---

In this research, histological changes of gill and kidney, as a basic and important organ in osmoregulation, during migration from Musa creek to Karun and Bahmanshir rivers was studied. Ten fish from each station of Persian gulf, Karun and Bahmanshir with similarity in  biometric of size and weight were collected. After fixation in Bouin's solutions, routine procedure of tissue preparation was done and 6μm sections stained with Hematoxylin and Eosin, studied by optical microscope equipped with a daynolit lenses. The greatest number and area of chloride cells were seen in sea samples but they decreased during migration to Karun and Bahmanshir rivers (p<0.05). But there was no significant difference between samples of Karun and Bahmanshir (p>0.05). In kidney, during migration from the sea to the river, the diameter of proximal and distal lumen increased. But in Karun samples, diameter of lumen of distal tubules were higher than others station. Also, there was no significant difference in other tubule between in Karun and Bahmanshir samples. Therefore according to this study, gill and kidney have active participation in osmoregulation. This fish can be a good model for study of osmoregulation in different salinities because of adaptation and response to different environmental salinities in gill that associated with histologic changes. 

---

According to morphological studies, Leptodius exaratus is one of the most common brachyuran species and the only known species of this genus which have been found in intertidal rocky shores along the Persian Gulf coast. Since the existence of hidden species among the crustacean species are very common, the aim of this study is to investigate the probability of hidden species identification among eight color morphotypes that were identified as Leptodius exaratus species in morphological studies. For this purpose, eight color morphotypes of Leptodius exaratus species were collected from rocky shores of Bushehr province. Then, the first male gonopods were separated and photographed by Scanning Electron Microscopy (SEM). For conducting molecular studies, DNA was extracted with a phenol-chloroform mixture and mitochondrial Cytochrome Oxidase subunit I (COI) gene was amplified and sequenced. The results showed that mitochondrial COI nucleotide sequences were not identical in all color morphotypes. These results were reflected in topology of evolutionary trees for both analyses (maximum likelihood, bayesian). Studying the ultrastructure of the apical segments of the first male gonopods showed fundamental differences in the apical organs of some color morphotypes. These results indicated that all samples did not belong to a single species and there were hidden species among them. Molecular evidence also showed that mitochondrial COI nucleotide sequences of eight color morphotypes of Leptodius exaratus from the Persian Gulf and the registered sequence of such species from Indonesia were in separate clusters. Identification based on the morphology of the first male gonopods using scanning electron microscopy (SEM) and the COI gene were useful research methods in the present study.
 

---

Sea cucumbers produce various bioactive compounds especially secondary metabolites including Saponin. The aim of this study was to investigate antibacterial activity of the Saponin extracted from the Persian Gulf sea cucumber (Stichopus hermanni) in some gram positive and negative bacteria. Extraction was first done by ethanol 70% and then Saponins were separated through column chromatography with silica gel. Extracted Saponins were identified through thin layer chromatography (TLC) and high performance thin layer chromatography (HPTLC). Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were investigated via tubular dilution. Two groups of steroidal and glycosides Saponins with R_f 0.9 and 0.05 were respectively identified in fractions extracted by TLC. HPTLC results for a sample of extracted Saponin confirmed the existence of Saponin in sea cucumber regarding the R_f. The gram negative bacterium Pseudomonas aeroginosa was the most resistant one to steroidal and glycosides- steroidal Saponins with MIC of 500 and 400 micg/ml, respectively. Steroidal and glycosides- steroidal Saponins showed no bactericidal effect in gram negative bacterium Pseudomonas aeroginosa. Among the gram positive bacteria, the highest MBC of 500 micg/ml belonged to glycosides- steroidal Saponin for Staphylococcus aureus. The higher MIC in gram negative bacteria approves the lower antibacterial property of the steroidal and glycosides Saponin extracted from sea cucumber Stichopus hermanni in these bacteria compared to the gram positive ones.
 

---

This study aimed to identify the effect of environmental conditions on the diversity and abundance of polychaetes in the southern shores of the Caspian Sea in Guilan, Iran. Sampling of seasonal sediments was done (June 2017 - May 2018) in 4 stations and three depths of 1, 5 and 10 m with the help of a Van Veen grab sampler with cross section of 225 cm² and three replications. Temperature, salinity, pH, dissolved oxygen, suspended solids and organic matter were recorded at each station. A total of 2711 polychaetes belonging to 5 species, 5 genera from 3 families were identified. The highest abundance was related to H. kowalewskii (39%) and the lowest abundance was related to H. invalida species (3%). The mean density in the four seasons of spring, summer, fall and winter was 67.66, 81.25, 48.66, and 28.33 worms per m², respectively. The highest species diversity was 2.16 species per square meter and the biomass of 62.75 mg/m² in spring and the lowest was in winter. The highest density, diversity and biomass were 80.41 worms per square meter, 2.33 species per square meter and 70.83 mg/m² on the shore of Chamkhaleh, respectively; and the lowest were 16.83 worms per square meter, 1.5 species per square meter, and 31.41 mg/m² on the shore of Chaboksar, respectively. The highest density, diversity and biomass were 127.87 per square meter, 3.68 species per square meter and 107 mg/m² at a depth of 10 meters, respectively. The correlation results showed that among environmental factors, only organic matter had a direct significant relationship with species diversity. Density had a significant direct relationship with temperature and organic matter. Other factors did not show a significant relationship. The results of stability level of different species in the southern shores of the Caspian Sea (Guilan province) showed that N. diversicolor and H. kowalewskii species exist temporarily and other species randomly.

---

Epixanthus frontalis is the only known species of the genus Epixanthus from the Persian Gulf which include color polymorphic specimens. Considering that cryptic species are common among crustaceans, the aim of this study was to identify the cryptic species complexes the collected samples from the Persian Gulf. For this purpose, 4 colorful morphotypes of E. frontalis collected from rocky shores along Bushehr province. Then the first male gonopods was separated and photographed by scanning electron microscopy (SEM). For molecular studies samples of DNA extracted by using phenol-chloroform and mitochondrial cytochrome oxidase subunit gene segment (COI) was PCR and sequenced. The results showed that the nucleotide sequence of this gene fragment of the mitochondrial (COI) is not identical in all colorful morphotypes. These results reflected in the evolution of tree topology for all two analyses (maximum likelihood, bayesian). The study of apical ultrastructure segments of the first male species showed that it can be seen fundamental differences in the apical organs in some colorful morphotypes. Results of this study indicated that although the specimens assigned to single species morphologically, they may belong to different species based on their genotype. Further molecular studies with specific markers are needed for molecular identifications.       

---

Introduction
Emerging anthropogenic pollutants in wastewater treatment plant (WWTP) effluents have been shown to pose a particular risk to aquatics and potentially affect the aquatic environment from the molecular to the ecosystem level (Santos et al., 2010). Micropollutants are emerging pollutants that include pharmaceuticals and personal care products (PPCPs). Due to the widespread consumption of a wide range of micropollutants and their discharge into wastewater worldwide, their occurrence and fate in aquatic environments has become a research topic in recent decades (Ryu et al., 2014). The common wastewater treatment process is not able to completely remove these compounds from the wastewater, and as a result, a significant amount of these compounds enters the aquatic water environment (Rehman et al., 2015). Dexamethasone is a potent synthetic glucocorticoid with anti-inflammatory and immunosuppressive effects that is used to treat many diseases including allergies, asthma, covid-19, rheumatic problems, and skin diseases (Sanders et al., 2020). Today, the increasing use of dexamethasone from sources such as hospitals is a global concern. High levels of this drug, which is a cortisone derivative, have been detected in wastewater (Herrero et al., 2013). As one of the important components of the innate immune system in ectothermic vertebrates, including fishes, the complement system has been less studied. Evaluation of serum complement activity is considered as a valuable tool to diagnose the health status of fish. Complement in the non-specific immune response to a compound can have a direct effect such as killing the pathogen by lysing it (Ellis, 1999).
Methodology
Fifteen mature Arabian sea bream, A. arabicus were collected from the Bahrakan Port in Abadan. Fish were transported to the 300 L tanks. After seven days of adaptation, the fish were anesthetized using 2-phenoxy ethanol (0.35 ml/L). Then, the fish were dissected under aseptic conditions and their spleens and kidney heads were separated (Wen et al., 2008). The separated tissues were immediately washed three times with 100 ml antibiotic medium (Leibovitz's L-15 (L-15) with 400 IU/ml penicillin, 400 µg/ml streptomycin and 200 µg/ml amphotericin B) each time for 30 minutes. The spleen cell culture was performed according to Huang et al. (2009). The head kidney cell culture followed the protocols described by Ribera et al. (2020). Dexamethasone cytotoxicity was assessed by MTT method according to Momeni et al. (2010). Cultivated spleen and head kidney cells were plated in 24-well microplates (10⁵ cells / ml L-15 medium / well) and incubated at 28°C for 24h. The cell viability was assessed using trypan blue exclusion test. Then, 200 μL of fresh L-15 culture medium with different experimental concentrations of dexamethasone was added to each well and microplates were then incubated at 28°C for 48 h. Each treatment runs in five replicates. For this purpose, the cell suspensions were collected at 0, 12, 24 and 48 hours of experiment and transferred to microtubes for further analysis. The level of C3 and C4 was measured using immunoturbidimetry method by a double–antibody sandwich ELISA kits. ACH50 was measured by hemolytic method according to Sunyer and Tort (1995).
Results
Measurement of dexamethasone in hospital wastewater: To measure dexamethasone in hospital wastewater, samples were taken from the wastewater of Taleghani Hospital in Abadan, and after scanning the dexamethasone samples and determining the maximum absorption wavelength at 213 nm for it, the concentration of dexamethasone was measured using an HPLC device. The obtained concentration was 86 ng/l. Primary culture of head kidney and spleen cells: Using the trypan blue test, the survival rate of spleen and head kidney cells was determined to be 95%. Then, cells derived from the spleen and head kidney were incubated in L-15 culture medium for two weeks at 28°C. After 48 hours, cells started to grow and colony formation started 5 days after incubation. The cells grew well and completely occupied the culture dish within 12 days. All the time, the cells were floating in the culture medium as round cells and did not stick to the bottom of the culture container. The trypan blue test showed that more than 90% of the cells were alive and healthy during the test. Twelve days after incubation, the cells were passaged. The second passage was performed fifteen days after incubation and then the cells were used for cytotoxicity assay. The C3 contents notably decreased in spleen cells exposed to all concentrations of dexamethasone except for the lowest concentration (3 μM; P < 0.05). In the head kidney, the C3 contents only notably decreased in cells treated with the higher concentrations of dexamethasone (3×10 and 3×10² μM; P < 0.05). The complement C4 has no obvious change following exposure of spleen and head kidney cells to all concentrations of dexamethasone except for the highest concentration. The C4 content of spleen and head kidney cells significantly decreased after 24 h exposure to highest concentration of dexamethasone (P < 0.05).  The concentrations of dexamethasone (except the highest concentration of dexamethasone) caused a significant change in the ACH50 content of spleen cells compared to the controls (P > 0.05). The exposure of spleen cells to 3 mM dexamethasone led to an 11% decrease in alternative complement activity after 48 hours of exposure (P < 0.05). ACH50 activity in head kidney cells treated with 3×10² nM and 3 μM of dexamethasone showed no difference compared to control (P > 0.05). A significant decrease in ACH50 was observed after exposure of head kidney cells to 3×10 and 3×10² μM of dexamethasone (P < 0.05). The amount of ACH50 activity in head kidney cells treated with 3 mM dexamethasone decreased by 27% after 48 hours of exposure (P < 0.05)
Discussion and Conclusion
This study is the first study that reports the suppressive effects of dexamethasone on the activity of complement components of head kidney cells and spleen of Arabian yellow fin sea bream (Acanthopagrus arabicus) in cell culture medium. Dexamethasone is commonly known as an immunosuppressant drug that is toxic to immune cells and inhibits the function of the immune system (Ribas et al., 2016). The complement system plays an important role in innate and acquired immunity by facilitating the function of phagocytizing cells and cell lysis. The components of this system, which are activated in a cascade manner, lead to the destruction of pathogens by targeting the membrane components of invading agents (Boshra et al., 2006). C3 is an important component of the complement system, whose stimulation activates other components. This component belongs to the acute phase cellular proteins and is the first component of the complement system to be activated (Bayne et al., 2001). In the present study, dexamethasone led to a significant decrease in C3 levels in spleen and head kidney cells. On the other hand, the level of C4 in spleen and head kidney cells was suppressed only by the highest concentration of dexamethasone. In the study of Zach et al. (1993), corticosteroids can suppress the expression level of genes related to the synthesis and activity of C3 through the glucocorticoid (GR) and mineralocorticoid (MR) receptors. Most Osteichthyes have three corticosteroid receptors, including two glucocorticoid receptors (GR1 and GR2) and one mineralocorticoid receptor (MR), which are encoded by three different genes (Zach et al., 1993). This can explain the decrease in complement levels under the influence of dexamethasone (corticosteroid) in the present study. In the present study, dexamethasone did not affect the activity of alternative complement component (ACH50) of spleen and head kidney cells, and only its highest concentration decreased ACH50 levels after 48 hours. Milla et al. (2018) did not observe any changes in gene expression, protein quantity or activity of proteins related to innate immunity (such as ACH50 level) up to two weeks after exposure of Perca fluviatilis to 11-deoxycorticosterone (corticosteroid). While after two weeks, a significant increase in the ACH50 level of the spleen was observed. They stated that acute exposure to corticosteroids (as immunosuppressants) does not seem to have a significant effect on some innate immune proteins (Milla et al., 2018).
Acknowledgment
This work was supported by the Khorramshahr University of Marine Science and Technology.