Monitoring cell viability in N-nitrosodiethylamine induced acute hepatitis and detection of hydrazine in solution and gas phase with Dual-function fluorescent probes.

The highly toxic N-nitrosodiethylamine (NDEA) and hydrazine (N2H4) caused severe environmental contamination and serious health risks. Herein, we designed the two-photon ratiometric fluorescent probe (Nap-2), emission maximum shifted from 466 nm to 571 nm, to monitor cell viability of NDEA induced acute hepatitis via esterase activity detection. Furthermore, the probe Nap-2 evaluate the hydrazine (N2H4) content in the solution and gas phase. It is worth mentioning that we used NDEA induced acute hepatitis in the mice and evaluated the negative correlation of esterase activity in the tissue cells and serum with Nap-2. The probe Nap-2 exhibited that acute hepatitis induced by NDEA decreased cell viability. Furthermore, we made convenient test papers using Nap-2 to detect N2H4 in solution and gas phase. After adding N2H4, the fluorescence color changed from blue to yellow and was visible to the naked eye. This work provides a convenient tool and method for evaluating the toxicity of NDEA induced acute hepatitis and detecting N2H4 in the environment.

Exploring the molecular and functional cellular response to hydrazine via transcriptomics and DNA repair mutant cell lines.

Hydrazine is a rodent carcinogen and is classified as a probable human carcinogen by IARC. Though hydrazine is positive in both in vitro and in vivo DNA strand break (comet) assays, hydrazine was reported to be negative in an in vitro mutation Muta Mouse lung epithelial cell (FE1) test, as well as in a regulatory-compliant, in vivo Big Blue mouse mutation test. In this article, mechanistic studies explored the cellular response to hydrazine. When tested in a regulatory-compliant mouse lymphoma assay, hydrazine yielded unusual, weakly positive results. This prompted an investigation into the transcriptional response to hydrazine in FE1 cells via RNA sequencing. Amongst the changes identified was a dose-dependent increase in G2/M DNA damage checkpoint activation associated genes. Flow cytometric experiments in FE1 cells revealed that hydrazine exposure led to S-phase cell cycle arrest. Clonogenic assays in a variety of cell lines harboring key DNA repair protein deficiencies indicated that hydrazine could sensitize cells lacking homology dependent repair proteins (Brca2 and Fancg). Lastly, hprt assays with hydrazine were conducted to determine whether a lack of DNA repair could lead to mutagenicity. However, no robust, dose-dependent induction of mutations was noted. The transcriptional and cell cycle response to hydrazine, coupled with functional investigations of DNA repair-deficient cell lines support the inconsistencies noted in the genetic toxicology regulatory battery. In summary, while hydrazine may be genotoxic, transcriptional and functional processes involved in cell cycle regulation and DNA repair appear to play a nuanced role in mediating the mutagenic potential.

Copper induces cell death by targeting lipoylated TCA cycle proteins.

Copper is an essential cofactor for all organisms, and yet it becomes toxic if concentrations exceed a threshold maintained by evolutionarily conserved homeostatic mechanisms. How excess copper induces cell death, however, is unknown. Here, we show in human cells that copper-dependent, regulated cell death is distinct from known death mechanisms and is dependent on mitochondrial respiration. We show that copper-dependent death occurs by means of direct binding of copper to lipoylated components of the tricarboxylic acid (TCA) cycle. This results in lipoylated protein aggregation and subsequent iron-sulfur cluster protein loss, which leads to proteotoxic stress and ultimately cell death. These findings may explain the need for ancient copper homeostatic mechanisms.

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

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

A novel colorimetric and ratiometric fluorescence probe based on 'C-CN' for detection of hydrazine and its imaging in living cells and mouse.

Hydrazine plays an important role in chemistry, pharmaceuticals, agriculture and aerospace. However, it is not to be underestimated and has been identified as harmful to the human body. Therefore, it is significant and urgent to develop the detection of hydrazine in vivo and in vitro. Here, the probe TAN was synthesized by using benzothiazole derivatives as the fluorophore and 2,3-diaminomaleonitrile as the identified group to detect hydrazine. The presence of hydrazine resulted in a colorimetric and ratiometric fluorescence response of the probe based on the formation of hydrazone. The detection limit of TAN was 0.31 µM for hydrazine. In addition, the probe TAN was successfully used to visualize hydrazine in living HepG-2 cells and mouse with low cytotoxicity and excellent biocompatibility.

Susceptibility of the Non-Targeted Crustacean Eurytemora affinis to the Endocrine Disruptor Tebufenozide: A Transcriptomic Approach.

Copepods are zooplanktonic crustaceans ubiquitously widespread in aquatic systems. Although they are not the target, copepods are exposed to a wide variety of pollutants such as insect growth regulators (IGRs). The aim of this study was to investigate the molecular response of a non-targeted organism, the copepod Eurytemora affinis, to an IGR. Adult males and females were exposed to two sub-lethal concentrations of tebufenozide (TEB). Our results indicate a sex-specific response with a higher sensitivity in males, potentially due to a differential activation of stress response pathways. In both sexes, exposure to TEB triggered similar pathways to those found in targeted species by modulating the transcription of early and late ecdysone responsive genes. Among them were genes involved in cuticle metabolism, muscle contraction, neurotransmission, and gametogenesis, whose mis-regulation could lead to moult, locomotor, and reproductive impairments. Furthermore, genes involved in epigenetic processes were found in both sexes, which highlights the potential impact of exposure to TEB on future generations. This work allows identification of (i) potential biomarkers of ecdysone agonists and (ii) further assessment of putative physiological responses to characterize the effects of TEB at higher biological levels. The present study reinforces the suitability of using E. affinis as an ecotoxicological model.

Role of Hydrazine-Related Chemicals in Cancer and Neurodegenerative Disease.

Hydrazine-related chemicals (HRCs) with carcinogenic and neurotoxic potential are found in certain mushrooms and plants used for food and in products employed in various industries, including aerospace. Their propensity to induce DNA damage (mostly O6-, N7- and 8-oxo-guanine lesions) resulting in multiple downstream effects is linked with both cancer and neurological disease. For cycling cells, unrepaired DNA damage leads to mutation and uncontrolled mitosis. By contrast, postmitotic neurons attempt to re-enter the cell cycle but undergo apoptosis or nonapoptotic cell death. Biomarkers of exposure to HRCs can be used to explore whether these substances are risk factors for sporadic amyotrophic laterals sclerosis and other noninherited neurodegenerative diseases, which is the focus of this paper.

Eltrombopag second-line therapy in adult patients with primary immune thrombocytopenia in an attempt to achieve sustained remission off-treatment: results of a phase II, multicentre, prospective study.

Up to 30% immune thrombocytopenia (ITP) patients achieve a sustained remission off-treatment (SROT) after discontinuation of thrombopoietin receptor agonists (TPO-RAs). Factors predictive of response are lacking. Patients aged ≥18 years with newly diagnosed or persistent ITP were treated with eltrombopag for 24 weeks. Primary end-point was SROT: the proportion of responders that were able to taper and discontinue eltrombopag maintaining the response during a period of observation (PO) of six months. Secondary end-points included the association between some immunological parameters (TPO serum levels, cytokines and lymphocyte subsets) and response. Fifty-one patients were evaluable. Primary end-point was achieved in 13/51 (25%) treated patients and 13/34 (38%) patients who started the tapering. Baseline TPO levels were not associated with response at week 24 nor with SROT. Higher baseline levels of IL-10, IL-4, TNF-α and osteopontin were negative factors predictive of response (P = 0·001, 0·008, 0·02 and 0·03 respectively). This study confirms that SROT is feasible for a proportion of ITP patients treated with eltrombopag. Some biological parameters were predictive of response.

Efficacy of biorational insecticides against Bemisia tabaci (Genn.) and their selectivity for its parasitoid Encarsia formosa Gahan on Bt cotton.

The toxicity of seven biorational insecticides [five insect growth regulators (Buprofezin, Fenoxycarb, Pyriproxyfen, Methoxyfenozide, and Tebufenozide) and two oil-extracts of neem and bitter gourd seeds] against Bemisia tabaci and their selectivity for its parasitoid, Encarsia formosa were evaluated in laboratory and field conditions for 2 years (2018-2019) in Pakistan. Toxicity results demonstrate that Pyriproxyfen, Buprofezin, and Fenoxycarb proved to be effective (80-91% mortality and 66.3-84.2% population-reduction) against B. tabaci followed by Methoxyfenozide, Tebufenozide (50-75% mortality and 47.8-52.4% population-reduction), and then oil-extracts of neem and bitter gourd (25-50% mortality and 36.5-39.8% population-reduction) in the laboratory [72 h post-application exposure interval (PAEI)] and field trails (168 h PAEI), respectively. All tested biorationals, except Methoxyfenozide [(slightly-harmful/Class-II), i.e., causing mortality of parasitoids between a range of 25-50%] and Tebufenozide [(moderately-harmful/Class-III), i.e., causing mortality of parasitoids between the ranges of 51-75%], proved harmless/Class-I biorationals at PAEI of 7-days in the field (parasitism-reduction < 25%) and 3-days in the lab (effect < 30%). In laboratory bioassays, exposure of parasitized-pseudopupae and adult-parasitoids to neem and bitter gourd oils demonstrated that these compounds proved harmless/Class-I biorationals (< 30% mortality). Alternatively, Pyriproxyfen, Buprofezin, Fenoxycarb, Methoxyfenozide, and Tebufenozide were slightly-harmful biorationals (30-79% mortality) against the respective stages of E. formosa. We conclude that most of the tested biorationals proved harmless or slightly harmful to E. formosa, except tebufenozide after PAEI of 7-days (168 h) in the field and, therefore, may be used strategically in Integrated Pest Management (IPM) of B. tabaci.

In vitro and in vivo mammalian mutation assays support a nonmutagenic mechanism of carcinogenicity for hydrazine.

Hydrazine has been described as a mutagenic, probable human carcinogen. It is mutagenic in in vitro systems such as bacterial reverse mutation (Ames) tests and some yeast systems, as well as in in vivo systems with drosophila. It was shown to cause chromosome damage both in vitro and in vivo but was negative in some well-validated mammalian mutation systems such as CHO HPRT assays. Importantly, there is only one in vivo gene mutation test reported, which was negative. Our objective was to determine if hydrazine is mutagenic in mammalian test systems. Thus, we conducted an in vitro gene mutation test in Muta™Mouse lung epithelial cells (FE1 cell assay) and a regulatory-compliant in vivo Big Blue® mouse test. Consistent with previous reports, an additional six-well Ames assay showed that hydrazine was mutagenic to bacteria. The FE1 cell assay was negative in conditions with and without metabolic activation when tested to cytotoxicity limits. In the Big Blue® mouse study, female mice received dosages of hydrazine up to 10.9 mg/kg via drinking water for 28 days. This dose is comparable to a dose used in a carcinogenicity study where female mice had significant increases in hepatocellular adenoma at 11.5 mg/kg. There were no increases in mutant frequency in liver and lung, two tissues sensitive to the carcinogenic effects of hydrazine in mice. Our research shows that hydrazine is not mutagenic in mammalian cells either in vitro or in vivo, indicating mutagenicity may not play a role in the carcinogenicity of hydrazine.

The Toxicity, Pathophysiology, and Treatment of Acute Hydrazine Propellant Exposure: A Systematic Review.

INTRODUCTION: Hydrazines are highly toxic inorganic liquids that are used as propellants in military and aviation industries, such as the U.S. Air Force F-16 Emergency Power Unit and SpaceX SuperDraco Rockets. The most commonly used derivatives include hydrazine, monomethylhydrazine, and 1,1-dimethylhydrazine (unsymmetrical dimethylhydrazine). Industrial workers in close contact with hydrazines during routine maintenance tasks can be exposed to levels well above the National Institute for Occupational Safety and Health relative exposure limits. MATERIALS AND METHODS: A systematic review was performed using PubMed, Web of Science, Google Scholar, National Aeronautics and Space Administration Technical Server, and Defense Technical Information Center, and data related to hydrazine exposures were searched from inception to April 2020. Publications or reports addressing hydrazine toxicity, pathophysiology, and treatment of hydrazine fuel exposure were selected. RESULTS: Acute toxic exposures to hydrazine and its derivatives are rare. There are few case reports of acute toxic exposure in humans, and data are largely based on animal studies. The initial search identified 741 articles, manuscripts, and government reports. After screening for eligibility, 51 were included in this review. Eight articles reported acute exposures to hydrazine propellant in humans, and an additional 14 articles reported relevant animal data. CONCLUSIONS: Exposure to small amounts of hydrazine and its derivatives can cause significant soft tissue injury, pulmonary injury, seizures, coma, and death. Neurologic presentations can vary based on exposure compound and dose. Decontamination is critical as treatment is mainly supportive. High-dose intravenous pyridoxine has been suggested as treatment for hydrazine-related neurologic toxicity, but this recommendation is based on limited human data. Despite recent research efforts to generate less toxic alternatives to hydrazine fuel, it will likely continue to have a role in military and aviation industries. Aerospace and military physicians should be aware of the toxicity associated with hydrazine exposure and be prepared to treat hydrazine toxicity in at-risk populations.

Warning on False or True Morels and Button Mushrooms with Potential Toxicity Linked to Hydrazinic Toxins: An Update.

Recently, consumption of the gyromitrin-containing neurotoxic mushroom Gyromitra sp. (false morel), as gourmet food was hypothesized to play a role in sporadic amyotrophic lateral sclerosis genesis. The present review analyses recent data on edibility and toxicity of false and true morels and Agaricus spp. Controversy about the toxic status of Gyromitra esculenta was due to variable toxin susceptibility within consumers. We suggest that Verpa bohemica, another false morel, is also inedible. We found a temporary neurological syndrome (NS) with cerebellar signs associated with high consumption of fresh or dried true morels Morchella sp. After ingestion of crude or poorly cooked fresh or dried morels, a gastrointestinal "haemolytic" syndrome was also observed. Agaritine, a water soluble hydrazinic toxin closely related to gyromitrin is present along with metabolites including diazonium ions and free radicals, in Agaricus spp. and A. bisporus, the button mushroom, and in mice after ingestion. It is a potential weak carcinogen in mice, but although no data are available for humans, a lifetime low cumulative extra cancer risk in humans can be estimated to be about 10-5. To conclude, a safety measure is to avoid consuming any true morels or button mushrooms when crude or poorly cooked, fresh or dried.

A dual-response ratiometric fluorescent probe for hypochlorite and hydrazine detection and its imaging in living cells.

In this work, a dual-response ratiometric fluorescent probe (E)-3-(5-(2-nitrovinyl)thiophen-2-yl)-9-phenyl-9H-carbazole (NTPC) for high selectivity and sensitivity detection of ClO- and N2H4 was successfully developed. This probe NTPC showed ratiometric fluorescent response to ClO- and N2H4, which induces obvious naked-eye color changes, respectively. In addition, the NTPC for ClO- and N2H4 detection displayed low detection limits of 71.4 nM and 0.6 µM, respectively. And the sensing mechanism of NTPC with ClO- and N2H4 was well confirmed by 1H NMR and HR-MS spectra. Moreover, this novel probe was applied to monitoring and differentiating ClO- and N2H4 in living cells, and exhibits good biocompatibility and low cytotoxicity.

In silico and in vivo anti-malarial investigation on 1-(heteroaryl)-2-((5-nitroheteroaryl)methylene) hydrazine derivatives.

BACKGROUND: Resistance of Plasmodium falciparum against common anti-malarial drugs emphasizes the need of alternative and more effective drugs. Synthetic derivatives of 1-(heteroaryl)-2-((5-nitroheteroaryl)methylene) hydrazine have showed in vitro anti-plasmodial activities. The present study aimed to evaluate the molecular binding and anti-plasmodial activity of synthetic compounds in vivo. METHODS: The molecular docking was used to study the binding of compounds to haem and Plasmodium falciparum lactate dehydrogenase (PfLDH). Acute toxicity of the synthetic compounds was evaluated based on the modified up & down method. The anti-plasmodial activity of the compounds was conducted by the two standard tests of Peters' and of Rane, using chloroquine-sensitive Plasmodium berghei in mice. Also, the toxicity to the internal organs of mice was evaluated on the seventh day after the treatment in addition to the histopathology of their liver. Compound 3 that showed high activity in the lowest dose was selected for further pharmacodynamic studies. RESULTS: According to the docking studies, the active site of PfLDH had at least four common residues, including Ala98, Ile54, Gly29, and Tyr97 to bind the compounds with the affinity, ranging from - 8.0 to - 8.4 kcal/mol. The binding mode of ligands to haem revealed an effective binding affinity, ranging from - 5.1 to - 5.5 kcal/mol. Compound 2 showed the highest  % suppression of parasitaemia (99.09%) at the dose of 125 mg/kg/day in Peters' test. Compound 3, with 79.42% suppression, was the best in Rane's test at the lowest dose (31 mg/kg/day). Compound 3 was confirmed by the pharmacodynamic study to have faster initial parasite elimination in the lowest concentration. The histopathology of the livers of mice did not reveal any focal necrosis of hepatocytes in the studied compounds. CONCLUSIONS: The docking studies verified Pf LDH inhibition and the inhibitory effect on the haemozoin formation for the studied compounds. Accordingly, some compounds may provide new avenues for the development of anti-malarial drugs without liver toxicity, although further studies are required to optimize their anti-plasmodial activity.

Comparative study of antibacterial activity between Schiff base nicotinic hydrazide derivative and its silver architected nanoparticles with atomic force microscopic study of bacterial cell wall.

The threat of multi-drug resistant bacterial pathogens evokes researchers to synthesized safe and effective chemotherapeutic agents for nano-drug delivery system. In current study, Schiff base of nicotinic hydrazide(NHD) and its silver nanoparticles(NHD-AgNPs) were synthesized and characterized. These compounds were investigated for cytotoxicity, antibacterial and AFM activity. The NHD showed LD50 at >1000µg/mL while NHD-AgNPs didn't exhibit toxicity at 1000µg/mL against 3T3 cell line. The NHD showed zone of inhibition against two strains of salmonella enteric (ATCC 14028 and 700408) 45.29±1.66 and 48.01±1.43mm respectively at 160µg/mL (p<0.01) while NHD-AgNPs exhibited 55.87±2.08 and 52.88±1.42 mm respectively at 130µg/mL (p<0.001) in disc diffusion method. NHD showed more than 70% growth inhibition for both strains at 85 and 125µg/ml (p<0.01) respectively, while NHD-AgNPs inhibit 80% and 75% respectively at 75 and 125 µg/ml (p<0.01, p<0.001) against Alamar blue antibacterial assay. For morphological changes in bacterial cell wall NHD and NHD-AgNPs treated bacterial cells were observed under atomic force microscope(AFM) and treated bacterial cells were severely damaged with leaked cytoplasmic contents as compare to untreated bacterial cell. These results validate that NHD-AgNPs were highly active as compared to NHD against both strains at their MIC concentrations. In future, comparative wound healing potential will be emphasized.

Targeted amino acid and related amines analysis based on iTRAQ®-LC-MS/MS for discovering potential hepatotoxicity biomarkers.

Drug induced liver injury (DILI) is a diverse set of liver injury occurring after exposure to any manufactured or naturally occurring chemical compounds. DILI has already become the leading cause of acute liver failure in developed countries. Early diagnosis is of great significance for the prevention and treatment of DILI, which can effectively avoid its progress to acute liver failure. In this paper, a novel targeted metabolomics method based on isobaric tagging reagent iTRAQ®-LC-MS/MS was developed for the exploring of 42 common amino acids and related amines alterations in rats treated with hydrazine, aiming to discover the potential biomarkers for early diagnosis of DILI. Forty-two amino acids and related amines were covered in this new method. Through derivatization by iTRAQ reagent, all derivatized amino acids and related amines can be separated and quantified in 16 min with excellent peak shape and good separation efficiency. Fragments related to reporter group (m/z 113 or 121) of iTRAQ reagents could be generated from all derivatized amino acids and related amines under general multiple reaction monitoring (MRM) parameters. Isotope dilution method was established for the quantification of amino acid, which significantly reduced the interference of matrix effect on quantitative accuracy. Using this iTRAQ®-LC-MS/MS method, the changes of amino acid metabolism were comprehensively investigated in rat serum and urine samples after DILI modeling by hydrazine. More significant changes of amino acids were observed in serum and urine samples with fold changes ranging from 0.5 to 193.7. Six significantly increased amino acids in serum, including L-citrulline, L-α-amino-adipic acid, L-tyrosine, L-glutamic acid, glycine and L-lysine, and ten amino acids and related amines in urine including L-citrulline, L-α-amino-adipic acid, L-tyrosine, taurine, ß-alanine, ethanolamine, argininosuccinic acid, D,L-ß-amino-isobutyric acid, γ-amino-n-butyric acid and L-glutamine, 3 of which were detected in both serum and urine. Except for L-lysine all these significantly increased amino acids and related amines possessed 92.5% to 100% specificity and sensitivity calculated at best cut-off points of their ROC curves in distinguishing control and DILI model group.

The effect of hydrazide linkers on hyaluronan hydrazone hydrogels.

The effect of hydrazide linkers on the formation and mechanical properties of hyaluronan hydrogels was intensively evaluated. The reaction kinetics of hydrazone formation was monitored by NMR spectroscopy under physiological conditions where polyaldehyde hyaluronan (unsaturated: ΔHA-CHO, saturated: HA-CHO) was reacted with various hydrazides to form hydrogels. Linear (adipic, oxalic dihydrazide) and branched (N,N´,N´´-tris(hexanoylhydrazide-6-yl)phosphoric triamide and 4-arm-PEG hydrazide) hydrazides were compared as crosslinking agents. The mechanical properties of hydrogels were also modified by attaching a hydrophobic chain to HA-CHO; however, it was found that this modification did not lead to an increase in hydrogel stiffness. Cytotoxicity tests showed that all tested hydrazide crosslinkers reduced the viability of cells only slightly, and that the final hyaluronan hydrogels were non-toxic materials.

Exposure to sublethal concentrations of methoxyfenozide disrupts honey bee colony activity and thermoregulation.

Methoxyfenozide is an insect growth regulator (IGR) commonly used in agriculture to simultaneously control pests and preserve beneficial insect populations; however, its impact on honey bees in not fully understood. We conducted field and laboratory experiments to investigate bee health in response to field-relevant concentrations of this pesticide. Significant effects were observed in honey bee colony flight activity and thermoregulation after being exposed over 9 weeks to supplemental protein patty containing methoxyfenozide. Compared to bee colonies in the control group, colonies fed pollen patty with 200 ppb methoxyfenozide (as measured by residue analysis) had: 1) a significantly reduced rate of weight loss due to forager departure in the morning; and 2) higher temperature variability during the winter. Colonies in the 100 ppb (as measured by residue analysis) treatment group had values between the 200 ppb group and control for both response variables. The dusk break point, which is the time associated with the end of forager return, differed among all treatment groups but may have been confounded with direction the hives were facing. Bee colony metrics of adult bee mass and brood surface area, and measurements of bee head weight, newly-emerged bee weight, and hypopharyngeal gland size were not significantly affected by methoxyfenozide exposure, suggesting that there may be significant effects on honey bee colony behavior and health in the field that are difficult to detect using standard methods for assessing bee colonies and individuals. The second experiment was continued into the following spring, using the same treatment groups as in the fall. Fewer differences were observed among groups in the spring than the fall, possibly because of abundant spring forage and consequent reduced treatment patty consumption. Residue analyses showed that: 1) observed methoxyfenozide concentrations in treatment patty were about 18-60% lower than the calculated concentrations; 2) no residues were observed in wax in any treatment; and 3) methoxyfenozide was detected in bee bread only in the 200 ppb treatment group, at about 1-2.5% of the observed patty concentration.

Transcriptional response of ATP-binding cassette (ABC) transporters to insecticides in the cotton bollworm, Helicoverpa armigera.

When any living organism is frequently exposed to any drugs or toxic substances, they evolve different detoxification mechanism to confront with toxicants during absorption and metabolism. Likewise, the insects have evolved detoxification mechanisms as they are frequently exposed to different toxic secondary plant metabolites and commercial insecticides. ABC transporter superfamily is one of the largest and ubiquitous group of proteins which play an important role in phase III of the detoxification process. However, knowledge about this gene family remains largely unknown. To help fill this gap, we have identified a total of 54 ABC transporters in the Helicoverpa armigera genome which are classified into eight subfamilies (A-H) by phylogenetic analysis. The temporal and spatial expression profiles of these 54 ABC transporters throughout H. armigera development stages and seven tissues and their responses to five different insecticides, were investigated using RNA-seq analysis. Furthermore, the mRNA expression of eight selected genes in different tissues and six genes responses to insecticides were confirmed by the quantitative real-time PCR (RT-qPCR). Moreover, H. armigera become more sensitive to abamectin and indoxacarb when P-gp was inhibited. These results provide a foundation for further studies of ABCs in H. armigera.