Comparative susceptibilities and immune reactions of wild and cultured populations of Caspian trout Salmo trutta caspius to VHSV.

Caspian trout Salmo trutta caspius is an endangered subspecies of brown trout Salmo trutta which is native to the Caspian Sea. Restocking programmes have been established, but recent introduction of the rhabdovirus viral haemorrhagic septicaemia virus (VHSV) into Iranian rainbow trout farms connected to waterbodies supporting wild Caspian trout may represent an additional threat to the declining stock. The susceptibility of wild and cultured populations of this endemic subspecies was demonstrated by performing controlled VHSV infection experiments (both by bath and injection challenges). Subsequently, VHSV infection in exposed fish was confirmed (CPE and quantitative PCR), virus levels were measured, and regulation of immune genes in exposed fish was investigated with a focus on the genes encoding IL-8, IFNγ, TGFß, TNFα, SAA, C3-4, CD8α, IgM, MHC I, MHC II, iNOS and IGF-1. The presence of IgM-, CD8α- and MHC II-positive cells in host organs was visualized by immunohistochemistry. Both wild and cultured trout strains proved to be VHSV-susceptible following experimental challenge, but the mortality curves and associated regulation of immune-related genes differed between the 2 trout types. Implications of the results for future management of Caspian trout populations are discussed.

Expression of growth hormone gene during early development of Siberian sturgeon (Acipenserbaerii).

The mRNA expression of growth hormone (GH) gene in early development stages of Siberian sturgeon was investigated using RT-PCR method. Samples were collected from unfertilized eggs up to 50 days post hatched (dph) larvae in 11 different times. Ribosomal protein L6 (RPL6) transcripts were used as the internal standard during quantification of GH mRNA expression. The results showed that the GH mRNA could be observed in the eyed eggs and even at unfertilized eggs of Siberian sturgeon. The highest amounts of GH mRNA were found at 25 and 50 dph larvae, while the lowest levels were detected at 1 and 3 dph larvae stage. These findings suggest that, the GH mRNA play a key role during developmental stages of Siberian sturgeon.

Distribution and composition pattern of polycyclic aromatic hydrocarbons in different tissues of sturgeons collected from Iranian coastline of the Caspian Sea.

The levels of 16 polycyclic aromatic hydrocarbons (PAHs) were determined in the liver, kidney, gills and muscle tissues of Persian sturgeon (Acipenser persicus; n=16), and Stellate sturgeon (Acipenser stellatus; n=7) collected from coastal waters of the South Caspian Sea from March and April 2011. The distribution and composition pattern of PAHs in the different tissues of sturgeons, and the effects of lipid content in sturgeon tissues and the octanol-water partition coefficient (Kow) of PAHs congeners on them were analyzed. The levels of total PAHs in the various tissues of Persian sturgeon and Stellate sturgeon ranged from 2.095 to 6.587 and 1.942 to 6.206 µg g(-1)dw, respectively. Stellate sturgeon showed significantly higher levels of heavy PAHs (⩾ 4-rings) than Persian sturgeon. The analysis has revealed a high degree of differential accumulation of the studied PAHs in the tissues of the both species. Low molecular weight PAHs predominated in the sturgeons, accounting for 81.89% of the total PAHs. Among the sixteen tested PAHs, naphthalene was the most dominant congener, followed by phenanthrene and fluorene. The PAHs levels and distribution in the tissues of sturgeons are dependent on both the Kow of PAH congeners and the lipid content in these tissues. There was a significant positive relationship (r=0.868, p<0.005) between lipid content and PAHs levels. The statistically significant negative relationships (p<0.01) were found between log Kow and log-transformed PAHs levels for muscle tissues of both sturgeon species.

Persian sturgeon insulin-like growth factor I: molecular cloning and expression during various nutritional conditions.

The effects of different periods of starvation (1, 2, 3, and 4 weeks) and subsequent re-feeding (over a 4 week) on the compensatory growth performance and insulin-like growth factor I (IGF-I) mRNA expression in liver and white muscle were investigated in juvenile Persian sturgeon (Acipenser persicus). First, a fragment of 617 nucleotides coding for IGF-I was cloned from liver, which included an open reading frame of 486 nucleotides, encoding a 162 amino acid preproIGF-I. This is composed of a 45 aa for signal peptide, a 117 aa for the mature peptide comprising the B, C, A, and D domains, and a 47 aa for E domain. The mature Persian sturgeon IGF-I exhibits high sequence identities with other sturgeon species and teleost, ranging between 68 and 95 %. The pattern of IGF-I mRNA expression in the liver and white muscle was measured in response to different periods of starvation and subsequent re-feeding. Nutritional status influenced IGF-I mRNA expression pattern in both liver and muscle. IGF-I mRNA expression in the liver increased during starvation, before decreasing after re-feeding. Furthermore, white muscle IGF-I mRNA expression showed better responses to nutritional status and decreased following starvation and increased by re-feeding. However, changes in the expression of IGF-I mRNA were not significantly different between any of the treatments in both tissues. These data suggest that muscle and liver IGF-I mRNA expression do not have a regulatory role for somatic growth induced by compensatory growth in Persain sturgeon.

Induction of meiotic gynogenesis in ship sturgeon Acipenser nudiventris using UV-irradiated heterologous sperm.

Diploid gynogenesis was induced in ship sturgeon Acipenser nudiventris using UV-irradiated sperm from Siberian sturgeon Acipenser baerii. The optimal condition for the retention of the second polar body in ship sturgeon was determined to be 10 min after activation/fertilization in experiments. The temperature of cold shock and its duration were 2.5 °C and 30 min, respectively. A total of 30 gynogens of known parentage from experimental treatments were screened using microsatellite DNA analysis, and uniparental transmission in meiogens was confirmed. The results show that heterologous Siberian sturgeon sperm is applicable as UV-irradiated sperm for the induction of gynogenesis in ship sturgeon. This technique may recover the critically endangered sturgeon species that are becoming extinct.

Bioaccumulation of Cd, Pb and Zn in the edible and inedible tissues of three sturgeon species in the Iranian coastline of the Caspian Sea.

The accumulations of Cd, Pb and Zn were determined in edible and inedible tissues of Persian sturgeon (Acipenser persicus; n=27), stellate sturgeon (Acipenser stellatus; n=5) and beluga (Huso huso; n=4) collected from coastal waters of the South Caspian Sea from March to April 2011. Concentrations of metals evaluated in the caviar, muscle, liver, kidney, gills, ovary and heart of the three species of sturgeons have been assessed using the flame and graphite furnace atomic absorption spectrophotometer. Concentrations of Zn, Pb and Cd in the edible and inedible tissues were apparently different among the three species of sturgeons. Especially, beluga heart showed the highest concentrations of Zn and Pb and Cd in Persian sturgeon liver. The analyzed metals were found in the caviar and muscle samples of Persian sturgeon and likewise muscle samples two other sturgeon species at mean concentrations under the permissible limits proposed by MAFF (2000).

Bioaccumulation of Zn, Cu and Mn in the caviar and muscle of Persian sturgeon (Acipenser persicus) from the Caspian Sea, Iran.

Concentrations of Zn, Cu and Mn were examined in caviar and muscle of the Persian sturgeon (Acipenser persicus) collected from coastal waters of south Caspian Sea during March and April, 2011. Mean Zn, Cu and Mn concentrations in caviar samples were 21.48, 2.05 and 1.66 µg g(-1) wet weight basis, respectively. The mean Zn, Cu and Mn concentrations in muscle tissues were 7.49, 1.00 and 0.34 µg g(-1) wet weight basis, respectively. The mean concentrations of Zn and Cu in caviar and muscle samples were under the permissible limits proposed by the United Kingdom's Ministry of Agriculture, Fisheries and Food (2000).

Morphology and fine structure of Acipenser persicus (Acipenseridae, Chondrostei) spermatozoon: Inter-species comparison in Acipenseriformes.

This study describes morphology and fine structure of the Persian sturgeon (Acipenser persicus) (Acipenseridae, Chondrostei) spermatozoon. The results show that the spermatozoon of A. persicus is differentiated into an elongated head (length: mean±SD: 7.1±0.5µm) with an acrosome (length: 1.2±0.2µm), a cylindrical midpiece (length: 1.8±0.5µm), a flagellum (length: 50.3±5.9µm) and a total length of 59.2±6.2µm. Ten posterolateral projections (PLPs) arise from the posterior edge of the acrosome and there were 3 endonuclear canals that traversed the nucleus from the acrosomal end to the basal nuclear fossa region. Three to six mitochondria were in peripheral midpiece and the proximal and distal centrioles were located near to "implantation fossa" and basement of the flagellum. The axoneme has a typical eukaryotic structure composed of 9 peripheral microtubules and a central pair of single microtubule surrounded by the plasma membrane. Lateral fins were observed along the flagellum. The fins started and ended at 0.5-1µm from midpiece and at 4-6µm from the end of flagellum. There were significant differences in the size of almost all measured morphological parameters between males and flagellar, midpiece and nucleus characters were more isolated parameters that can be considered for detecting inter-individual variations. This study showed that sperm morphology and fine structure are similar among sturgeon species, but the dimensions of the parameters may differ.

Metallothionein as potential biomarker of cadmium exposure in Persian sturgeon (Acipenser persicus).

Metallothionein (MT) concentration in gills, liver, and kidney tissues of Persian sturgeon (Acipenser persicus) were determined following exposure to sublethal levels of waterborne cadmium (Cd) (50, 400, and 1,000 µg l(-1)) after 1, 2, 4, and 14 days. The increases of MT from background levels were 4.6-, 3-, and 2.8-fold for kidney, liver, and gills, respectively. The results showed that MT level change in the kidney is time and concentration dependent. Also, cortisol measurement revealed elevation at the day 1 of exposure and followed by MT increase in the liver. Cd concentrations in the cytosol of experimental tissues were measured, and the results indicated that Cd levels in the cytosol of liver, kidney, and gills increased 240.71-, 32.05-, and 40.16-fold, respectively, 14 days after exposure to 1,000 µg l(-1) Cd. The accumulation of Cd in cytosol of tissues is in the order of liver > gills > kidney. Pearson correlation coefficients showed that the MT content in kidney is correlated with Cd concentration, the value of which is more than in liver and gills. Thus, kidney can be considered as a tissue indicator in A. persicus for waterborne Cd contamination.

Effects of triploidy on the Caspian salmon Salmo trutta caspius haematology.

The aim of this study was a comparison of key haematological features of diploid (2n) and triploid (3n) Caspian salmon (Salmo trutta caspius). Morphometric indices of erythrocytes were determined on blood smears by light microscopy. Triploidy significantly (P < 0.001) increased all morphometric indices measured in the erythrocytes including cell size, cell surface area, and cell volume. The increase in cell size was larger for the major (27%) axis than for the minor (22%) axis, thus making erythrocytes of 3n Caspian salmon more ellipsoidal. The estimated increase in erythrocyte nuclear volume (87%) was bigger than the theoretical expected 50% increase. Haematological indices were measured manually by hemocytometry. Triploids had lower numbers of red blood cells (RBC: 1,120,000 cells/mL in 2n vs. 700,000 cells/mL in 3n; P < 0.001) but they were larger in size (mean erythrocytic volume [MEV]: 363.1 nm3 in 2n vs. 483.3 nm3 in 3n; P < 0.001). The decrease in RBC number was not compensated by the increase in MEV and, thus, triploidy affected the haematocrit (Hct: 38.8% in 2n vs. 33.06% in 3n; P < 0.05). Total blood hemoglobin concentration was lower in triploid fish (Hb: 9.9 g/dL in 2n vs. 8.9 g/dL in 3n; P < 0.05). In contrast, mean erythrocytic hemoglobin (MEH: 95 mug in 2n vs. 133.2 mug in 3n; P < 0.001) was higher for 3n Caspian salmon as a result of their larger erythrocytes, although MEH concentration (MEHC: 0.26 g/dL in 2n vs. 0.27 g/dL in 3n) did not significantly differ (P > 0.05). White blood cell (WBC) counts (lymphocytes and neutrophiles) were measured and WBC/RBC ratios were calculated. There were no significant differences in WBC (15,710 cells/mL in 2n vs. 12,683 cells/mL in 3n; P > 0.05), lymphocytes, and neutrophils as %WBC as well as WBC/RBC ratios between two ploidy levels (P > 0.05). Triploid Caspian salmon showed higher erythrocyte abnormalities such as 'twisted', 'tailed', and 'anucleated' cells as well as high portions of immature RBC in blood smears in comparison with diploids (P < 0.001).

Concentrations of trace elements in muscle of sturgeons in the Caspian Sea.

Concentrations of 21 trace elements were determined in muscle of beluga (Huso huso), Persian sturgeon (Acipenser persicus), Russian sturgeon (Acipenser gueldenstaedtii), ship sturgeon (Acipenser nudiventris) and stellate sturgeon (Acipenser stellatus) collected from coastal regions of Azerbaijan, Iran, Kazakhstan and Turkmenistan, surrounding the Caspian Sea during 2000-2001. Concentrations of Mn, Co, Cu, Mo, Sn, Hg, Pb and Bi in the muscle were apparently different among the five species of sturgeons. Especially, beluga showed the highest concentrations of Hg, Pb and Mn in all the five species. In addition, more than half of the individuals of beluga exceeded the guideline level (0.3 microg/g wet wt.) of Hg for food in UK. However, V, Cr, Zn, Ga, Rb, Sr and Ba concentrations were similar among five sturgeons. Growth-dependent increase in Hg in beluga, Cu in Persian sturgeon, and Zn in Russian sturgeon were observed, whereas V, Mn, Co, Cu, Ga, Sr, Mo, Sn, Hg and Bi concentrations in Persian sturgeon, Pb in ship sturgeon, and Cr, Mn, and Rb in stellate sturgeon were negatively correlated with body length. All species of sturgeon in Azerbaijan showed the highest concentration of Sr and the lowest concentration of Rb, while the five sturgeons from Iran showed opposite trends. Concentration of V, which is present in oil, in sturgeons in the Caspian Sea was considerably lower than that of fish from Kuwait, but was comparable to that of Cambodia and the Gulf and Gulf of Oman. To our knowledge, this study provides the first extensive data on multielemental accumulation in sturgeons of the Caspian Sea.

Contamination by organochlorine compounds in sturgeons from Caspian Sea during 2001 and 2002.

Organochlorine compounds (OCs) were determined in the five species of sturgeons collected from coastal waters of Caspian Sea in Kazakhstan, Azerbaijan, Turkmenistan and Iran during 2001 and 2002 to understand their status of contamination and accumulation features. Among OCs examined, concentrations of DDTs (DDT and its metabolites) were predominant in all the sturgeon samples with concentrations ranging from 73 to 31,000 ng/g on lipid weight basis, followed by PCBs, CHLs, HCHs, HCB, dieldrin, TCPMOH, and heptachlor epoxide in order. The concentrations of OCs in beluga (Huso huso) were the highest among all the five species. When comparing residue levels among same species, OC residues were highest in sturgeons from Azerbaijan and Kazakhstan and lowest in Turkmenistan. However, the concentrations of HCHs, dieldrin, and heptachlor epoxide in sturgeons showed less geographical variability.