Ecotoxicological risk assessment of 14 pesticides and corresponding metabolites to groundwater and soil organisms using China-PEARL model and RQ approach.

Global use of pesticides brings uncertain risks to human and nontarget species via environmental matrix. Currently, various models for exposure risk assessment are developed and widely used to forecast the impact of pesticides on environmental organisms. In this study, five commonly used insecticides, seven herbicides and three fungicides were chosen to analyze the subsequent risks in groundwater in simulated scenarios using China-PEARL (Pesticide Emission Assessment at Regional and Local Scales) model. In addition, their exposure risks to soil organisms were characterized based on risk quotient (RQ) approach. The results indicated that 23.3% of the total 528 predicted environmental concentrations (PECs) of pesticides and respective metabolites in groundwater from six Chinese simulated locations with ten crops were above 10 µg L-1. Furthermore, acceptable human risks of pesticides in groundwater were observed for all simulation scenarios (RQ < 1). Based on the derived PECs in soil short-term and long-term exposure simulation scenarios, all compounds were evaluated to be with acceptable risks to soil organisms, except that imidacloprid was estimated to be with unacceptable chronic risk (RQ = 27.5) to earthworms. Overall, the present findings provide an opportunity for a more-comprehensive understanding of exposure toxicity risks of pesticides leaching into groundwater and soil.

Genotoxicity response of Vicia faba seedlings to cadmium in soils as characterized by direct soil exposure and micronucleus test.

To overcome the drawbacks of the Vicia faba root tip micronucleus test in soil using the solution extract method, we conducted a potting experiment by direct soil exposure. Cadmium was spiked into 3 typical soils (brown soil, red soil, and black soil) to simulate environmental concentrations (0.625, 1.25, 2.5, 5, and 10 mg kg-1). Multiple Vicia faba tissues (primary root tips, secondary root tips, and leaf tips) were sampled, and mitotic index (MI), chromosome aberration frequency (CA), and micronucleus frequency (MN) were used as endpoints after a seedling period of 5 days. The results showed a response between Cd concentrations and multiple sampling tissues of Vicia faba, and the secondary root tips responded to Cd stress the most, followed by primary root tips and leaf tips. Soil physicochemical properties (e.g., pH, total phosphorus, total organic carbon, etc.) influenced the genotoxicity of Cd, and pH was the dominant factor, which resulted in the genetic toxicity response of Cd in soils in the order: red soil > brown soil > black soil. The lowest observable effect concentration (LOEC) of Cd was 1.25 mg kg-1 for both brown soil and red soil and 2.5 mg kg-1 for black soil. In view of this, we suggested that soil properties should be considered in evaluating genotoxicity risk of Cd in soil, especially with soil pH range, and the secondary root tips should be taken as suitable test tissues in the MN test due to its more sensible response feature to Cd stress in soil.

Toxicity of six insecticides to predatory mite Amblyseius cucumeris (Oudemans) (Acari: Phytoseiidae) in- and off-field.

Amblyseius cucumeris (Oudemans) is a beneficial non-target arthropod (NTA) and a key predator of tetranychid mites in integrated pest management (IPM) programs across China. Evaluating the toxic effects of insecticides on such predatory mites is essential for the success and development of IPM. We tested six insecticides to determine the risk of neonicotinoid insecticide toxicity to predatory mites, using the 'open glass plate method' and adult female A. cucumeris in a "worst case laboratory exposure" scenario. A 48-h toxicity test was performed using the hazard quotient (HQ) approach to evaluate the risk of each insecticide. The LR50 values (application rate that caused 50% mortality) of acetamiprid, thiamethoxam, imidacloprid, and dinotefuran were 76.4, 104.5, 84.9, and 224.6 g active ingredient (a.i.) ha-1, respectively, with in-field HQ values of 0.40, 1.28, 0.49, and 0.82, respectively. The HQ values were lower than the trigger value of 2, and were consistent with off-field values. The risks of the four neonicotinoid insecticides to adult female A. cucumeris were acceptable in two exposure scenarios in field and off field. The 48-h LR50 values for bifenthrin and malathion were 0.008 and 0.062 g. a.i. ha-1, respectively, which were much lower than the recommended field application rates. The HQ values were much higher than the trigger values for both in- and off-field, indicating that the risks of these two insecticides were unacceptable. Bifenthrin and malathion posed an extremely high risk to the test species, and their use should be restricted to reduce risks to the field with augmentative releases of A. cucumeris.

Ecotoxicity and Risk to Zooplankton of a Novel Insecticide Used in Rice Paddies in a Simulated Ecosystem.

Diamide insecticides are widely used in rice paddies and pose a potential threat to aquatic organisms. However, the risk research related to their application in major rice-producing areas is very limited, especially mesocosm research to simulate the impact on aquatic ecosystems of long-term exposure, as well as exposure analysis based on local models and local scenarios. To assess potential risks from a novel diamide insecticide (tetrachlorantraniliprole) to aquatic nontarget organisms in the field over long-term exposure, an outdoor mesocosm study was performed, and the environmental concentrations were predicted by the multimedia paddy-pond model (TOPRICE). The mesocosm experiment showed that tetrachlorantraniliprole mainly stayed in the aqueous phase after entering the water body. Although the chemical dissipated quickly in the aqueous phase (half-life of 0.79-1.5 days), it showed toxic effects on zooplankton communities. Cladocerans, represented by Simocephalus vetulus, were most sensitive to tetrachlorantraniliprole stress. Significant short-term toxicity to cladocerans occurred in all treatment groups, but all recovered within 8 weeks except for the highest concentration group (30.0 µg /L). Based on the ecological recovery results, 7.74 µg tetrachlorantraniliprole/L (nominal concentration, 10.0 µg /L) is suggested to be the no-observed-ecological-adverse-effect concentration (NOEAEC) for the zooplankton community. When this NOEAEC was compared with predicted environmental concentrations (PECs; the PECs in natural ponds simulated by the TOPRICE model for 148 application scheme combinations in major rice-producing areas), a relatively high risk of applying tetrachlorantraniliprole during the rice tillering stage was found. The present study makes a positive contribution to the hypothesis that the current Tier 1 approaches for global acute risk assessment have a sufficient protective effect for assessing the risk of tetrachlorantraniliprole to aquatic organisms. Also, the present results should help us to gain a fuller understanding of the ecological risk of diamide insecticides in aquatic ecosystems and their rational application schemes. Environ Toxicol Chem 2024;43:429-439. © 2023 SETAC.

A Specific High Toxicity of Xinjunan (Dioctyldiethylenetriamine) to Xanthomonas by Affecting the Iron Metabolism.

Xanthomonas spp. encompass a wide range of phytopathogens that brings great economic losses to various crops. Rational use of pesticides is one of the effective means to control the diseases. Xinjunan (Dioctyldiethylenetriamine) is structurally unrelated to traditional bactericides, and is used to control fungal, bacterial, and viral diseases with their unknown mode of actions. Here, we found that Xinjunan had a specific high toxicity toward Xanthomonas spp., especially to the Xanthomonas oryzae pv. oryzae (Xoo), the causal agent of rice bacterial leaf blight. Transmission electron microscope (TEM) confirmed its bactericidal effect by morphological changes, including cytoplasmic vacuolation and cell wall degradation. DNA synthesis was significantly inhibited, and the inhibitory effect enhanced with the increase of the chemical concentration. However, the synthesis of protein and EPS was not affected. RNA-seq revealed differentially expressed genes (DEGs) particularly enriched in iron uptake, which was subsequently confirmed by siderophore detection, intracellular Fe content and iron-uptake related genes transcriptional level. The laser confocal scanning microscopy and growth curve monitoring of the cell viability in response to different Fe condition proved that the Xinjunan activity relied on the addition of iron. Taken together, we speculated that Xinjunan exerted bactericidal effect by affecting cellular iron metabolism as a novel mode of action. IMPORTANCE Sustainable chemical control for rice bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae need to be developed due to limited bactericides with high efficiency, low cost, and low toxicity in China. This present study verified a broad-spectrum fungicide named Xinjunan possessing a specific high toxicity to Xanthomonas pathogens, which were further confirmed by affecting the cellular iron metabolism of Xoo as a novel mode of action. These findings will contribute to the application of the compound in the field control of Xanthomonas spp.-caused diseases, and be directive for future development of novel specific drugs for the control of severe bacterial diseases based on this novel mode of action.

Abiotic transformation of kresoxim-methyl in aquatic environments: Structure elucidation of transformation products by LC-HRMS and toxicity assessment.

In this study, abiotic transformation of an important strobilurin fungicide, kresoxim-methyl, was investigated under controlled laboratory conditions for the first time by studying its kinetics of hydrolysis and photolysis, degradation pathways and toxicity of possibly formed transformation products (TPs). The results indicated that kresoxim-methyl showed a fast degradation in pH9 solutions with DT50 of 0.5 d but relatively stable under neutral or acidic environments in the dark. It was prone to photochemical reactions under simulated sunlight, and the photolysis behavior was easily affected by different natural substances such as humic acid (HA), Fe3+and NO3-which are ubiquitous in natural water, showing the complexity of degradation mechanisms and pathways of this chemical compound. The potential multiple photo-transformation pathways via photoisomerization, hydrolyzation of methyl ester, hydroxylation, cleavage of oxime ether and cleavage of benzyl ether were observed. 18 TPs generated from these transformations were structurally elucidated based on an integrated workflow combining suspect and nontarget screening by high resolution mass spectrum (HRMS), and two of them were confirmed with reference standards. Most of TPs, as far as we know, have never been described before. The in-silico toxicity assessment showed that some of TPs were still toxic or very toxic to aquatic organisms, although they exhibit lower aquatic toxicity compared to the parent compound. Therefore, the potential hazards of the TPs of kresoxim-methyl merits further evaluation.

Accumulation of epoxiconazole from soil via oleic acid-embedded cellulose acetate membranes and bioavailability evaluation in earthworms (Eisenia fetida).

A passive sampler in the soil environment is a relatively novel technique and has had quite limited applications, especially for pesticides. Oleic acid-embedded cellulose acetate membranes (OECAMs) were developed to evaluate the bioavailability of epoxiconazole (EPO) to earthworms (Eisenia fetida). The uptake of EPO by OECAMs (R2 = 0.975) and earthworms (R2 = 0.938) was compared and found to follow a two-compartment kinetic model. EPO sampling by OECAMs reached equilibrium (94%) within 2 d. OECAM could be used to determine the concentration of EPO in soil porewater. Furthermore, a significant linear relationship (R2 = 0.990) was observed between the EPO concentrations in earthworms and the OECAMs. The EPO concentrations in the porewater and OECAMs were lower in soils with a higher organic matter (OM) content. The EPO concentrations in the porewater, earthworms, and OECAMs decreased by 64.4, 49.0, and 56.1%, respectively, in the presence of 0.5% biochar, compared with the control. Furthermore, the use of OECAMs versus earthworms for soil testing also allows you to avoid factors that increase variance in organisms, such as avoidance behaviors or feeding. Therefore, OECAMs show good potential for use as a passive sampler to evaluate the bioavailability of EPO.

Microplastics reduce the bioaccumulation and oxidative stress damage of triazole fungicides in fish.

Microplastics (MPs) and pesticides are typical representatives of harmful chemicals in polluted waters. It is understood that the combined toxicity may differ from that of a single toxic substance. Although their combined toxicities on aquatic organisms have practical significance and research value, they have received little attention due to their complicated interaction, and the mechanism has rarely been reported. In this paper, we designed a study to investigate the single and combined effects of polystyrene microplastics (PS-MPs) and the triazole fungicide difenoconazole on zebrafish, and to explore the mechanism of this effect. The results showed that PS-MPs could reduce the bioaccumulation of difenoconazole in zebrafish to a certain extent and alleviate the oxidative stress damage of difenoconazole in the zebrafish liver. The transcriptome and qRT-PCR data revealed the association of multiple pathways in the difenoconazole response, while the presence of PS-MPs ameliorated this effect in gene expression changes. Due to the properties of PS-MPs and the interaction between them, the toxic effect of difenoconazole when combined with PS-MPs is more prominent. These results provide a novel aspect to understand the environmental behavior of MPs and to evaluate the combined effect of MPs and pesticides on aquatic food.

Degradation of difenoconazole in water and soil: Kinetics, degradation pathways, transformation products identification and ecotoxicity assessment.

Difenoconazole is a widely used triazole fungicide that has been frequently detected in the environment, but comprehensive study about its environmental fate and toxicity of potential transformation products (TPs) is still lacking. Here, laboratory experiments were conducted to investigate the degradation kinetics, pathways, and toxicity of transformation products of difenoconazole. 12, 4 and 4 TPs generated by photolysis, hydrolysis and soil degradation were identified via UHPLC-QTOF/MS and the UNIFI software. Four intermediates TP295, TP295A, TP354A and TP387A reported for the first time were confirmed by purchase or synthesis of their standards, and they were further quantified using UHPLC-MS/MS in all tested samples. The main transformation reactions observed for difenoconazole were oxidation, dechlorination and hydroxylation in the environment. ECOSAR prediction and laboratory tests showed that the acute toxicities of four novel TPs on Brachydanio rerio, Daphnia magna and Selenastrum capricornutum are substantially lower than that of difenoconazole, while all the TPs except for TP277C were predicted chronically very toxic to fish, which may pose a potential threat to aquatic ecosystems. The results are important for elucidating the environmental fate of difenoconazole and assessing the environmental risks, and further provide guidance for scientific and reasonable use.

Risk assessment for pesticide mixtures on aquatic ecosystems in China: a proposed framework.

BACKGROUND: With over 12 000 registered mixture formulations in China, environmental risk assessment of co-formulated pesticide mixtures to aquatic ecosystems is of increasing interest to scientists, risk assessors, and risk managers. This study proposed a pragmatic approach based on widely accepted scientific concepts and strategies. RESULTS: The proposed approach has three tiers, a preliminary Tier 0 stage, a Tier 1 stage, and a refinement stage as Tier 2, which are demonstrated with one case study. Because of the large number of co-formulated pesticide mixtures available in China, a unique Tier 0 stage is proposed with practicable criteria to identify mixtures requiring further evaluation. For Tier-1 assessment, exposure is assessed based on the application scheme and local scenarios in China. Hazard is evaluated using the concentration addition model as the default approach, which was developed a century ago and was adopted by the majority of the regulators globally. Given the unique characteristics of mixtures in the environment, data variability and ratio changes of active ingredients are also evaluated for hazard assessment. More accurate assessments with decreased uncertainties can be achieved by higher tier refinements in Tier-2, such as alternative models, consideration of co-formulants, and higher tier effect and exposure analyses. CONCLUSION: Widely accepted concepts, methodologies, and criteria should be used for aquatic risk assessment for pesticide mixtures in China. To avoid unnecessary workload and a more accurate assessment, unique perspectives such as local agricultural practice, local scenarios and a preliminary screening stage (Tier 0) should be implemented. © 2019 Society of Chemical Industry.

Joint Toxicity of Acetamiprid and Co-Applied Pesticide Adjuvants on Honeybees under Semifield and Laboratory Conditions.

The evaluation of adverse effects of pesticides, pesticide adjuvants, and their combination on honeybees is hampered by a lack of colony-level bioassays reflecting productivity and survival over longer term exposure. In the present study, the joint toxicity of acetamiprid and co-applied pesticide adjuvants (N-methyl pyrrolidone [NMP], Silwet L-77, and Triton X-100) to honeybees was determined both in the laboratory and under semifield conditions. The 3 pesticide adjuvants caused no significant acute toxicity to honeybees by themselves; however, in the laboratory tests, they significantly increased the acute contact toxicity of acetamiprid to honeybees. For the semifield tests, in the T2 group (treatment with 5% acetamiprid soluble concentrate [SL] containing 10% Silwet L-77), the mortality of honeybees was significantly higher (p < 0.05) than that of the blank control on the fourth day after application (DAA + 4), that of the T1 group (5% acetamiprid SL containing 10% NMP) on DAA + 4 and DAA + 7 (seventh day after application), and that of the T3 group (5% acetamiprid SL containing 10% Triton X-100) on DAA + 4. Furthermore, the flight intensity in the T2 group on DAA + 7, the colony intensity on DAA + 28 (28th day after application), and the mean areas covered by pupae on DAA + 15 (15th day after application) were significantly lower (p < 0.05) than those of the blank control. Therefore, pesticide adjuvants may be important factors in increasing the toxicity of neonicotinoids to honeybees. Measures should be taken to manage the environmental risk of pesticide adjuvants during the process of formulation development and registration. Environ Toxicol Chem 2019;38:1940-1946. © 2019 SETAC.

Assessing the risk of resistance in Pseudoperonospora cubensis to the fungicide flumorph in vitro.

BACKGROUND: The oomycete fungicide flumorph is a recently introduced carboxylic acid amide (CAA) fungicide. In order to evaluate the risk of developing field resistance to flumorph, the authors compared it with dimethomorph and azoxystrobin with respect to the ease of obtaining resistant isolates to these fungicides, the level of resistance and their fitness in the laboratory. RESULTS: Mutants with a high level of resistance to azoxystrobin were isolated readily by adaptation and UV irradiation, and their fitness was as good as that of the parent isolates. Attempts to generate mutants of Pseudoperonospora cubensis (Burk. & MA Curtis) Rostovsev resistant to flumorph and dimethomorph by sporangia adaptation on fungicide-treated leaves were unsuccessful. However, moderately resistant mutants were isolated using UV mutagenesis, but their resistance level [maximum resistance factor (MRF) < 100] was much lower than that of the azoxystrobin-resistant mutant (MRF = 733). With the exception of stability of resistance, all mutants showed low pathogenicity and sporulation compared with wild-type isolates and azoxystrobin-resistant mutants. There is cross-resistance between flumorph and dimethomorph, suggesting that they have the same resistance mechanism. CONCLUSION: The above results suggest that the resistance risk of flumorph may be similar to that of dimethomorph but lower than that of azoxystrobin and can be classified as moderate. Thus, it can be managed by appropriate product use strategies.

Vegetative Compatibility of Fusarium graminearum Isolates and Genetic Study on Their Carbendazim-Resistance Recombination in China.

ABSTRACT Monoconidial isolates of 33 carbendazim-sensitive isolates and 31 carbendazim-resistant isolates of Fusarium graminearum were selected from three regions of China for vegetative compatibility group (VCG) analysis. A total of 213 and 224 nit mutants were recovered from the 33 sensitive and the 31 resistant isolates, respectively. Of all the nit mutants, the frequency of the different phenotypes was 44.6, 46.5, 5.7, and 3.2% for nit1, nit3, nitM, and nitA, respectively. VCG analysis identified 30 different VCGs among the 33 sensitive- and the 31 carbendazim-resistant isolates, with VCG diversity 0.91 and 0.97, respectively. Both, a carbendazim-sensitive and a -resistant isolate from the same field belonged to the same VCG. In all then, a total of 59 VCGs were identified among the 64 isolates with an overall VCG diversity 0.92. Direct hyphal fusion was observed in six pairs of vegetatively compatible complements, which is evidence of heterokaryon formation. It was hypothesized that carbendazim resistance could not be transferred by hyphal fusion or there is a small chance to be transferred between two compatible isolates. Three stable sexual recombinants of F. graminearum were randomly chosen from each of the three genetic crosses to study their biological properties. There were no significant differences in mycelial linear growth and pathogenicity between recombinants and their parents, but they differ in sporulation ability and capacity to produce perithecia. We concluded that sexual recombination presumably played a role in the development of carbendazim resistance under field conditions.

Characterization and fitness of carbendazim-resistant strains of Fusarium graminearum (wheat scab).

BACKGROUND: Carbendazim (MBC) has failed to control wheat scab, caused by Fusarium graminearum Schwabe, on the eastern coast of China in recent years after about 30 years of application. RESULTS: MBC resistance was found to be common in pathogen populations on the eastern coast and along areas of the Yangtze River. EC(50) and minimum inhibitive concentration (MIC) values of MBC inhibiting mycelium growth of wild-type isolates were less than 0.9 and 1.4 microg mL(-1) respectively, while EC(50) values of resistant collections averaged 7.02 +/- 11.86 microg mL(-1). The slope of the MBC dosage-response curve (DRC) for resistant isolates of F. graminearum was flat: 1 < b < 2.8 for resistant isolates and 3.5 < b < 11 for sensitive isolates). Both field resistant and sensitive MBC strains shared similar temperature sensitivity, fitness and virulence on ears. Field resistant strains and UV-induced mutants showed positive cross-resistance to other benzimidazole derivatives and were mainly at intermediate MBC resistance level. Highly resistant field MBC strains rarely appeared, but only some of the highly resistant MBC UV mutants were insensitive to N-phenylaminecarbamates. No mutation in beta-tubulin was found in F. graminearum, in contrast to mutation in this tubulin which has led to MBC resistance in other plant pathogens. CONCLUSION: MBC(R) isolates have high fitness and competition in field, conferred by a novel molecular mechanism.

Simultaneous enantioselective determination of phenylpyrazole insecticide flufiprole and its chiral metabolite in paddy field ecosystem by ultra-high performance liquid chromatography/tandem mass spectrometry.

A novel and sensitive ultra-high performance liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) method was developed and validated for simultaneous enantioselective determination of flufiprole and its hydrolysis metabolite in paddy field ecosystem. The separation and determination were performed using reversed-phase chromatography on a novel cellulose chiral stationary phase, a Lux Cellulose-4 (150 mm × 2.0 mm) column, under isocratic conditions at 0.25 mL/min flow rate. The effects of other four different polysaccharide-based chiral stationary phases (CSPs) on the separation and simultaneous enantioseparation of the two target compounds were also evaluated. The elution orders of the eluting enantiomers were identified by an optical rotation detector. Modified QuEChERS (acronym for Quick, Easy, Cheap, Effective, Rugged and Safe) method and solid-phase extraction (SPE) were used for the enrichment and cleanup of paddy water, rice straw, brown rice and paddy soil samples, respectively. Parameters including the matrix effect, linearity, precision, accuracy and stability were evaluated. Under the optimal conditions, the mean recoveries for all enantiomers from the above four sample matrix were ranged from 83.6% to 107%, with relative standard deviations (RSD) in the range of 1.0-5.8%. Coefficients of determination R(2)≥0.998 were achieved for each enantiomer in paddy water, rice straw, brown rice and paddy soil matrix calibration curves within the range of 5-500 µg/kg. The limits of quantification (LOQ) for all stereoisomers in the above four matrices were all below 2.0 µg/kg. The methodology was successfully applied for simultaneously enantioselective analysis of flufiprole enantiomers and their chiral metabolite in the real samples, indicating its efficacy in investigating the environmental stereochemistry of flufiprole in paddy field ecosystem.

[Induction and genetic analysis of laboratory mutants of Gibberella zeae resistance to carbendazim].

The mutants of wild-type Gibberella zeae resistance to carbendazim were generated in laboratory by ultra-violet (UV) irradiating and fungicide taming. Two levels of resistance (low, LR; high, HR) were identified among these mutants. LR mutants could grow at the critical concentration of 1.4 +/- microg/mL of carbendazim while completely inhibited above 10 microg/ mL concentration, but this phenotype of isolates was not detected in the field. HR mutants could grow at 100 microg/mL and showed negative cross-resistance with N-phenylcarbamate fungicides, such as N-( 3,5-dichlorophenyl) carbamate (MDPC) whereas the LR mutants were not. Moderate resistant (MR) mutants which could grow fast at 10 microg/mL, but completely inhibited at 100 microg/mL, were not generated in laboratory. HR mutants of field MR isolate were also generated by fungicide taming on 100 microg/mL of carbendazim, but these mutants did not express negative cross-resistance to MDPC. The genetic study suggested that the carbendazim-resistance in these mutants could be steadily inherited by asexual and selfed reproduction, and the resistance was controlled by the same single major gene both in laboratory mutants and field resistant isolates, the different levels of resistance or the same resistant level in different strains maybe conferred by mutations at different sites or one site with different allelic mutations. The gene compensates for the sensitivity to MDPC was allelic to that governs the carbendazim resistance, but the mutation for increasing sensitivity to MDPC in this gene could get the pathogen highly resistant to carbendazim.

[Genetics of resistance to carbendazim in Gibberella zeae].

According to the ability of the field isolates of Gibberella zeae to grow on the PSA with varying carbendazim(MBC) concentrations, three sensitivity levels of isolates were determined in vitro. The sensitive isolates(S) could grow at 0.5 microgram/ml, but were completely inhibited at 1.4 micrograms/ml. The moderate resistant isolates (MR) could grow fast at 1.4 micrograms/ml and slow at 50 micrograms/ml, but could not grow at 100 micrograms/ml. The high resistant isolates(HR) could grow faster than R at 50 micrograms/ml, and also could grow at 100 micrograms/ml. No low resistant isolates, that could grow fast at 1.4 micrograms/ml but could not grow at 50 micrograms/ml, were found among the field isolates. The genetics of resistance to carbendazim in G. zeae was investigated by analyzing the sensitivity of sexual outcrossed progeny to MBC. The nitrate non-utilizing mutant (nit) as an another added genetic marker was used to select the out-crossed perithecium from self-crossed perithecia. Seven crosses were tested in all, including cross between S x S, MR x S, MR x MR, HR x S, as well as HR x MR. The results showed that no recombinant phenotype was found among all progeny of seven crosses. Progeny segregation of crosses between the parents with different sensitive levels(i.e. MR x S, HR x S, HR x MR) clearly fit a 1:1 ratio of two parental phenotypes, and no segregation was found in the crosses of S x S and MR x MR. So it can be concluded that the two levels of MBC resistance in G. zeae are conferred by two loci mutations or one locus with different allelic mutations that constitute a polymorphic series in a single Mendelian gene. In these isolates, the MBC resistance is not affected by modifying genes or cytoplasmic components.