Characterization of Heavy-Metal-Removal Bacteria From The Persian Gulf

The study was carried out to isolate and screen high heavy metals resistant bacteria from Persian Gulf and enclosed industrial areas within 241,000 km2. A total of 35 heavy metal resistant bacteria strains were identified from sediment and water samples collected. The resistance and biological capacity of the isolated bacteria were tested in a new formulated media, minimal salt solution (M.S.S), that mimics seawater. Isolated bacteria responded to media supplemented in range 0.5 to 2 mM of Zn, Cd, Cu and Pb by showing a prolonged lag phase and by decreasing growth rate. Bacteria isolates, in the form of free or immobilized cells, are able to remove lead, copper, zinc and cadmium from solution. Removal of lead and cadmium from solution by some bacteria was very efficient, about 120 mglg dry weight as high as 90%. Isolates tested presented distinct uptake capacities and the best results were obtained for Delftia tsuruhatensis and Pseudornonas AU3411 respectively.The diversity of isolated bacteria was examined by the phylogenetic analysis of 16s rRNA gene sequences. The phylogenetic analyses of the sequences revealed seven main taxonomic lineages. The phylogenetic tree illustrated discrimination between isolated bacteria from wastewater, industrials areas and marine environment. Some Pseudornonas strains isolated from marine environment were well differentiated from those of industrial wastewater. Members of the genera Delftia and Bacterium formed a monophyletic group within the subdivision of the class. There was a clear differentiation between two groups of Pseudornonas and other groups of bacteria in the phylogenetic tree. The isolated bacteria were tested for the occurrence of plasmid using the modified alkaline lysate method. The study revealed that the frequency of the occurrence of plasmid in the heavy metals resistance bacteria was more than in the common bacteria. Multiple forms of plasmids were observed in 66% of the plasmidcarrying strains. Isolates bacteria from industrials wastewater showed the highest plasmid incidence (84.6%). In the marine environment there was a slightly higher incidence of plasmid in bacteria isolated from sediments (55.5%) compared to the water sample of the same origin (53.8%). Scanning Electron Microscope (SEM) analyses showed Pseudornonas sp. accumulated heavy metals in the cell wall and along the external cell surfaces. This suggested that heavy metals uptake involves both surface phenomena and diffusion. Energy Dispersive X-ray (EDX) analysis confirmed heavy metals on the bacteria cell surface which was reported by SEM

Evaluation of antioxidant activity of the purified peptides from hydrolysis of rotifer (Brachionus plicatilis)

Rotifers are an important group of zooplankton in aquatic ecosystems that contain relatively high amounts of unsaturated fatty acids, proteins and peptides. Rotifers, especially Brachionus plicatilis species, are one of the important live food sources for marine fish larvae in aquaculture. The aim of this study was to evaluate the antioxidant activities of purified peptides from B. plicatilis. Antioxidant peptides of the B. plicatilis have been hydrolyzed by Alcalase, α¬Chymotrypsin, Papain, Neutrase, Pepsin, and Trypsin. Their antioxidant activity was evaluated by the free radical inhibitory effect of diphenylpicrylhydrazyl (DPPH). Also the sequential chromatography method was used for extraction and purification of the peptides. The results showed that peptides obtained from the pepsin hydrolysate have a higher inhibitory effect than other peptides. Pepsin showed 58% inhibitory effect in 0.015 mM DPPH. Finally, the pepsin hydrolysate constitutes were purified and isolated by gel-filtration chromatography (Sephadex G-250) and reverse-phase liquid chromatography on Eurospher C18 column (250×4.6 mm), respectively. The results of this study have been recognized the high antioxidant activities of extracted hydrolysate from B. plicatilis and their feasibility of using them in food industries as a food complement

Searching the genome of beluga (Huso huso) for sex markers based on targeted bulked segregant analysis (BSA)

In sturgeon aquaculture, where the main purpose is caviar production, a reliable method is needed to separate fish according to gender. Currently, due to the lack of external sexual dimorphism, the fish are sexed by an invasive surgical examination of the gonads. Development of a non-invasive procedure for sexing fish based on genetic markers is of special interest. In the present study we employed Bulked Segregant Analysis (BSA) methodology to search for DNA markers associated with the sex of the beluga sturgeon (Huso huso). DNA bulks (male and female) were created by combining equal amounts of genomic DNA from 10 fish of both sexes. A total of 101 decamer primers associated with the sex-specific sequences in non-sturgeon species was used for targeted screening of the bulks, resulting in 2846 bands that all of them were present in both sexes. Our results showed that sex chromosomes are weakly differentiated in the sturgeon genome and comprised sequences not complementary to the sex-specific primers in non-sturgeon species

Effects of different salinity on number and area of chloride cells in gill of juvenile grouper (Epinephelus coioides)

The purpose of this study was to evaluate the ability of osmoregulation by mitochondria-rich cells in grouper fish with high tolerance of salinity variation levels. For this purpose, groupers were transferred from salt water with an average salinity of 40 ppt to the waters with 10 ppt, 20 ppt and 60 ppt rates of salinity. Changes in the number and area of chloride cells in the different treatments were observed during two months adjustment period. Experiment was carried out by sampling three fish from each tank within 8 phases at the moment of the transition, 12 hours, day 1, day 3, day 7, day 14, day 30 and day 60. Histomorphological and immunohistochemical observations were done after fixing in Bouin's solution for 24 hours. Samples were dehydrated with increasing series of ethanol, followed by paraffin, and cleared by xylene. Paraffin blocks were cut at 5 microns and stained by hematoxylin – eosin. Changes in the number and area of chloride cells in 5% level were very different at high and brackish salinity. So in the early hours of the transfer, number and area of chloride cells were low in the brackish water than to that in high salinity. From second week towards the end of period, number of chloride cells in 60 ppt was more than that in control treatment from seventh days towards the end of period, the area of chloride cells was more than that in control group. The high ability and adaptability of this species in response to different environmental salinities might be associated with changes in gill histological evaluation and model osmoregulation mechanisms at different salinities

Molecular identification and phylogeny of Nerita᾽s species on rocky shores of the Northern Persian Gulf

Neritas are among the most dominant groups of Gastropoda in the Persian Gulf. There is no previous study in relation to molecular and phylogeny of Neritas in the study area. The molecular identification of Nerita species have been studied for the first time in the northern rocky coastal zones of Persian Gulf during 2013 and 2014. After morphological identifications of species, DNA extraction, amplifying partial of cytochrome oxidase COI and 16S rRNA and sequencing procedure were done in the laboratory. In this study, 6 COI and 6 16S rRNA sequences, belonging to 3 species, were obtained. Also, phylogeny analyses with drawing phylogeny trees of Maximum Likelihood and Bayesian were done using MEGA6 and BEAST softwares. Morphological and molecular identification results were similar for 2 species and dissimilar for one species. This discrepancy shows that a combination of morphological and molecular studies is more reliable for species identification than either of them solely. The results also showed that Nerita species are monophyletic

Study of genetic diversity of hara trees on the coast of Hormozgan Province using rapid molecular method

In order to study the genetic diversity of Hara trees in the coastal waters of Hormozgan province, using 40 RAPD molecular samples, 40 samples of Hara leaves were collected at different stations (ports of Jask, Dumitry, Tiba and Qeshm Island) and transferred to the laboratory. After extraction of DNA They were examined by 30 primers in polymerase chain reaction. The results showed that the highest mean genetic variation in Qeshm population was 0 458 with a standard deviation of 0.033 and the lowest mean of genetic diversity in Jask population was 0.423 with a standard deviation of 0.056. Also, using the PopGene software, the average Shannon information index was 0.643 for four populations. The value of Fst index was 011/0 and the gene flow rate was 25.854, and the UPGMA dendrogram, based on the Nei genetic distance, did not show a clear separation between the populations. Also, molecular variance analysis showed that there is a high variation (99%) within the studied populations. The results of this research can provide useful information for conservation and management programs of this kind with indigenous value of Iran

Morphological and molecular identification of brown algae, Padina sp. in Lengeh Port, Persian Gulf

Brown algae, Padina sp., are found across tropical coastal areas as well as inter-tidal and sub-tidal regions. The aim of this research was to identify the morphological and molecular characteristics of the Padina species distributed across Port Lengeh in the Persian Gulf using morphological examinations and rbcL chloroplast gene sequencing. For this purpose, morphological features were undertaken using valid identification keys. For the molecular analysis, genomic DNA was extracted through slightly modified CTAB. The amplification of fragments was carried out using rbcL primers. The analysis of genome sequences was undertaken using Chromas, BioEdit and MEGA6 and the phylogenetic trees were constructed through Neighbor Joining (NJ) and Maximum Likelihood (ML). The results indicated that there was a 99% chance that the two identified Padina species (P. boergessenii and P. australis) belonged to the same cluster and that there were large genetic similarities among the compared Padina species registered in GenBank

Isolation, identification and cloning of growth hormone gene from Epinephlus coioides in E. coli

According to the role of the growth hormone gene in the aquaculture, in this research cloning growth hormone coioides (Epinephlus coioides) was evaluated in vector pTZ57R/T. After PCR product purification by kit QIA quick Gel Extraction, growth hormone gene was integrated into pTZ57R/T vector. Constructed vector was transferred in E. coli strain DH5α competent cells. The white colonies was recombinant bacteria, from that that plasmid extraction was performed. Cloning obtained conformed using with direct PCR and sequencing methods. Growth hormone cDNA from E. coioide has an open reading frame of 615 nucleotides and 204 amino acids respectively. The calculated molecular weight and predicted of isoelectric point of the growth hormone protein were 014/23 kDa and 9.6 respectively. The results of this study showed that growth hormone gene in pTZ57R/T vector successfully had been cloned and can be used to growth hormone gene expression and protein production. The operation of gene sequencing with growth hormone gene from E. coioides in the Gen Bank was showed similarity between them

Genetic diversity of Crassostrea genus using 16S rRNA from coastal of Emam Khomayni Port

Nucleotide sequence polymorphism associated with 16S rRNA gene of mitochondrial genome of the Persian Gulf Oyster (Crassostrea sp.), examined in 30 samples collected from Bandar-e Emam Khomeini. Polymerase chain reaction was performed by a pair of primer for all samples and PCR products were sequenced using dideoxy chain termination method . Then the obtained data were analyzed usig MEGA Ver 5.0, BioEdit Ver 7.0 and DnaSP Ver 5.0 programs and software. The results showed 2 mutation and Nucleotide diversity (Pi) 0.00061 observed in studied genes. Among the 30 samples, 3 haplotypes were detected and average haplotype diversity (Hd) 0.195 were calculated. The haplotypes obtained in this study were submitted in the Gene Bank for the first time. Phylogeny trees derived from 16S rRNA gene sequencing showed no significant separation of the samples in the Imam Khomeini port. The results of this study also showed that there is low sequence differentiation in the study population in Imam Khomeini port

Heavy metals (Hg, Cd, Pb and Cu) bioaccumulation in the oyster Crassostrea gigas of Imam Khomeini Port

Heavy metals are major group of dangerous contaminants in marine environment. Due to their persistence, they may enter aquatic food chains and transfer from one trophic level to another. They also may be harmful for human being which is placed at the top of the food chain. Imam Khomeini’s Port is located in the north west end of Persian Gulf and ships traffic and several industries especially petrochemical industries around of the port, discharge various organic and non-organic contaminants such as heavy metals into this ecosystem. In this study in order to investigate the levels of heavy metals (Hg, Cd, Pb and Cu) in soft tissue of oyster Crassostrea gigas from Imam Khomeini port, oysters of the same size were sampled on March 2010 from 5 different locations including harbors of Petroshimi, Doc Sorsoreh, 15, 28 and 33 in the Imam Khomeini port. The samples were freeze dried and acid digested in order to determine their heavy metal content with atomic absorption spectrophotometry. Results indicated that the content of Hg, Cd, Pb and Cu in the oysters’ soft tissue were 3.12±2.97, 7.56±3.62, 7.1±3.30 and 440.57±66.17 µg/g of dry weight, respectively. The highest level of Hg, Pb and Cu were recorded in harbor of Petroshimi while the highest level of Cd was measured in harbor no 15.The concentration of Hg, Cd, Cu and Pb in bivalve soft tissue was more than standard limits. This can be dangerous for public health therefore regular monitoring of heavy metals in the area is recommended