Are fish fed with cyanobacteria safe, nutritious and delicious? A laboratory study.

Toxic cyanobacterial blooms, which produce cyclic heptapeptide toxins known as microcystins, are worldwide environmental problems. On the other hand, the cyanobacteria protein (30-50%) has been recommended as substitute protein for aquaculture. The present laboratory study verified the feasibility of cyanobacteria protein substitution and risk assessment. Goldfish were fed diets supplemented lyophilised cyanobacteria powder for 16 weeks with the various doses: 0% (control), 10%, 20%, 30% and 40%. Low doses (10% and 20%) promoted growth whereas high doses (30% and 40%) inhibited growth. In cyanobacteria treated fish, the proximate composition of ash, crude fat content and crude protein content decreased in 16 weeks; the saturated fatty acid (SFA) content significantly increased; the n-3 polyunsaturated fatty acid content, collagen content and muscle pH significantly decreased; cooking loss percents increased significantly. Muscle fiber diameter and myofibril length were negatively correlation. Additionally, flavour compounds (e.g., amino acids, nucleotides, organic acids and carnosine) changed significantly in the treated fish, and odour compounds geosmin and 2-methylisoborneol increased significantly. The estimated daily intake (EDI) of microcystins in muscle was close to or exceeded the World Health Organization (WHO) tolerable daily intake (TDI), representing a great health risk. Cyanobacterie is not feasible for protein sources use in aquaculture.

The suppression of hematopoiesis function in Balb/c mice induced by prolonged exposure of microcystin-LR.

Microcystins (MCs) cause normocytic anemia in patients in a hemodialysis unit in Caruaru, Brazil in 1996, but the underlying mechanisms are still unclear. In the present study, Balb/c mice were intraperitoneally injected with microcystin-LR (MC-LR) at the doses of 0.5, 2 and 8 µg/kg body weight (bw) every 48 h for 30 d. After the prolonged exposure of MC-LR, significant decreases of red blood cell count (RBC), hemoglobin (Hb) and hematocrit (Ht) were observed in 2 and 8 µg/kg bw groups, but erythrocyte mean corpuscular volume (MCV) showed no significant changes. Significantly elevated micronucleus frequency was observed in bone marrow cells (BMCs) in all MC-LR treatments. The proliferation of BMCs significantly declined in both 2 and 8 µg/kg bw groups. Serum levels of some hematopoietic growth factors significantly changed in 8 µg/kg bw group, mainly including granulocyte-macrophage (GM-CSF), erythropoietin (EPO), interleukin-3 (IL-3) and TNF-α. The transcriptional levels of these 4 genes in BMCs were also significantly changed in 8 µg/kg bw group. MC-LR exposure significantly increased the apoptosis rates in all MC-LR treatments. The present study indicates prolonged exposure of MC-LR induces normocytic anemia, and the disturbed hematopoietic growth factors and BMCs apoptosis are responsible for this normocytic anemia.

The interactive effects of cytoskeleton disruption and mitochondria dysfunction lead to reproductive toxicity induced by microcystin-LR.

The worldwide occurrence of cyanobacterial blooms evokes profound concerns. The presence of microcystins (MCs) in waters and aquatic food increases the risk to human health. Some recent studies have suggested that the gonad is the second most important target organ of MCs, however, the potential toxicity mechanisms are still unclear. For a better understanding of reproductive toxicity of MCs on animals, we conducted the present experimental investigation. Male rats were intraperitoneally injected with MC-LR for 50 d with the doses of 1 and 10 µg/kg body weight per day. After prolonged exposure to MC-LR, the testes index significantly decreased in 10 µg/kg group. Light microscope observation indicated that the space between the seminiferous tubules was increased. Ultrastructural observation showed some histopathological characteristics, including cytoplasmic shrinkage, cell membrane blebbing, swollen mitochondria and deformed nucleus. Using Q-PCR methods, the transcriptional levels of some cytoskeletal and mitochondrial genes were determined. MC-LR exposure affected the homeostasis of the expression of cytoskeletal genes, causing possible dysfunction of cytoskeleton assembly. In MC-LR treatments, all the 8 mitochondrial genes related with oxidative phosphorylation (OXPHOS) significantly increased. The reactive oxygen species (ROS) level significantly increased in 10 µg/kg group. The mitochondria swelling and DNA damage were also determined in 10 µg/kg group. Hormone levels of testis significantly changed. The present study verified that both cytoskeleton disruption possibly due to cytoskeletal reorganization or depolymerization and mitochondria dysfunction interact with each other through inducing of reactive oxygen species and oxidative phosphorylation, and jointly result in testis impairment after exposure to MC-LR.

Apoptotic responses of zebrafish (Danio rerio) after exposure with microcystin-LR under different ambient temperatures.

Microcystins (MCs) can cause evident hepatic apoptosis. In vitro studies indicated that uptake of MC by isolated hepatocytes was dramatically reduced as ambient temperature dropped, and some studies presented a hypothesis that differences in core body temperatures in animals result in diverse uptake of MC, as well as different toxic effects. Thus far, however, few in vivo studies have been conducted to investigate the effects of temperature on MC-induced hepatocyte apoptosis in fish, a typical poikilotherm. In the present study, zebrafish were treated with MC-LR, an MC metabolite, at three water temperatures (12, 22 and 32 °C), and evident differences in apoptotic profiles were observed. Damage to liver ultrastructures revealed temperature-dependent early-stage patterns of apoptosis. Flow-cytometric analysis indicated that hepatocyte apoptotic rates varied with a temperature-dependent effect. The transcription levels of some apoptosis-related genes were determined using quantitative real-time polymerase chain reaction, and significantly elevated gene expressions of P53, Bcl-2, Bax and caspase-3 were found in the 12 and 32 °C groups. Results of the present study indicate that different ambient temperatures can lead to various toxic effects of MCs on hepatic apoptosis in fish.

Effect of cyanobacteria on immune function of crucian carp (Carassius auratus) via chronic exposure in diet.

Cyanobacterial blooms caused by water eutrophication have become a worldwide problem. Microcystins (MCs) released during cyanobacterial blooms exert toxicity on fish. Up to now, immunotoxicity of MCs on fish has been rarely reported. The present study investigated immune response of crucian carp (Carassius auratus) to cyanobacteria via chronic exposure in diet. Fish were fed with diets containing 20% (low dose group) and 40% (high dose group) of cyanobacteria lyophilized powder. After exposure of 30 d, a batch of assays was determined for assessing immunotoxicity of MCs. The head kidney and spleen indexes significantly increased in high dose group. Blood nitroblue tetrazolium activity in high dose group was nearly twice as much as that in control group with no cyanobacteria additive. Marked haemorrhage and hyperemia were observed in kidney and spleen in high dose group. The edematous mitochondria, deformation of the nucleus and compaction of chromatin occurred in lymphocytes of head kidney and spleen in both cyanobacteria groups. Lysozyme activity showed an obvious increase in low dose group but a sharp decrease in high dose group. Significant increase of macrophage bactericidal activity was detected in low dose group. The present findings indicate that via chronic diet exposure of different cyanobacteria levels, fish exhibit various immune responses. Fish immunity tends to proceed toward the direction of immunostimulative response at low MCs concentrations but toward the trend of immunosuppressive answer at high MCs concentrations.

Erythrocyte damage of crucian carp (Carassius auratus) caused by microcystin-LR: in vitro study.

Fish suffer from anemia and hypovolemic hypotensive shock after in vivo exposure with microcystins.However, except for in vivo causes for anemia and hypotension, an in vitro study of fish erythrocytes exposed to MC is necessary. For a better understanding of hematology toxicity of MC, the main aim of the present study was to investigate the toxic effects of microcystin on fish erythrocytes in vitro. Crucian carp erythrocytes were incubated in vitro with microcystin-LR (MC-LR) at doses of 0, 1, 10, 100 and 1,000 nM.The level of lipid peroxidate significantly increased in MC-LR treatment groups. Glutathione decreased after exposure to MC-LR. The activities of antioxidative enzymes, including superoxide dismutase, catalase,glutathione peroxidase and glutathione-S-transferase,were significantly increased after exposure with MC-LR.The hemolysis was significantly increased, while the activities of acetylcholinesterase, Na?­K?-ATPase and Ca2?­Mg2?-ATPase were significantly decreased. In addition, pathological alterations in agglomerated and jagged erythrocytes were observed in blood smears. The findings indicate that damages to erythrocytes should also be responsible for anemia and hypotensive shock or even death.