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The morphology, morphogenesis and molecular phylogeny of a hypotrichous ciliate, Lamtostyla granulifera sinensis subsp. nov., isolated from northern China, were investigated. This population appeared highly similar in morphology to L. granulifera Foissner, 1997. However, on detailed investigation some non-overlapping features were identified, i.e., the body shape and the arrangement of the cortical granules. These differences suggested the separation at subspecies level. Furthermore, the morphogenesis of the new subspecies is described, which is characterized by: (1) the posterior part of the parental adoral zone of membranelles is renewed; (2) the amphisiellid median cirral row is formed from two anlagen; and (3) the frontoventral-transverse cirral anlagen II to VI generate one transverse cirrus each. Phylogenetic analyses based on SSU rDNA sequence data show that Lamtostyla species are scattered in different clades. The monophyly of the genus Lamtostyla is also rejected by the AU test in this study.

A homogeneous fluorescence quenching immunoassay is described for simultaneous separation and detection of aflatoxin M1 (AFM1) in milk. The novel assay relies on monoclonal antibody (mAb) functionalized Fe3O4 decorated reduced-graphene oxide (rGO-Fe3O4-mAb) as both capture probe and energy acceptor, combined with tetramethylrhodamine cadaverine-labeled aflatoxin B1 (AFB1-TRCA) as the energy donor. In the assay, AFB1-TRCA binds to rGO-Fe3O4-mAb in the absence of AFM1, quenching the fluorescence of TRCA by resonance energy transfer. Significantly, the immunoassay integrates sample preparation and detection into a single step, by using magnetic graphene composites to avoid washing and centrifugation steps, and the assay can be completed within 10 min. Under optimized conditions, the visual and quantitative detection limits of the assay for AFM1 were 50 and 3.8 ng L−1, respectively, which were significantly lower than those obtained by fluorescence polarization immunoassay using the same immunoreagents. Owing to its operation and highly sensitivity, the proposed assay provides a powerful tool for the detection of AFM1.

A homogeneous fluorescence quenching immunoassay is described for simultaneous separation and detection of aflatoxin M

Microcystin-LR (MC-LR) is regarded as one of the most toxic microcystins (MCs) isoforms. Microcystins could cause multiple organs dysfunction, and more attention has been drawn to the toxic effects on the gastrointestinal disorder. By using ex vivo everted gut sac model in 6 fish (Carassius auratus, Megalobrama amblycephala, Hypophthalmichthys molitrix, Aristichthys nobilis, Ctenopharyngodon idellus and Cyprinus carpio) and determining the accumulation of MC-LR in zebrafish intestine, we found a dose-dependent manner in the absorption and accumulation of MC-LR. Until now, little studies have been reported concerning the gut microbiota composition caused by different MC-LR exposure. The present study is the first time characterized the phylogenetic composition and taxonomic of the bacterial communities growth in the intestines of zebrafish treated with MC-LR using 16S rRNA pyrosequencing. After 30 days of treatment with 0, 1, 5 or 20 μg/L MC-LR, the alpha and beta diversity did not generate significant differences, indicating the existence of a core microbiota. However, db-RDA analysis showed that treatment with 20 μg/L MC-LR changed the characteristics of high abundances microbiota. The expression of Oatp2b1, stress related enzyme activities in gut and their associations with gut microbiota were also determined. The identified phylotypes including Actinobacteria, Lactobacillus and some opportunistic pathogens highlight the increasing risks of pathogen invasion and recovery tendency via potential probiotics resistance in zebrafish exposed to MC-LR.

Obtaining oocytes from the adult female zebrafish (Danio rerio) ovary has enormous importance in the studies of developmental biology, toxicology, and genetics. It is vital to establish a simple and effective approach to ensure the quantity and quality of oocytes, which will enable the success of follow-up experimental investigation finally. Usually, oocytes are separated with mechanical or enzymatic methods, however, little studies have been done with concerns about the comparative effects. The present study separated zebrafish oocytes of Stage III with five frequently used methods, including stripping, pipetting, hyaluronidase (1.6 mg/ml), collagenase (0.4 mg/ml), and trypsin (0.1%). The cell viability, oxidative stress, mitogen-activated protein kinase (MAPK) protein phosphorylation, and apoptosis levels were selected as main biomarkers to evaluate the oocytes health status. The results showed that both trypsin and hyaluronidase isolation significantly upregulated germinal vesicle breakdown (GVBD) rates and downregulated p38 MAPK activity simultaneously. GVBD rates and survival rates were decreased notably in oocytes separated by the collagenase method. Above results indicate that zebrafish oocytes in vitro are sensitive to enzymatic treatments and the enzymatic isolation is not the suitable mean for collecting zebrafish oocytes although it is time-saving. The mechanical strategy of pipetting remarkably increased the reactive oxygen species and malondialdehyde level in isolated oocytes. Interestingly, oocytes separated with stripping show less physiological and biochemical damages. Therefore, stripping isolation is comparatively recommended as the optimum method for separating and collecting numerous intact and healthy zebrafish oocytes in vitro for the subsequent developmental research.

The essence of epigenetics is the study of heritable changes in gene expression and function that occur without a change in DNA sequence. Epigenomics is the study of epigenetic variation at the genomic level. Herb epigenomics focuses on molecular mechanisms underlying secondary metabolism, growth and development, and stress responses in medicinal plants and fungi on a genome-wide scale by employing molecular biology, bioinformatics, and “omics” technologies as well as pharmacological mechanisms underlying epigenetic modifications brought about by active ingredients of traditional Chinese medicine (TCM) (Dao-di). Herb epigenomics will provide a theoretical basis for analyzing environmental factors responsible for the genuineness of TCM and new pharmacological mechanisms underlying it. Comparative epigenetics is the key method used in herb epigenomics. This study summarizes the current status of research pertaining to herb epigenomics and discusses the prospect of herb epigenomics for clarifying biological and new pharmacological mechanisms underlying TCM.

The biosynthesis of sialic acid (Neu5Ac) leads to the intracellular production of cytidine-5'-monophospho-N-acetylneuraminic acid (CMP-Neu5Ac), the active sialic acid donor to nascent glycans (glycoproteins and glycolipids) in the Golgi. UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase myopathy is a rare autosomal recessive muscular disease characterized by progressive muscle weakness and atrophy. To quantify the intracellular levels of CMP-Neu5Ac as well as N-acetylmannosamine (ManNAc) and Neu5Ac in human leukocytes, we developed and validated robust liquid chromatography-tandem mass spectrometry methods. A fit-for-purpose approach was implemented for method validation. Hydrophilic interaction chromatography was used to retain three hydrophilic analytes. The human leukocyte pellets were lysed and extracted in a methanol-water mixture and the leukocyte extract was used for LC-MS/MS analysis. The lower limits of quantitation for ManNAc, Neu5Ac and CMP-Neu5Ac were 25.0, 25.0 and 10.0 ng/ml, respectively. These validated methods were applied to a clinical study.

Cerebrospinal fluid (CSF) culture is the gold standard for diagnosing postoperative central nervous system infection. The time to positivity (TTP) of an automated continuous blood culture system may indicate the original concentration of the organism. Coagulase-negative Staphylococcus (CoNS), the common organism recovered in CSF, poses difficulty in differentiating infection from contamination. This study investigated the TTP of CSF culture with CoNS and its relationship to clinical parameters and prognosis.Adult neurosurgical patients with CoNS who recovered via the use of CSF culture in BacT/ALERT Pediatric FAN blood culture bottles and were admitted from September 2013 to July 2015 were enrolled. The demographics, clinical and microbiological data, and treatment were reviewed, and the TTP of each culture was retrieved.Thirty-nine adult patients with CoNS recovered from CSF culture were included. The TTP ranged from 7.68 to 57.36 hours. A univariate logistic regression analysis indicated patients with rapid TTP (21.5 hours) compared with those with longer TTP were more likely to be female, show an effective response to antibiotic therapy within 7 days, have clean-contaminated surgical incisions, and show CSF leak. A multivariate logistic regression analysis indicated that being female, an effective antibiotic therapy within 7 days, and clean-contaminated surgical incisions were independent predictors of rapid TTP.Targeted antibiotic therapy was more likely to be beneficial to patients with a rapid TTP within 7 days, which suggested that CoNS with a rapid TTP represents the pathogen of central nervous system infection rather than contamination in neurosurgical patients.

Background: Toxic cyanobacterial blooms result in the production of an organic biomass containing cyanotoxins (e.g. microcystins) and an elevated ammonia concentration in the water environment. The ingestion of toxic cyanobacteria and exposure to ammonia are grave hazards for fish. The present study assessed the effects of dietary toxic cyanobacteria and ammonia exposure on the flesh quality of blunt snout bream (Megalobrama amblycephala).; Results: Dietary toxic cyanobacteria and ammonia exposure had no impact on fish growth performance, fillet proximate composition and drip loss, whereas it significantly decreased fillet total amino acids, total essential amino acids, hardness and gumminess, and increased fillet ultimate pH as well as malondialdehyde content. However, there was no significant interaction between dietary toxic cyanobacteria and ammonia exposure on these parameters. Additionally, dietary toxic cyanobacteria significantly increased fillet initial pH, thaw loss and protein carbonyl content, whereas ammonia exposure did not.; Conclusion: The results of the present study indicate that dietary toxic cyanobacteria and ammonia exposure reduced the quality of blunt snout bream fillet. © 2016 Society of Chemical Industry.; © 2016 Society of Chemical Industry.

IgZ is considered to be the last immunoglobulin discovered in vertebrate species. In this study, the structure of secreted form of blunt snout bream (Megalabrama amblycephala) IgZ (sIgZ) heavy chain gene is VH-Cζ1-Cζ2-Cζ3-Cζ4, in which Cζ4 provides the specificity to the IgZ isotype. The deduced amino acid sequence of sIgZ shows highest similarity with that of Ctenopharyngodon idella (79%). The ontogeny of sIgZ gene expression showed a V-shape change pattern: decreased initially from unfertilized egg stage to 16-cell embryos and increased significantly from blastula stage, which exhibited maternal transmission effects. Compared with the juvenile fish, sIgZ mRNA level was higher in head kidney, spleen, trunk kidney, liver, intestine and gill of adult fish. In both juvenile and adult fish, sIgZ mRNA was detected in intestine and gill. Aeromonas hydrophila challenge experiment showed that sIgZ transcription significantly increased in skin, gill and intestine, indicating a prominent mucosal immune response. The results of Western blot also verified the protein alterations of sIgZ in mucosal organs in M. amblycephal. Our studies indicate a prominent role of IgZ in mucosa-associated lymphoid tissue immunity and further support the specialized role of IgZ in teleost mucosal immune responses.

This study aimed to improve the sensitivity of lateral flow immunoassay (LFIA) using two strategies, including using a high sensitivity label and an indirect-labeling format, which were simultaneously applied to develop a highly sensitive LFIA to detect 22 β-lactams. In the new format, β-lactam receptors were used without any labeling, and anti-receptor monoclonal antibodies (mAbs) were labeled with amorphous carbon nanoparticles (ACNs). The new LFIA is based on the following steps: 1) screening mAbs against the receptors for antigen-antibody binding sites away from the active site of the receptors; and 2) detecting immune complexes using receptor-mAb-ACNs. The cut-off values against ampicillin, amoxicillin, benzylpenicillin, dicloxacillin, cloxacillin, oxacillin, nafcillin, cefquinome, cefoperazone, cefapirin, ceftazidime, cephalothin, cefepime, cefoxitin, cefazolin, ceftriaxone, ceftiofur, cefotaxime, cefalexin, imipenem, meropenem, and ertapenem were 1, 2, 2, 10, 10, 10, 10, 3, 3, 3, 5, 10, 10, 20, 20, 20, 30, 30, 100, 10, 15, and 15 ng/mL, respectively, which is significantly lower than that of receptor-mAb-colloidal gold-LFIA using the same immune reagents.

Cyanobacterial blooms and their secondary metabolites, microcystins (MCs), are not only toxic to aquatic organisms, but also to humans. MCs exert reproductive toxicity in female fish by affecting the oocyte development. However, the mechanism behind MC-LR interference in oocyte development remains largely unknown. In our study, adult female zebrafish were exposed to MC-LR (0, 1, 5, 20 μg/L) for 30 d. After exposure to MC-LR for 30 d, fertilized eggs from the treated females and healthy males were collected and cultured in water without MC-LR. Histomorphological observations showed pathological damage in the ovary after MC-LR exposure, which was mainly characterized by enlarged intercellular spaces, detachment of follicular cells from oocytes, and vacuolation of parenchymal tissues. The 20 μg/L MC-LR treatment caused a remarkable increase in the rate of the zebrafish oocytes germinal vesicle breakdown (GVBD) and a significant decrease in the levels of cyclic adenosine monophosphate (cAMP) and vitellogenin (VTG). In addition, the phosphorylation levels of the extracellular signal-regulated kinases (ERK) were elevated in ovaries from zebrafish exposed to 5 and 20 μg/L MC-LR, and cyclinB phosphorylation levels were also upregulated notably in the 20 μg/L MC-LR group. However, MC-LR exposure did not cause any change in the levels of cAMP-dependent protein kinase (PKA) protein and cdc2 phosphorylation in all the treatments. All the doses of MC-LR reduced the number of eggs, prematurely hatched the fertilized eggs and increased the abnormal rate of offspring generation. In summary, the present study demonstrates that MC-LR promotes oocyte maturation by activating the ERK1/2 and MPF signaling pathways, and cAMP is involved in this process.

The existence of microcystins (MCs), the secondary metabolite of cyanobacteria, has become a growing public health concern. Previous researches have proved that MCs can trigger endoplasmic reticulum stress (ERS), but the underlying mechanisms remain unclear. In the present study, adult female zebrafish were exposed to MC-LR (0, 1, 5 and 20 μg/L) for 30 d, and the offspring derived from the treated females and healthy males were cultured in water without MC-LR until 96 h post fertilization (hpf). Our data suggested that MC-LR causes a significant increase in the eif2s1a, atf4, and eif2ak3 transcription levels in the liver and ovary. The mRNA levels of atf4, atf6, bcl-2, hspa5, eif2s1a and eif2ak3 upregulated notably in the offspring. JNK phosphorylation level and cleaved-caspase3 protein expression elevated obviously in the liver and ovary, but had no remarkable change in the offspring. Furthermore, TUNEL results showed that MC-LR significantly induced apoptosis in the liver and ovary, while acridine orange (AO) staining indicated that MC-LR did not cause abnormal apoptosis in offspring. Concisely, the present study indicated that MC-LR leads to apoptosis through different ERS pathways in the liver, ovary and offspring, and also provides a new perspective for understanding the apoptosis caused by MC-LR.

Harmful cyanobacteria and their production of microcystins (MCs) exert significant toxicity on reproduction of fish, especially the process of oogenesis. Our previous studies demonstrated that MCs have negative impacts on the quantity and quality of mature oocytes in female zebrafish. However, the underlying mechanisms of MCs disrupting oocyte maturation (OM) have been rarely reported. In the present study, in vitro oocytes (immature) were separated from zebrafish and treated with 1, 10, 100 μg/L MC-LR. The serine/threonine protein phosphatase 2A (PP2A) activity was downregulated significantly in oocytes exposed to 10 and 100 μg/L MC-LR for both 2 and 4 h. The phosphorylation levels of mitogen-activated protein kinase (MAPK) were detected without noticeable change in all oocytes treated with MC-LR for 2 h, whereas the activated levels of MAPK subtypes (ERK, p38 and JNK) increased remarkably in the 100 μg/L MC-LR treatment of 4 h. In the oocytes exposed to 100 μg/L MC-LR for 4 h, germinal vesicle breakdown (GVBD) rates changed abnormally and maturation-promoting factor (MPF) activity increased significantly, in accordance with the upregulation of Cyclin B protein levels. Moreover, the MAPK inhibitors (10 μM) were applied to explore the role of MAPK subtypes during MC-LR influencing OM and results showed that ERK inhibitor U0126 and p38 inhibitor SB203580 mitigated the effects of 100 μg/L MC-LR-induced MAPK hyper-phosphorylation and elevated GVBD in the oocytes. In conclusion, the present study indicates that microcystins disrupt the meiotic maturation by the pathway of MC-PP2A-MAPK-OM due to the phosphorylation disorder in oocytes.

Immunoglobulins (Igs), which bind antigens with high specificity, are essential molecules in adaptive immune system of jawed vertebrates. In this study, cDNA encoding the secreted form of the immunoglobulin heavy chain of IgM (sIgM) was cloned from the mesonephros of blunt snout bream (Megalabrama amblycephala) using RT-PCR and rapid amplification of cDNA ends (RACE). The full-length cDNA of sIgM heavy chain gene has 1961 nucleotides encoding a putative protein of 569 amino acids, constant region shares high amino acid identity with that of Ctenopharyngodon idella (80%), Carassius auratus langsdorfii (65%) and Danio rerio (59%). Multiple protein sequence alignment revealed that blunt snout bream sIgM was clustered with the homologues of cyprinid fish and constructed one clade. Using quantitative real-time PCR (qRT-PCR) analysis, the level of sIgM mRNA was determined, with a V-shape change pattern: decreased initially from unfertilized egg stage to 4 cells stage and increased from 16 cells stage to prelarva. This sharp drop indicates that sIgM mRNA is maternally transferred, and was continuously degraded until 16 cells stage. The drastic rising in sIgM level from blastula stage to prelarva might be attributed to embryonic stem cell differentiation procedure. Compared with juvenile fish, the expression of sIgM was significantly higher in pronephros, liver, spleen, gill and muscle of adult fish. After the injection of Aeromonas hydrophila, the expression pattern of sIgM was found first down-regulated at 4 h, then up-regulated and reached the peak at 7 d and 21 d in mesonephros, spleen, liver and gill, respectively.

Microcystins (MCs) released during cyanobacterial blooms exert varied toxicity on fish. Up to now, the reproductive toxicity of MCs on fish has rarely been reported. The present study investigated the reproductive toxicity of microcystin-LR (MC-LR) on male and female zebrafish (Danio rerio) by subchronic immersion in 1, 5, 20 μg/L for 30 d. After MC-LR exposure, the hatchability and the 17 beta-estradiol (E2) concentration in gonads significantly decreased in the 20 μg/L group. In the 5 and 20 μg/L groups, the whole body vitellogenin (VTG) levels significantly increased in females, while considerably decreased in males. The VTG1 transcriptional level significantly reduced in the liver of both female and male treated fish. Marked histological lesions were observed in the livers, ovaries and testes in MC-LR treated fish. Apoptotic rate in the ovaries significantly increased. Significant down-regulation of Bcl-2 transcriptional level was found in the gonads of all MC-LR treated fish, while marked up-regulation of Bax transcription level was determined in the 20 μg/L female treatment group, but a significant down-regulation in males. Although the transcriptional level of caspase-3 dropped in ovaries of the 5 and 20 μg/L treatment groups, the significant increase of caspase-3 activation levels in the ovaries and testes were detected. The present findings indicate that MC-LR exposure exerts diverse reproductive toxicity in zebrafish with females exhibiting more sensitivity than males. The present study also confirmed for the first time that MC-LR does not cause any estrogenic effects in adult zebrafish.

While microcystins (MCs) have been reported to exert reproductive toxicity on fish with a sex-dependent effect, the underlying mechanism has been rarely investigated. In the present study, zebrafish were exposed to 1, 5 and 20 μg/L MC-LR for 30 d. The gonad-somatic index declined in all treated males. 17β-estradiol (E2), testosterone (T), 11-keto testosterone (11-KT) and follicle-stimulating hormone (FSH) levels increased in serum from all treated females, while T, FSH and luteinizing hormone (LH) levels changed in all treated males. Histomorphological observation showed that MC-LR exposure evidently retarded oogenesis and spermatogenesis. Transcriptional changes of 22 genes of the hypothalamic-pituitary-gonad (HPG) axis exhibited sex-specific responses and the relationship between gene transcriptions and gametogenesis was evaluated by principle component analysis (PCA). Major contributors to PC1 (gnrh2, gnrhr3, ar, lhr, hmgra, hmgrb and cyp19a) were positively correlated with the number of post-vitellogenic oocytes, while PC1 (gnrh2, lhβ, erβ, fshr, cyp11a and 17βhsd) were positively correlated with the number of spermatozoa. The protein levels of 17βHSD and CYP19a were affected in both females and males. In conclusion, this study first investigated the sex-dependent effects of microcystins on fish reproduction and revealed some important molecular biomarkers related to gametogenesis in zebrafish suffered from MC-LR.

Cyanobacterial blooms caused by water eutrophication have become a worldwide problem. Microcystins (MCs), especially microcystin-LR (MC-LR), released during cyanobacterial blooms exert great toxicity on fish and even lead to massive death. The present study mainly investigated the pathological damage and immune response of spleen, gut and gill in zebrafish exposed to MC-LR. Fish were exposed to 0, 1, 5 and 20 μg/L of MC-LR for 30 d. In zebrafish exposed to 5 and 20 μg/L MC-LR, edematous mitochondria, deformation of the nucleus and compaction of chromatin were observed in lymphocyte of spleen; frayed gut villi, exfoliation of epithelial cells and widespread cell lyses were observed in intestines; hyperemia in gill lamellae, epithelial tissue edema and uplift and lamellar fusion were observed in gill. Varied changed gene expression was observed in spleen, intestine and gill of zebrafish. The transcriptional levels of IFN-1 and IL-8 in spleen significantly up-regulated in 20 μg/L group, and the transcription of IL-1β and TNFα in spleen increased in 1 μg/L MC-LR treated fish. In addition, the mRNA levels of IFN-1, IL-1β, IL-8, TGF-β and TNF-α dramatically increased in intestine and gill in all MC-LR treated groups. The present studies indicated that MC-LR exposure caused marked pathological damage, however, fish could adjust actively the expression of innate immune-related genes to resist the tissue damage. Our findings provided strong evidence of the recovery potential of fish exposed to microcystins.

Toxic cyanobacterial blooms result in the production of an organic biomass containing cyanotoxins (e.g. microcystins) and an elevated ammonia concentration in the water environment. The ingestion of toxic cyanobacteria and exposure to ammonia are grave hazards for fish. The present study assessed the effects of dietary toxic cyanobacteria and ammonia exposure on the flesh quality of blunt snout bream (Megalobrama amblycephala).Dietary toxic cyanobacteria and ammonia exposure had no impact on fish growth performance, fillet proximate composition and drip loss, whereas it significantly decreased fillet total amino acids, total essential amino acids, hardness and gumminess, and increased fillet ultimate pH as well as malondialdehyde content. However, there was no significant interaction between dietary toxic cyanobacteria and ammonia exposure on these parameters. Additionally, dietary toxic cyanobacteria significantly increased fillet initial pH, thaw loss and protein carbonyl content, whereas ammonia exposure did not.The results of the present study indicate that dietary toxic cyanobacteria and ammonia exposure reduced the quality of blunt snout bream fillet. © 2016 Society of Chemical Industry.

The function of IgD in fish and mammals has not been fully understood since its discovery. In this study, we have isolated and characterized the cDNA that encodes membrane-bound form of the immunoglobulin D heavy chain gene (mIgD) of blunt snout bream (Megalobrama amblycephala) using RT-PCR and rapid amplification of cDNA ends (RACE). The full-length cDNA of mIgD consisted of 3313 bp, encoding a putative protein of 943 amino acids. The structure of blunt snout bream mIgD is VDJ-μ1-δ1-δ2-δ3-δ4-δ5-δ6-δ7-TM. Multiple alignment and phylogenetic analyses indicated that blunt snout bream mIgD clusters with the homologues of cyprinid fish and that its highest identity is with that of C. idella (82%). The mIgD expression in early different developmental stages showed that the level of mIgD mRNA decreased dramatically from the unfertilized egg stage to the 32-cell stage, suggesting that mIgD mRNA was maternally transferred. As cell differentiation initially took place in the blastula stage, the mIgD expression increased significantly from the blastula stage to prelarva, which might be attributed to embryonic stem cell differentiation processes. Compared with juvenile fish, the expression and tissue distribution patterns of mIgD in adult individuals exhibited considerable variation. After the injection of Aeromonas hydrophila, mIgD expression was up-regulated in various tissues, reaching the peak expression at 5 d, 14 d or 21 d (depending on the tissue type). The present study provides a theoretical basis for further research of the teleost immune system.

The outbreak of cyanobacterial blooms induces the production and release of microcystins (MCs) into water, representing a health hazard to aquatic organisms and even humans. Some recent studies have suggested that kidney is another important target organ of MCs except liver, however, the potential toxicity mechanisms are still unclear. In this study, we first investigated the collaborative effect of oxidative stress and cytoskeletal disruption in microcystin-induced apoptosis in CIK (Ctenopharyngodon idellus kidney) cells in vitro. CIK cells were treated with 0, 1, 10, and 100 mu g/L microcystin-LR (MC-LR) for 24 and 48 h. Cell viability was increased by MC-LR in 1 mu g/L group, while decreased in 100 mu g/L group at 48 h. Cell cycle assay showed that 1 and 10 mu g/L MC-LR induced cell cycle through G(1) into S and G(2)/M phases, while 100 mu g/L MC-LR reduced G2/M phase population. MC-LR markedly induced apoptosis in 10 and 100 mu g/L groups. Elevated reactive oxygen species (ROS) production, increased malondialdehyde (MDA) contents, decreased glutathione (GSH) levels, and modulated antioxidant enzymes including catalase (CAT) and superoxide dismutase (SOD) were observed in CIK cells exposed to MC-LR. These alterations were more pronounced at higher doses (10 and 100 mu g/L), indicating that oxidative stress was induced by MC-LR. Laser scanning confocal microscope observation showed aggregation and collapse of microfilaments (MFs) and microtubules (MTs) in CIK cells, and even loss of some cytoskeleton structure. Moreover, transcriptional changes of cytoskeletal genes (beta-actin, Ic3a, and keratin) were also determined, which have a high probability with cytoskeleton structure damage. Our data suggest that oxidative stress and cytoskeletal disruption may interact with each other and jointly lead to apoptosis and renal toxicity induced by MCs. (C) 2015 Elsevier B.V. All rights reserved.

Microcystins (MCs) are algal toxins produced intracellularly within the cyanobacteria cells. MCs exposure exerts great harm to the reproductive system of fish and deteriorates the quality of eggs and sperms, and has further adverse effects on early developmental stages of fish. Whether the MC toxicity can be parentally transmitted to offspring, even though the embryos and larvae are free of MC exposure? In the present study, adult zebrafish were continuously exposed to MC-LR (with dose of 1, 5 and 20 μg/L) for 30 days. After MC-LR exposure, fertilized eggs were collected and the following F1 generation was reared in water containing no MC-LR until 60 days post fertilization (dpf). In F1 offspring, both body weight and body length were evidently dropped. Some growth and immune related genes were detected using the real-time PCR. The transcriptional levels of these genes significantly decreased in F1 offspring of zebrafish whose parents were treated with 5 and 20 μg/L MC-LR. The activities of some antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) significantly dropped in 5 and 20 μg/L MC-LR groups, and the malondialdehyde (MDA) levels markedly increased in all the three treatment groups. Furthermore, distinct pathological changes in liver were observed in F1 zebrafish. Our findings show that the MC-LR exposure to parental zebrafish results in liver damage and evidently influences the growth and immune function in F1 offspring. We consider this damage as a parental transmission effect of microcystin toxicity. Further mechanism studies are necessary to elucidate this transmission effect.