Gill alterations of whiteleg shrimp (Litopenaeus vannamei) exposed to cadmium under different experimental salinities.

BACKGROUND: Heavy metals are one of the most important environmental pollutants in marine coastal ecosystems. Cadmium is a heavy metal that enters to marine environments via industrial wastes and oil production activities. OBJECTIVES: This study were done to determine the toxicity of cadmium to Litopenaeus vannamei and to evaluate the histological changes in gill tissues after exposure to sublethal concentrations of cadmium at different salinities. METHODS: For this reason, toxicity test was done to determine the lethal concentration (LC50) of cadmium for whiteleg shrimp. According to the calculated LC50 amount, sublethal doses of cadmium were used to determine its histological effects in different salinity during 2 weeks exposing period. RESULTS: LC50 of cadmium for 96 h for whiteleg shrimp was 6.56 mg/L. Histological alterations in the gill were observed in L. vannamei after 14 days exposure to different concentrations of cadmium and salinity. Histopathological index was increased in a dose-dependent manner. CONCLUSION: Our findings showed that doses lower than 2 mg/L have repairable effects on gill structure, but the concentration of 2 mg/L cadmium leaves irreparable and destructive effects on the gill tissue.

Phenanthrene disrupting effects on the thyroid system of Arabian seabream, Acanthopagrus arabicus: In situ and in vivo study.

Phenanthrene, a polycyclic aromatic hydrocarbon (PAH), is one of the endocrine disrupting chemicals (EDCs). The present study aimed to evaluate the effects of phenanthrene on histophysiology of thyroid in Arabian seabream (Acanthopagrus arabicus). In this regards, different concentrations of phenanthrene (2, 20 and 40 pg/gbw) were injected to Acanthopagrus arabicus and changes in thyroid tissue structure and the serum levels of triiodothyronine (T3) and Thyroxine (T4) were assessed. The experiment lasted 21 days. Alterations in thyroid tissue structure and T3 and T4 serum levels also were assessed in Acanthopagrus arabicus caught from different stations of the Persian Gulf (Jafari, Samail, Arvand, Zangi, Bahrakan). In addition, the concentration of phenanthrene was measured in the fish muscle and sediment samples from the stations. Phenanthrene concentration reached the maximum level in the muscle of all injected fish after 4 days and then decreased by the end of the experiment. The highest and lowest concentrations of phenanthrene were recorded in the fish muscle and sediment samples collected from Jafari and Bahrakan, respectively. The levels of T3 and T4 decreased dose dependently in phenanthrene-injected fish up to day 7 and then increased by the end of the experiment. The serum level of T3 and T4 in fish collected from different stations was as follows: Jafari

Interactions of oxidative DNA damage and CYP1A gene expression with the liver enzymes in Klunzinger's mullet exposed to benzo[a]pyrene.

Benzo[a]pyrene (B[a]P) is an important contaminant whose liver biotransformation is dependent on the species, the route of exposure and the concentration. The goal of this study was to assess the interactions of oxidative DNA damage and CYP1A gene expression with the liver enzymes in Klunzinger's mullet (Liza klunzingeri) exposed to benzo[a]pyrene. Sublethal doses of B[a]P (5, 10 and 50 mg/kg) were intraperitoneally administered to the fish for 14 days. The alterations in antioxidant enzymes' activity (SOD, CAT, and GPX), hepatic enzymes' activity (ALT, AST and ALP), DNA damage (measured by comet assay and cellProfiler software) and CYP1A gene expression in the fish liver were studied on the 1st, 3rd, 7th and 14th days. The determination of these parameters in the liver showed that most of these parameters significantly increased mostly in a time-dependent manner. Multiple regression analysis showed that DNA damage and CYP1A gene expression had positive correlations with the liver enzymes in this fish species intraperitoneally exposed to these concentrations. Moreover, these interactions indicated that theses parameters are sensitive biomarkers for the exposure to B[a]P in Klunzinger's mullet. However, other possible factors and B[a]P metabolites should be considered in future studies for better elucidating the biotransformation mechanisms and introducing better biomarkers of B[a]P.

Short-term induction of vitellogenesis in the immature male yellowfin seabream ( Acanthopagrus latus) exposed to bisphenol A and 17 ß-estradiol.

Bisphenol A (BPA) is a known environmental endocrine-disrupting chemical that is widely used in plastics manufacturing. BPA enters in the aquatic environment mainly through urban and industrial sewage effluents, thereby posing a potential threat to organisms living in these ecosystems. This study was conducted to investigate the effect of BPA on VTG production with direct (sodium dodecyl sulfate-polyarylamide gel electrophoresis) and indirect (alkali-labile phosphate (ALP), total plasma calcium and protein) methods in immature male yellowfin seabream ( Acanthopagrus latus) as a marine fish model. Fish were randomly distributed into seven groups that were administered 1, 10, 50, and 100 µg g-1 week-1 of BPA and 2 µg g-1week-1 of 17ß-estradiol (E2) over a period of 2 weeks. Solvent controls received olive oil, whereas controls were not injected. The fish were sampled on days 0, 7, and 14, and their blood plasma and liver were obtained. The results showed that the hepatosomatic index of all treated fish was elevated in comparison with controls. Direct and indirect indicators showed that fish VTG protein was induced by BPA and E2 exposure. The protein was found to have two bands with molecular weights around 210 and 190 KDa. ALP, total plasma calcium and protein levels were increased in dose- and time-dependent manners. The results of this study demonstrated that short-term exposure of yellowfin seabream to BPA induced adverse effects in the reproductive system of hermaphrodite fish.

Detection of benzo[a]pyrene-induced immunotoxicity in orange spotted grouper (Epinephelus coioides).

This study aimed to investigate the effects of benzo[a]pyrene (BaP) on immune status of orange spotted grouper (Epinephelus coioides). Fish were injected with 2, 20 and 35 mg/kg-bw of BaP and were kept under laboratory conditions for 14 days. Blood samples were taken at days 1, 4, 7, and 14 and changes in total WBC and RBC, phagocytosis, lysozyme activity, lysosomal membrane stability, immunoglobulin M (IgM) level and antibacterial activity were evaluated. Also BaP bioaccumulation in fish muscle was measured. BaP concentration in the muscle of treated fish reached a maximum level after 4 days (P < 0.05). Exposure of fish to BaP resulted in a significant decrease of total RBC and WBC, lysozyme activity, lysosomal membrane stability, IgM level and antibacterial activity after 4 days and phagocytosis after 7 days of the experiment (P < 0.05). Totally, the results revealed BaP ability to suppress the fish immune function.

Application of biomarkers in Epaulet grouper (Epinephelus stoliczkae) to assess chromium pollution in the Chabahar Bay and Gulf of Oman.

In the present study plasma levels of erythropoietin (EPO) hormone and kidney histological changes were assessed as biomarkers to evaluate the chromium pollution in Epaulet grouper (Epinephelus stoliczkae) obtained from different areas in Chabahar Bay and Gulf of Oman. Chemical analyses included the evaluation of chromium in the fish kidney tissue and sediment. The highest and lowest concentrations of chromium were measured in the kidney of fish and sediment collected from Konarak (47.53±1.23 and 110±1.4 µg g(-1)) and the mouth of Chabahar Bay (3.43±1.31 and 13.5±3.23 µg g(-1)), respectively. There was significant difference (P<0.05) of EPO plasma levels between Konarak, Beris and Ramin stations with the mouth of Chabahar Bay as the cleanest station. Reduction of Bowman's space, occlusion of the tubule lumen, leukocytes infiltration, melanomacrophage centers aggregation, nuclear vacuolation, nuclei hypertrophy of tubule cells, necrosis of the tubules, lifting of the tubular basement membrane and dilatation of glomerular capillaries were the most alterations observed in the kidney. The HAI values of Epaulet grouper from Konarak, Beris and Ramin stations were significantly higher than other stations (P<0.05). It was concluded that selected biomarkers as a first investigation could be useful tools to environmental biomonitoring programs.

Induction of vitellogenin synthesis in immature male yellowfin seabream (Acanthopagrus latus) exposed to 4-nonylphenol and 17ß-estradiol.

In the present study, the effects of xenoestrogen 4-nonylphenol (4-NP) and 17ß-estradiol (E2) on the synthesis of vitellogenin (VTG) in immature male yellowfin seabream (Acanthoparus latus) were assessed to determine the potential use of this species as a biological model for studying endocrine disrupters in the Musa estuary in southern seawaters of Iran. A total number of 104 fish were injected by 10, 50, 100, 150 and 200 µg g(-) (1) week(-) (1) of 4-NP and 2 µg g(-) (1) week(-) (1) of E2, and their blood plasma was obtained for biochemical analysis on days 0, 7 and 14. The hepatosomatic index (HSI) of each fish was also recorded at each sampling time. The results showed that 4-NP significantly induced VTG synthesis in A. latus, which was detected by gel electrophoresis as two major protein bands (∼210 and 191 KDa). The induction of VTG was also determined indirectly by measuring the alkali-labile phosphate, total plasma calcium and protein. All of these VTG indicators significantly increased in the plasma of 4-NP- and E2-treated fish. Meanwhile, a significant dose-dependent increase was observed in HSI, indicating the activation of hepatic VTG production in 4-NP- and E2-injected fish. Furthermore, the plasma alanine aminotransferase and aspartate aminotransferase concentrations significantly increased in 4-NP-treated fish, presumably reflecting a situation of hepatic tissue damage due to 4-NP toxicity. Based on the results, it is suggested that A. latus could be used successfully as a model species for future studies of endocrine disrupting contaminants in the Musa estuary.

An assessment of hematological and biochemical responses in the tropical fish Epinephelus stoliczkae of Chabahar Bay and Gulf of Oman under chromium exposure: ecological and experimental tests.

The aim of this investigation was to evaluate the effect of chromium on hematological and biochemical parameters in Epaulet Grouper, Epinephelus stoliczkae of Chabahar Bay and Gulf of Oman by ecological and experimental tests. Spatial evaluation of ecological test results showed these parameters had significant difference among some sampling sites. Examination of hematological and biochemical profiles on Epaulet Grouper was performed after 0.5, 1, 7, 14, and 21 days of chromium exposure (3.6, 7.31 and 14.6 mg/L). Experimental test results of chromium induce indicated the significant decrease in MCV, MCH, neutrophils, basophils, plasma protein and significant increase in MCHC, lymphocytes, monocytes, eosinophils and a biphasic trend in Hb, Ht, RBC, WBC, and glucose (p < 0.05). Cellular and nuclear axis, cytoplasmic volume, cell and nuclear volume, and surface area were significantly different for ecological and experimental results (p < 0.05). It was concluded that these parameters are sensitive in monitoring the toxicity of chromium concentrations.

Effects of 4-nonylphenol on balance of steroid and thyroid hormones in sexually immature male yellowfin seabream (Acanthopagrus latus).

Nonylphenol (NP) is an endocrine disrupting chemical which has been shown to be able to modulate the endocrine system of various organisms by different mechanisms. The objective of this study was to investigate the potential effects of 4-NP on steroid and thyroid hormone levels in sexually immature male yellowfin seabream (Acanthopagrus latus), a protandrous hermaphrodite species. For this, the fish were injected with ascending doses (10, 50, 100, and 200 µg g(-1) body weight) of 4-nonylphenol (4-NP) or vehicle during 2 weeks. After 7 and 14 days the fish were anesthetized, blood sample were collected and plasma steroid and thyroid hormone concentrations were quantified by radioimmunoassay. The result showed that 4-NP induced a significant increase in 17ß-etradiol levels at dose 10 µg g(-1) , while the levels of this hormone in the higher doses decreased compared with the control group. However, 4-NP treatment did not have any significant effect on plasma levels of testosterone. In addition, it was observed that 4-NP affect the level of thyroid hormones in fish. Plasma thyroxine levels increased in a dose-dependent manner after 7 and 14 days of the exposure. In contrast, a significant decrease in triiodothyronine levels was observed during the experiment period. Moreover, no significant change was detected for thyroid stimulating hormone levels in 4-NP-treated fish. These results indicated that 4-NP could lead to disturb the balance of steroid and thyroid hormones with potential consequences for sexually immature male yellowfin seabream.

Assessment of gill pathological responses in the tropical fish yellowfin seabream of Persian Gulf under mercury exposure.

Gill histomorphological alterations were used to assess the effects of chronic exposure to HgCl2 on the yellowfin seabream, Acanthopagrus latus. In this regard, 90 A. latus were exposed to sublethal concentrations of HgCl2 (10, 20, 35 and 50 µg/L) for 3 weeks. Treated fish were erratic and showed respiratory distress. The most common morphological abnormalities included: filaments disorganization, increase of mucus secretion, debris and blood plaques on the filaments, losing or shortening of some filaments. The most frequent histopathological changes detected in the gills included extensive lifting of the lamellar epithelium and edema of lamellae with enlarged sub-epithelial spaces, exfoliated epithelium of lamellae, telangiectasia, hypertrophy and hyperplasia of the epithelial cell resulted in partial fusion of the secondary lamellae and a reduction of the water space, club shaping of gill lamellae, blood congestion. Some more severe alternations found in the gill of fish exposed to higher levels of HgCl2 (35 and 50 µg/L) included lamellar aneurysm and hemorrhages with rupture of the lamellar epithelium. According to the results of the present study, mercuric chloride could cause major histomorphological changes in the gill of A. latus, decreasing its gas exchange capability. Two mercury concentrations (10 and 20 µg/L) used in the present study were in agreement with the concentration of mercury in the water of different parts of Mahshahr creeks (the north of Persian Gulf) (3.66 to 15 µg/L). Therefore, based on the results the presence of pathological alteration in A. latus inhibited in the natural environment (Mahshahr creeks) seems to be logical.

The relationship between heavy metal (Cd, Co, Cu, Ni and Pb) levels and the size of benthic, benthopelagic and pelagic fish species, Persian Gulf.

The concentration of heavy metals was determined in tissues of sole (Euryglossa orientalis), mullet (Liza abu) and croacker fish (Johnius belangerii) from Musa estuary. Generally, the highest concentration of the studied metals in the three species was found in the liver tissue. The levels of Cd and Cu in fish liver were J. belangerii = E. orientalis > L. abu and E. orientalis > L. abu > J. belangerii respectively. The concentrations of Cd and Cu in fish gills were E. orientalis > L. abu = J. belangerii and E. orientalis > L. abu > J. belangerii, respectively, and the level of Cu in muscle was E. orientalis > L. abu = J. belangerii. The results of linear regression analysis indicated that highly significant (p < 0.001) negative relationships were found between fish size and Cd concentrations in the liver of L. abu and Pb in the gills of J. belangerii.

Heavy metal concentration in sediment, benthic, benthopelagic, and pelagic fish species from Musa Estuary (Persian Gulf).

The concentration of Cd, Co, Cu, Ni, and Pb was measured in sediment and three fish species collected from Musa Estuary, Persian Gulf. The concentration order of heavy metals in sediment were Ni > Co > Cu > Pb > Cd >. Concentrations of the heavy metals in the fish were apparently different among the three species. The concentrations of Cd and Ni in fish were Johnius belangerii > Euryglossa orientalis > Liza abu, while the Co and Cu levels were L. abu > E. orientalis > J. belangerii and E. orientalis > L. abu > J. belangerii, respectively. Result of regression analysis showed that there were no significant relationships between metal concentration in fish tissues and sediment, except for Ni concentration in the J. belangerii liver. The concentrations of studied metals in fish muscle were below the permissible limits proposed by FAO, WHO, and EC.

Using histopathological changes as a biomarker to trace contamination loading of Musa Creeks (Persian Gulf).

Gill histological changes were used as a biomarker to evaluate the health of yellowfin seabream, Acanthopagrus latus (A. latus), collected from different sites in Musa Creeks including: (A) Petrochemical, (B) Jaafari, (C) Ghazaleh, (D) Majidieh, and (E) Zangi creeks. Also Genaveh with relatively clean water was selected as reference. The gills of 60 A. latus were fixed in Bouin's solution for 12 hr, dehydrated in a graded series of ethanol, and embedded in paraffin. Five micrometer thick sections were stained in H&E and Periodic Acid Schiff for microscopic examination. Also, the presence of gill histological alterations was assessed by the histopathologic alteration index (HAI). The gill morphological abnormalities include mucus secretion increase, debris, blood plaque, and shortening of filaments. The gill pathological changes included lamellar cells hyperplasia, aneurysm, lamellar fusion, and epithelial lifting. The HAI means were varied from moderate to severe in sites A and B, thus these sites are considered as being of low quality. Some severe pathological alterations were observed in site D, but their distribution was lower than sites A and B. The least HAI means of sites C and E demonstrated their good environmental quality. The results suggest that there is close relation between amounts of pathological alterations and environmental contamination.

Effect of sublethal dose of mercury toxicity on liver cells and tissue of yellowfin seabream.

The aim of the present study was to provide baseline data on the prevalence of histopathological liver lesions in Acanthopagrus latus under experimental mercury exposure. Experimental study was at seawater recirculatory tanks. Mercury concentrations were determined using a standard cold vapour atomic absorption. Histopathological analyses were done in tissue processor and the slides were stained with haematoxylin and counterstained with eosin. There were many liver lesions including enlarged and lateral nuclei, nuclear degeneration and vacuolation; oncotic, apoptic, focal, massive, centrilobular and periportal necrosis; atrophy, lipidosis, hydropic and cloudy swelling, oval cell proliferation; bile stagnation, dilation of sinusoid, intracellular oedema and dark granules in both field and laboratory conditions. In conclusion the present investigation indicated that mercury is a toxic substance in yellowfin seabream and the sublethal concentrations of mercury may cause several changes in the histological indices of the studied fish, and we can use these changes as biomarkers of mercury detection.

Serum hormone and biochemical activity as biomarkers of mercury toxicity in the yellowfin seabream Acanthopagrus latus.

This study was undertaken in two separate environmental and experimental conditions. The environmental test was carried out at creeks in the Persian Gulf and the experimental test was undertaken in seawater re-circulatory tanks. Mercury concentrations were determined using a standard cold vapour atomic absorption method. Serum enzymes were determined with Diagnostics Infinity reagent kit. Serum glucose and total protein were measured photometrically. Serum hormones were assayed using diagnostic ELISA kits. Results of the present investigation indicated that the sub-acute and chronic mercury concentrations tested may cause several changes in the biochemical and hormone parameters of the studied fish and we can use these changes as biomarkers for mercury detection. The range of mercury concentrations found in the creek water and specially sediments along the Mahshahr coast was higher than the other marine environment. It was declared that between enzymatic indices serum acid phosphatase (ACP) amount and ACP activity, between hormonal indices testosterone, T3 and T3/T4 and between metabolite index total protein can be considered as suitable and effective biomarkers of mercury pollution in yellowfin seabream.

Characterization of a chitinase with antifungal activity from a native Serratia marcescens B4A

Chitinases have the ability of chitin digestion that constitutes a main compound of the cell wall in many of the phytopathogens such as fungi. In the following investigation, a novel chitinase with antifungal activity was characterized from a native Serratia marcescens B4A. Partially purified enzyme had an apparent molecular mass of 54 kDa. It indicated an optimum activity in pH 5 at 45ºC. Enzyme was stable in 55ºC for 20 min and at a pH range of 3-9 for 90 min at 25ºC. When the temperature was raised to 60ºC, it might affect the structure of enzymes lead to reduction of chitinase activity. Moreover, the Km and Vmax values for chitin were 8.3 mg/ml and 2.4 mmol/min, respectively. Additionally, the effect of some cations and chemical compounds were found to stimulate the chitinase activity. In addition, Iodoacetamide and Idoacetic acid did not inhibit enzyme activity, indicating that cysteine residues are not part of the catalytic site of chitinase. Finally, chitinase activity was further monitored by scanning electronic microscopy data in which progressive changes in chitin porosity appeared upon treatment with chitinase. This enzyme exhibited antifungal activity against Rhizoctonia solani, Bipolaris sp, Alternaria raphani, Alternaria brassicicola, revealing a potential application for the industry with potentially exploitable significance. Fungal chitin shows some special features, in particular with respect to chemical structure. Difference in chitinolytic ability must result from the subsite structure in the enzyme binding cleft. This implies that why the enzyme didn't have significant antifungal activity against other Fungi.

Characterization of a chitinase with antifungal activity from a native Serratia marcescens B4A.

Chitinases have the ability of chitin digestion that constitutes a main compound of the cell wall in many of the phytopathogens such as fungi. In the following investigation, a novel chitinase with antifungal activity was characterized from a native Serratia marcescens B4A. Partially purified enzyme had an apparent molecular mass of 54 kDa. It indicated an optimum activity in pH 5 at 45°C. Enzyme was stable in 55°C for 20 min and at a pH range of 3-9 for 90 min at 25°C. When the temperature was raised to 60°C, it might affect the structure of enzymes lead to reduction of chitinase activity. Moreover, the Km and Vmax values for chitin were 8.3 mg/ml and 2.4 mmol/min, respectively. Additionally, the effect of some cations and chemical compounds were found to stimulate the chitinase activity. In addition, Iodoacetamide and Idoacetic acid did not inhibit enzyme activity, indicating that cysteine residues are not part of the catalytic site of chitinase. Finally, chitinase activity was further monitored by scanning electronic microscopy data in which progressive changes in chitin porosity appeared upon treatment with chitinase. This enzyme exhibited antifungal activity against Rhizoctonia solani, Bipolaris sp, Alternaria raphani, Alternaria brassicicola, revealing a potential application for the industry with potentially exploitable significance. Fungal chitin shows some special features, in particular with respect to chemical structure. Difference in chitinolytic ability must result from the subsite structure in the enzyme binding cleft. This implies that why the enzyme didn't have significant antifungal activity against other Fungi.