Misgurnus Anguillicaudatus Research Articles

Characterization of neural stem cells and proliferating cells in the brain of loach (Misgurnus anguillicaudatus)

Loach (Misgurnus anguillicaudatus) is an essential commercial fish in China. We aimed to give some notes on brain histology of loach fish and characterize the sites of stem cells in the brain. As well as, we showed the sites of proliferating cells with detection of their glial property. To this end, we histologically observed the structure of the brain via hematoxylin and eosin stain. We also used fluorescent in situ hybridization to evaluate the localization of Msi1, Bmi1, and Sox2. They are essential genes expressed in brain stem cells of loach. Proliferating cells were evaluated by PCNA immunofluorescence. As well as GFAP immunofluorescence was used for detection of the gial property of proliferating cells. Our results indicated that the neural stem cells were located in the granular cell layer of the cerebellum, vagal lobe, facial lobe, optic tectum, and beside the third ventricle because these sites exhibited expression of Sox2. There were several areas of the brain that contained proliferating cells, including the cerebrum, in the torus longitudinalis, and in between the optic tectum and cerebellum. In the cerebellum, the proliferating cells were present in the molecular cell layer, around the intracerebellar ventricle, in the lobus caudalis cerebella, in the eminentia granularis, and around the rhombencephalic ventricle. Proliferating cells were also observed in the three layers of the vagal lobe and throughout the facial lobe. These proliferating cells were all negative for GFAP. The present study provides valuable insight into neural stem cells in loach brain, and these findings can be used to guide future studies.

R-spondin1 in loach (Misgurnus anguillicaudatus): Identification, characterization, and analysis of its expression patterns and DNA methylation in response to high-temperature stress.

With a well-understood function in mammals, R-spondin1 (Rspo1) is an important regulator of ovarian development via the Wnt/β-catenin pathway. Rspo1 deficiency causes retardation of ovarian development in XX fish, and increases Rspo1 function induces femininity and sex reversal in XY fish. In this study, Rspo1 was successfully cloned from loach (Misgurnus anguillicaudatus), and its expression profile was analyzed. The full-length cDNA of Misgurnus anguillicaudatus Rspo1 (MaRspo1) comprised 1322bp and included an open reading frame (ORF) of 795bp, which encoded a predicted polypeptide measuring 264 amino acids in length. Phylogenetic and gene structure analyses showed a highly conserved sequence of MaRspo1 (identical to the Rspo1 genes of other species), consisting of an N-terminal signal peptide (SP), two furin-like cysteine-rich domains (FU1 and FU2), a thrombospondin type 1 repeat (TSP1) and a C-terminal region. Real-time PCR revealed the female-biased expression profile of MaRspo1, with the highest expression level among tested tissues detected in ovary. Investigation of MaRspo1 expression levels throughout the early development stage (10-60days post hatching) under three temperature treatments (25°C, 28°C, and 31°C) revealed significantly differential expression of MaRspo1 among the three temperature groups, with decreased MaRspo1 expression in the high-temperature (31°C) group. The results of DNA methylation analysis indicated that exposure to high temperature during early development can increase the average promoter methylation level of MaRspo1 in both females and males. Taken together, the results of this study provide the basis for the further investigation of the molecular mechanism of Rspo1 in response to temperature.

Toxic effects of zinc and cadium on the benthic organisms in sediments of Lake Poyang and verification of quality guideline

以泥鳅（Misgurnus anguillicaudatus）、铜锈环棱螺（Bellamya aeruginosa）、河蚬（Corbicula fluminea）为受试生物，以死亡率、渗血率、体质量变化、行动迟钝率、挖洞率为测试指标，探究沉积物中Zn和Cd对底栖生物的21 d慢性毒性效应，并对鄱阳湖Zn和Cd的沉积物基准值进行了验证.结果表明，沉积物中Cd对底栖生物的毒性效应强于Zn，沉积物中Zn、Cd对泥鳅、铜锈环棱螺、河蚬的LC₅₀（半数致死浓度）分别为644和61 mg/kg、751和54 mg/kg、652和81 mg/kg，EC₅₀（半数效应浓度）值分别为526和49 mg/kg、589和35 mg/kg、474和45 mg/kg.经验证，沉积物Zn和Cd的TEL（临界效应浓度）、PEL（必然效应浓度）值比较合理，具有一定适用性，即当Zn、Cd的加标含量低于TEL值时，Q_TEL<1，不具有毒性；当加标含量高于PEL值时，Q_PEL>1，毒性明显.;The 21d chronic toxic effects of Zn and Cd in sediments on benthic organisms were investigated by using Misgurnus anguillicaudatus, Bellamya aeruginosa and Corbicula fluminea as the testing organisms, using mortality rate, bleeding rate, average body weight change, sluggishness rate and burrowing rate as the testing indexes, and verified by the sediment reference values of Zn and Cd in Lake Poyang. The results showed that the toxicity of Cd to benthic organisms in sediments was stronger than that of Zn, and the LC₅₀(50% lethal concentration) of Zn and Cd to Misgurnus anguillicaudatus, Bellamya aeruginosa and Corbicula fluminea were 644 and 61 mg/kg, 751 and 54 mg/kg, 652 and 81 mg/kg, and the EC₅₀(50% effective concentration) values were 526 and 49 mg/kg, 589 and 35 mg/kg, 474 and 45 mg/kg, respectively. It has been verified that the TEL (threshold effect level) and PEL (probable effect level) values of the sediments Zn and Cd are reasonable and have certain applicability. When the additive concentration of Zn and Cd is lower than the TEL value, Q_TEL<1 is not toxic. When the additive concentration was higher than PEL value, Q_PEL>1 showed obvious toxicity.

Biotic differentiation in headwater creeks after the massive introduction of non-native freshwater aquarium fish in the Paraíba do Sul River basin, Brazil

Abstract This study evaluated fish beta diversity in six headwater creeks located in the area affected by the largest ornamental aquaculture center implemented in the Minas Gerais State, southeastern Brazil. We sampled fish assemblages in 2017 and 2018 to investigate changes in assemblage structure (species richness and beta diversity), comparing these data with the historic species pool. We recorded 60 fish species, of which 16 were native and 44 non-native with 19 translocated, and 25 exotic. The exotics Poecilia reticulata, Xiphophorus maculatus, X. variatus, Danio rerio, and Misgurnus anguillicaudatus were the most widely distributed in the headwater creeks. The Contamination Index showed that most creeks had high proportional contamination by exotic species (above 60%). Beta diversity increased from historical to contemporary periods in all creeks due to the introduction and differential colonization pressure of several non-native translocated and exotic species, indicating biotic differentiation. Temperature and number of ponds were the main preditors of change in beta diversity in the headwater creeks during the contemporary period. In summary, we observed that invaders have induced substantial changes to fish communities under influence of environmental filters. Our results support the hipothesis that aquaculture is a main driver of fish non-native fish introduction and native biodiversity loss in the Neotropics.

A comparison study of test organism species and methodologies for combined toxicity assay of copper ions and zinc ions.

The general toxicity assays for evaluating the risk of aquatic environment were commonly based on single-species test organism models. Thus, the lack and conflict of the different responses among species had hindered researchers to assess the real toxicity of a target toxicant. Therefore, the difference between the test species and their corresponding methodologies was investigated in this work and three species, Escherichia coli, Saccharomyces cerevisiae and Misgurnus anguillicaudatus (a fish), were chosen as the test organism for typical prokaryotes, eukaryotes, and vertebrates, respectively. More specifically, we investigated (i) the individual and combined toxicity of Cu2+ and Zn2+ by the three test organisms; (ii) the different evaluation manners for the test organisms, including IC50 and toxic unit (TU) model for microorganisms by respiratory toxicity assay and enzyme-substrate assay, while survival time for fish; and (iii) the states of test organism, including suspended and immobilized states for microorganisms. The combined effects, includingsynergistic (Vt < Vp), antagonistic (Vt > Vp) and additive effects for the three species, were complex as that they were usually dose-dependent and could be changed by the different evaluation manners. The present work was useful for enriching of the associated theory and the insights from this work could open the way for further practical risk assessments.

Development and characterization of polyclonal antibodies against subtype specific vitellogenin of the dojo loach, Misgurnus anguillicaudatus

Fish vitellogenin (Vtg) has become an important biomarker for assessing the estrogenic potency of chemicals and the exposure of animals to estrogenic contaminants present in aquatic environments. In the current study, the polyclonal antibodies against subtype-specific Vtgs of the dojo loach (Misgurnus anguillicaudatus) were developed and identified by gene recombination and expression. Immuno-biochemical analyses revealed that VtgAo1 protein appeared to be the major Vtg type in this species. Enhanced chemiluminescent Western blotting was developed using the antiserum against VtgAo1. Exposure of male loach to 17α-ethinylestradiol (EE2) and 17β-estradiol (E2) via water induced the VtgAo1 protein induction with a lowest-observed-effect concentration (LOEC) by 100 ng L−1 and 1000 ng L−1group at 7 day post-initiation (dpi), respectively. The results indicated that this method provided a valuable tool for detecting estrogenic activities in aquatic environments and excluded any difficulties expected in purification procedures to separate a highly pure Vtg subtype from circulating proteins, and the other Vtg subtypes.

Comparative analysis of mitochondrial genomes in genetically distinct groups of the dojo loach Misgurnus anguillicaudatus

Dojo loach (Misgurnus anguillicaudatus) that inhabit Japan are composed of two genetically divergent groups (A and B). Although most individual loach reproduce bisexually, clone lineages exist in certain populations that reproduce gynogenetically. To investigate the molecular phylogenetic relationships among the M. anguillicaudatus groups and clone lineages, complete mitogenomes of members from groups A and B and a clone lineage were sequenced using long range PCR and primer walking methods. The three groups of mitogenomes shared the same gene order and had similar base compositions and codon usage patterns. Phylogenetic analysis indicated group A and the clone lineage were genetically close with group B being genetically divergent.

A C-type lectin with antibacterial activity in weather loach, Misgurnus anguillicaudatus.

C-type lectins are carbohydrate-binding proteins that play important roles in immunity by serving as pattern recognition receptors. In the present study, a novel nattectin-like C-type lectin was obtained from the weather loach, Misgurnus anguillicaudatus, designated as MaCTL. MaCTL encodes a peptide with 165 amino acids, with a signal peptide and a single C-type lectin domain (CTLD), containing a galactose-specific QPD motif and a conserved Ca2+ -binding site. Transcripts of MaCTL were significantly upregulated after immune challenge with its pathogen A.hydrophila. In vitro assays with recombinant MaCTL protein revealed that it exhibited hemagglutinating and bacterial agglutinating activities, in a Ca2+ -dependent manner. MaCTL was found to bind to a wide range of bacteria, as well as bind to bacterial polysaccharides LPS and PGN. Moreover, MaCTL displayed antimicrobial activity by inhibiting the growth of bacteria. These results collectively suggest that MaCTL is involved in the antibacterial defence of weather loach.

Environmental DNA analysis provides an overview of distribution patterns of two dojo loach species within the Naka-ikemi Wetland, Fukui Prefecture, Japan

In the Naka-ikemi Wetland, a cryptic species of the dojo loach, Misgurnus sp. Type I (called Type I loach herein), inhabits along with the common nominal species, Misgurnus anguillicaudatus (called Type II loach herein). In this study, the distribution patterns of two loaches were investigated by environmental DNA (eDNA) analysis and previously suggested habitat segregation between two sympatric loaches was discussed. Type I loach tended to have a limited distribution, mainly inhabiting the northern part of the wetland; Type II loach tended to be widely distributed across the central to southern parts of the wetland. Both loaches occurred sympatrically in some areas throughout the sampling periods, indicating that they did not completely segregate their habitats.

Influence of feeding frequency on diurnal variations in liver and muscle RNA/DNA ratios in Misgurnus anguillicaudatus

We investigated the effect of the feeding regimen (group 1: fed twice daily at 8:00 and 18:00 for 28 days followed by 5 days of fasting; group 2: fed twice daily at 8:00 and 18:00 for 28 days and then fed once daily at 8:00 for 5 days; and group 3: fed twice daily at 8:00 and 18:00 for 33 days) on the circadian rhythms of the RNA/DNA ratio in the liver and muscle of the loach Misgurnus anguillicaudatus. The experiment was carried out with a photoperiod of 14 hr (light):10 hr (dark), with light from 07:30 to 21:30, followed by sampling. Cosine rhythm analysis and parametric analysis revealed obvious diurnal variation regularity in all three groups. The amplitude and phase of diurnal regularity in the RNA/DNA ratios increased after feeding. The highest value in group 2 was higher than that in group 3. Correlation analysis showed a correlation coefficient between the RNA/DNA ratios of the two tissues of 0.85. The coefficients of variation in the maximum RNA/DNA ratio and two cycles of chi-square values in liver tissue were lower than those in muscle tissue, but the stability of the ratio was more reliable and representative in liver than in muscle. These results suggest the occurrence of some altered circadian rhythms of the RNA/DNA ratios in the liver and muscle of the loach M. anguillicaudatus and that the best sampling time was after 4 hr of daily feeding.

Sequence and phylogenetic analysis of the mitochondrial genome for the Poyang Lake, Misgurnus anguillicaudatus (natural diploid loach)

Misgurnus anguillicaudatus is a highly valued, aquaculture-relevant food fish in East Asian countries. In this study, the complete mitochondrial genome of Poyang loach was obtained by PCR. The genome is 16,646 bp in length, including 2 ribosomal RNA genes. 13 protein-coding genes, 22 transfer RNA genes, and a non-coding control region, the gene composition and order of the species were similar to most reported from other vertebrates. The phylogenetic tree showed that Misgurnus family got together for one branch, which includes Poyang M. anguillicaudatus, and the other loaches had their own branches.

Fertility and gamete ploidy of allopentaploid offspring of tetraploid dojo loach Misgurnus anguillicaudatus females and large-scale loach Paramisgurnus dabryanus males

Artificial interspecific hybrids (3n) of tetraploid dojo loach Misgurnus anguillicaudatus (TTTT) and diploid large-scale loach Paramisgurnus dabryanus (PP) were viable. Allopentaploid loaches (TTTTP) were produced by cold-shocking the fertilized eggs of TTTT females fertilized with sperm of PP males. The TTTTP were viable and well developed. As there are currently no reports on the fertility and gamete ploidy of TTTTP, artificial propagation and ploidy analyses were conducted to determine them. The ovaries of TTTTP were well developed with primary oocytes and secondary oocytes; the testes were sterile with low-density sperm-like cells. By backcrossing with mature TTTT, TTTTP were found to be female fertile and male sterile. The fertility of female TTTTP was low; they produced few viable diploid eggs and a few inviable triploid and aneuploid eggs. Ploidy analysis of cold-shock-induced progenies of TTTTP × TTTT indicated that TTTTP could generate viable diploid eggs, along with diploid second polar bodies. The results of this study are helpful for further understanding of loach polyploidization and should encourage more investigations on hybridization and gamete formation in loaches.

Comparative Histological Analysis of Intestines of Loach, Grass Carp and Catfish Provide Insights into Adaptive Characteristics in Air-Breathing Fish

Abstract Accessory respiratory is generally accepted to have evolved independently on numerous occasions in adaption to aquatic hypoxia in freshwater habitats. In general, the air-breathing organ in fish is believed to be structurally modified to supplement respiration. In this study, intuitive evidence for elaborate structural modifications of the intestine, an air-breathing organ in mud loach (Misgurnus anguillicaudatus), compared with two other obligate aquatic breathers, grass carp (Ctenopharyngodon idellus) and yellow catfish (Pelteobagrus fulvidraco), were directly provided by histological and morphometric methods. As a result, a sharply decreasing height of mucosal folds and thickness of muscularis were manifested in loach intestine from its anterior to posterior region. Compared with grass carp and yellow catfish, loach had the smallest ratios of mucosal fold height/muscularis thickness to intestinal lumen radius in the posterior intestine. These suggested that the posterior intestine is the air-breathing location for the loach. Furthermore, length density of capillary (0.46±0.05 μm−2) in the posterior intestine of the loach was significantly higher than those of grass carp and yellow catfish. Meanwhile, diffusion distance of air-blood barrier (1.34±0.04 μm) in the posterior intestine of the loach was significantly smaller than those of the other two fish species. In summary, the characteristics of highly vascularized, short diffusion distance of air-blood barrier, thinned and flattened made the posterior intestine a perfect air-breathing location for the loach.

Transcriptome Analysis of Maternal Gene Transcripts in Unfertilized Eggs of Misgurnus anguillicaudatus and Identification of Immune-Related Maternal Genes.

Maternal genes are important in directing early development and determining egg quality in fish. We here report the de novo transcriptome from four tissue libraries of the cyprinid loach, Misgurnus anguillicaudatus, and for the first time identified maternal gene transcripts in unfertilized eggs and suggest their immune system involvement. Expression profiles and functional enrichment revealed a total 24,116 transcripts were expressed as maternal transcripts in unfertilized eggs, which were involved in a wide range of biological functions and pathways. Comparison expression profiles and analysis of tissue specificity revealed that the large numbers of maternal transcripts were stored in unfertilized eggs near the late phase of ovarian maturation and before ovulation. Functional classification showed a total of 279 maternal immune-related transcripts classified with immune system process GO term and immune system KEGG pathway. qPCR analysis showed that transcript levels of identified maternal immune-related candidate genes were dynamically modulated during development and early ontogeny of M. anguillicaudatus. Taken together, this study could not only provide knowledge on the protective roles of maternal immune-related genes during early life stage of M. anguillicaudatus but could also be a valuable transcriptomic/genomic resource for further analysis of maternally provisioned genes in M. anguillicaudatus and other related teleost fishes.

Full-length transcriptome analysis of Misgurnus anguillicaudatus.

Faithful analysis of transcripts represents a critical component of the gene discovery, genomic characterization and annotation process in species without a reference genome. The present study generated 28,001 full-length transcripts of Misgurnus anguillicaudatus using the Pacific Bioscience (PacBio) single-molecule real-time (SMRT) sequencing system. A total of 77,346 consensus isoforms were identified, and 18,991 complete open reading frames (ORFs) detected. In total, 1345 lncRNAs were identified with high-confidence, with additional identification of a number of well-known transcription factors. The present study may facilitate additional exploration of the genomic signatures of M. anguillicaudatus.

Risk screening of the potential invasiveness of non-native freshwater fishes in the River Ob basin (West Siberian Plain, Russia)

To inform regional managers of potentially invasive non-native (NN) freshwater fishes in the principal hydrosystem that drains the West Siberian Plain, the River Ob basin, 31 extant and potential future NN fish species were screened using the Aquatic Species Invasiveness Screening Kit (AS-ISK) with respect to current and projected future climate conditions. Calibration of the AS-ISK scores, using receiver operating characteristic curve analysis, identified ‘basic risk assessment’ and ‘climate change assessment’ threshold scores of 27.5 and 34.75, respectively, with which to distinguish species that pose a high risk of being invasive in the Ob basin and those that pose a low-to-medium risk. Of the species screened, 12 ranked as high risk (black bullhead Ameiurus melas, brown bullhead Ameiurus nebulosus, grass carp Ctenopharyngodon idella, common carp Cyprinus carpio, eastern mosquitofish Gambusia holbrooki, silver carp Hypophthalmichthys molitrix, oriental weatherfish Misgurnus anguillicaudatus, rainbow trout Oncorhynchus mykiss, Chinese (Amur) sleeper Perccottus glenii, topmouth gudgeon Pseudorasbora parva, brown trout Salmo trutta, pikeperch Sander lucioperca and rudd Scardinius erythrophthalmus). The remaining species ranked as medium or low risk. Although the risk scores increased in 68% of species under climate change conditions, this affected the risk rankings of only two species: Salmo trutta decreased in rank from high to medium and Sander lucioperca increased in rank from medium to high. The outcomes of the present study, which identified 12 species for which full risk assessments are recommended, serves to inform the development of NN species policy and management in Russia.

First Isolation and Characterization of Chryseobacterium cucumeris SKNUCL01, Isolated from Diseased Pond loach (Misgurnus anguillicaudatus) in Korea.

Loaches are widely distributed throughout the natural environment and are consumed for medicinal purposes in East Asia. Usually, loaches are cultured in ponds where the water conditions can easily cause bacterial infections. Infections due to bacterial pathogens such as Aeromonas have been well described in cultured loaches; however, there is no report regarding Chryseobacterium infection. This study focused on the elucidation of the pathogenic and antibiotic resistance characteristics of C. cucumeris, SKNUCL01, isolated from diseased loaches (Misgurnus anguillicaudatus). SKNUCL01 forms a biofilm, which is associated with its virulence. Koch’s postulates were satisfied with a lethal dose 50 (LD50) of 8.52 × 107 colony-forming units (CFU)/ml. Abrasion facilitates the mortality of the fish, which makes it a possible infection route for C. cucumeris. The strain showed resistance to nearly all tested antibiotics, such as trimethoprim/sulfamethoxazole, levofloxacin, and ciprofloxacin, formerly considered effective treatments. Phenotypic analyses for antibiotic resistance—the combined disk test, double-disk synergy test, modified Hodge test, and efflux pump inhibition test—revealed that the resistance of SKNUCL01 originated from metallo-beta lactamases (MBLs) and efflux pumps. Our findings provide evidence that could result in a breakthrough against multidrug-resistant Chryseobacterium infection in the aquaculture industry; the antibiotic resistance-related genes can be elucidated through future study.

A triple WAP domain containing protein acts in antibacterial immunity of weather loach, Misgurnus anguillicaudatus

Whey acidic protein domain (WAPD) occurs in a variety of proteins in animals and many of WAPD-containing proteins are involved in immunity. In the present study, a novel protein containing three WAPDs was identified from the weather loach, Misgurnus anguillicaudatus, designated as MaTWD. MaTWD share high identity with TWDs from fish but low identity with TWDs from other animal phyla. MaTWD transcripts mainly distributed in gills and head kidney responded to bacterial challenge with significant upregulation. In vitro assay with recombinant MaTWD protein revealed that MaTWD had antiprotease activity against bacterial proteases. Moreover, MaTWD exhibited bacterial binding capacity and antimicrobial activity. Most importantly, exogenous MaTWD protected loach against bacterial infection by reducing loach mortality. We infer that MaTWD participates in the antibacterial immunity of loach via its antiprotease and antimicrobial activities.

An effective live attenuated vaccine against Aeromonas veronii infection in the loach (Misgurnus anguillicaudatus)

Aeromonas veronii is a major pathogenic bacterium in humans and animals. When it causes outbreaks, there are enormous economic losses to the aquaculture industry. An effective live attenuated vaccine strain, ΔhisJ, was obtained in our previous studies by gene knockout in Aeromonas veronii TH0426 using the suicide vector pRE112. Here, we evaluated whether the live attenuated vaccine ΔhisJ was suitable for prevention of Aeromonas veronii infection by injection and immersion in loaches. Compared with that of the TH0426 wild-type strain, the virulence of the live vaccine was significantly weakened. Vaccine safety assessment results also indicated that 1 × 107 CFU/mL live vaccine was safe and did not induce clinical symptoms or obvious pathological changes. Additionally, after challenging loaches with Aeromonas veronii TH0426, the relative percent survival of the IN3 injection group was 65.66%, and that of the IM group was 50.78%. Our data show that the live attenuated vaccine ΔhisJ can improve the immune protection rate of loaches. Furthermore, increased enzyme activity parameters (SOD, LZM, ACP, and AKP) in the skin mucus, increased enzyme activity parameters (SOD, LZM, ACP, AKP, and GPx) in the serum, increased specific IgM antibodies and cytokine IL-1β contents in the serum, and increased cytokine (IL-15, pIgR, IL-1β, and TNF-α) expression in the liver and spleen were observed. These data are the first to indicate that the live attenuated vaccine ΔhisJ is suitable for the development of a safe and effective vaccine against Aeromonas veronii infection in loach aquaculture.

A full evaluation of chiral phenylpyrazole pesticide flufiprole and the metabolites to non-target organism in paddy field

Pesticides applied to paddy fields may pose considerable danger to non-target aquatic organisms and further threaten human health. Flufiprole is a pesticide used in rice fields; considering the widespread existence of rice-fish-farming ecosystems, the acute toxicities of flufiprole enantiomers and its six metabolites (fipronil, flufiprole sulfide, flufiprole sulfone, detrifluoromethylsulfinyl flufiprole, desulfinyl flufiprole, and flufiprole amide) to four common aquatic organisms in rice fields including Misgurnus anguillicaudatus (pond loach), Carassius gibelio (Prussian carp), Pelophylax nigromaculatus (black-spotted frog), and Daphnia magna (water flea) were investigated. Genotoxicity, pathological changes and the effects on the antioxidant system of M. anguillicaudatus were also evaluated after exposure. The LC50 (EC50) values showed that fipronil and desulfinyl flufiprole were the most toxic compounds and were approximately about six times as toxic as flufiprole. No enantioselective toxicity was observed between the two enantiomers. The activity of antioxidant defense enzymes and the content of malondialdehyde (MDA) in the liver and gills of M. anguillicaudatus were significantly increased by the chemicals in most cases. In addition, fipronil and desulfinyl flufiprole were found to induce an increase in the micronucleus rate in M. anguillicaudatus. Histopathological analysis showed that the liver of M. anguillicaudatus was not significantly affected by flufiprole. Our study demonstrated a potential negative effect on flufiprole-treated aquatic organisms. As an alternative to fipronil, the environmental risk of flufiprole and its metabolites to non-target organisms in rice fields cannot be ignored.