This modelling revealed median lethal dose estimates for female S. macroura of 669.1 mg kg-1 (95% CI = 550-990 mg kg-1; assuming death at 990 mg kg-1) and 990 mg kg-1 (95% CI = 544.7-1470 mg kg-1; assuming survival at 990 mg kg-1). These median lethal dose estimates are 3-10-fold higher than available LD50 values of 94 mg kg-1 for a similarly sized eutherian mammal, Mus musculus (L. 1758) and 97 mg kg-1 for Rattus norvegicus (Birkenhout 1769). Implications for pesticide risk assessments in Australia are discussed.

Combined metabolic and target-site resistance mechanisms confer fipronil and deltamethrin resistance in field-collected German cockroaches (Blattodea: Ectobiidae).

Despite insecticide resistance issues, pyrethroids and fipronil have continued to be used extensively to control the German cockroach, Blattella germanica (L.) (Blattodea: Ectobiidae) for more than two decades. We evaluated the physiological insecticide resistance in five German cockroach populations collected from 2018 to 2020 and measured the extent of metabolic detoxification and target-site insensitivity resistance mechanisms. Topically applied doses of the 3 x LD95 of deltamethrin, fipronil, DDT, or dieldrin of a susceptible strain (UCR, Diagnostic Dose) failed to cause >23% mortality, and the 10 x LD95 of deltamethrin or fipronil failed to cause >53% mortality. All field-collected strains possessed a combination of metabolic and target-site insensitivity mechanisms that cause reduced susceptibility. Elevated activities of esterase and glutathione S-transferase were measured, and the synergists piperonyl butoxide or S,S,S-tributyl phosphorotrithioate increased topical mortality up to 100% for deltamethrin and 93% for fipronil 10 x LD95. The target-site mutations L993F of the para-homologous sodium channel and A302S of the GABA-gated chloride channel associated with pyrethroid and fipronil resistance, respectively, were found at ~80-100% frequency in field populations. Pyrethroid and fipronil spray formulations also were ineffective in a choice box assay against field-collected strains suggesting that these treatments would fail to control cockroaches under field conditions.

P-coumaric acid ameliorates fipronil induced liver injury in mice through attenuation of structural changes, oxidative stress and inflammation.

Fipronil is a broad-spectrum phenylpyrazole insecticide and has been used effectively in the agriculture. Due to its widespread use and bioaccumulation in the environment, it possesses significant threat to human and animals. P-coumaric acid is a natural dietary polyphenolic compound that has anti-oxidant and anti-inflammatory property. The present study was aim to investigate the ameliorative effect of p-coumaric acid on fipronil induced liver injury. The mice were divided into five groups (SHAM, FPN, FPN/PCA/50, FPN/PCA/100 and PCA/100) and challenged with fipronil @ 25 mg/kg bw (half of LD50). Haematological, liver function biomarkers (ALT, AST, ALP, GGT), biochemical parameters (MPO, oxidative, nitrosative stress and anti-oxidant enzyme activity), levels of serum and liver inflammatory cytokines (TNF-α, IL-1ß and IL-10), histopathology were monitored. Fipronil administration caused a significant increase in liver enzymes with concomitant significant increase in inflammatory cytokines (TNF-α, IL-1ß, IL-10) and myeloperoxidase activity. A significant increase in oxidative stress (lipid peroxidation, nitric oxide) as well as down regulation of anti-oxidant enzymes like superoxide dismutase (SOD) and catalase (CAT) along with histopathological changes such as microsteatosis, hypertrophy of the hepatocytes and necrosis were observed on fipronil administration. Administration of p-coumaric acid against fipronil caused decreased serum liver enzymes, inflammatory cytokines, myeloperoxidase activity and oxidative stress along with improvement in anti-oxidant enzyme levels and structural changes induced by fipronil. Thus p-coumaric acid ameliorates the FPN induced liver injury in mice through attenuation of structural changes, oxidative stress, and inflammation.