Effects of artificial photoperiods on antigen-dependent immune responses in rainbow trout (Oncorhynchus mykiss).

In this study, we investigated the effects of the artificial photoperiods that mimic summer (16L:8D; 16 h Light: 8 h Dark) and winter (8L:16D) solstices, equinoxes (12L:12D), and the artificial 24-h light regimen (24L:0D) on the leukocyte populations and the T helper and regulatory type responses on rainbow trout (Oncorhynchus mykiss). Using flow cytometry analysis, we found that photoperiod induces changes in head kidney leukocyte subsets. The lymphoid subset increased in the 16L:8D summer solstice regime. The analysis using antibodies against B and T cells showed the increase of CD4-1+ T lymphocytes and other unidentified lymphoid cells, with no changes in the B cells. To investigate the modulatory influence of the photoperiod on the fish T cell response, we quantified in the head kidney the transcript levels of genes involved in the Th1 type response (t-bet, ifn-Æ´, il-12p35, il-12p40c), Th2 type response (gata3, il-4/13a), Th17 response (ror-Æ´t, il-17a/f), T regulatory response (foxp3α, il-10a, tgf-ß1), and the T cell growth factor il-2. The results showed that the seasonal photoperiod alone has a limited influence on the expression of these genes, as the only difference was observed in il-14/13a and il-10a transcripts of fish kept on the 16L:8D regimen. In addition, the 24L:0D treatment used in aquaculture produces a reduction of il-14/13a and il-17a/f. We also evaluated the effect of photoperiod in the presence of an antigenic stimulus. Thus, in fish immunized with the recombinant viral protein 1 (rVP1) of infectious pancreatic necrosis virus (IPNV), the photoperiod had a striking influence on the type of adaptive immune response. Each photoperiod fosters a unique immune signature of antigenic response. A classical type 1 response is observed in fish subjected to the 16L:8D photoperiod. In contrast, fish in the 12L:12D photoperiod showed only the upregulation of il-12p40c. Furthermore, none of the cytokines were increased in fish maintained on the artificial 24L:0D regimen, and a decrease in the master transcription factors (t-bet, ror-Æ´t, and foxp3α) was observed. Thus, fish on the 12L:12D and 24L:0D photoperiod appear hyporesponsive regarding the T cell response. Altogether, this study showed that photoperiods modify the magnitude and quality of the T-helper response in rainbow trout and thus impact essential mechanisms for the generation of immune memory and protection against microorganisms.

In vitro fertilization with cryopreserved spermatozoa in small pieces of gonad of the scallop Argopecten purpuratus (Lamarck, 1819).

The aim of this work was to evaluate the fertilization rates through the first cleavage, obtained with fresh oocytes inseminated with sperm cryopreserved at different freezing rates (-8.8,-10 and -12 °C/min) and at two thawing rates, using cryoprotectant solution (Me2SO, at 1 M, 2 M, 3 M, 4 M and 5 M with or without egg yolk and sucrose). Sperm contained in small pieces of male gonad were frozen at the three freezing rates, stored in liquid nitrogen and later the samples were thawed at two rates by immersing the samples in water at 50 °C (rapid) or 30 °C. Control fertilization rates ranged from 69.2 ± 2.8%-45.5% ± 1.6%. To determine the best concentration of the cryoprotectant (between 1 M and 5 M), in a first step, a freezing of -15 °C/min and a rapid thawing was used. Fertilization rates ranged between 9.6 ± 2.5% and 34.6± 12.2% and the highest percentages of fertilized oocytes (34.6%) was obtained with 3 M concentration with cryoaditive. The second step, using 3 M Me2SO with cryoadditive, determined that the freezing rate -8.8 °C/min produces the best result 29 ± 2.9% of fertilized oocytes corresponding to 59.2 ± 9.1% compared to controls. Although there were no significant differences among the different freezing rates, the fertilization rate tended to be higher with a lower freezing rate. Comparing the results of the present study, which used a cryoprotective solution composed of Me2SO and a cryoadditive, to those of other studies that used Me2SO without cryoadditives, suggests that the addition of a cryoadditive to the cryoprotectant Me2SO improves the fertilizing capacity of the sperm of Argopecten purpuratus after being cryopreserved.

Transcriptional Evaluation of Neuropeptides, Hormones, and Tissue Repair Modulators in the Skin of Gilthead Sea Bream (Sparus aurata L.) Subjected to Mechanical Damage.

The skin of bony fish is the first physical barrier and is responsible for maintaining the integrity of the fish. Lesions make the skin vulnerable to potential infection by pathogens present in the aquatic environment. In this way, wound repair has barely been studied in gilthead sea bream. Thus, this study investigated the modulation of peripheral neuro-endocrine and tissue repair markers at the transcriptional level in the skin of teleost fish subjected to mechanical damage above or below the lateral line (dorsal and ventral lesions, respectively). Samples were evaluated using RT-qPCR at 2-, 4-, and 20-days post-injury. Fish with a ventral lesion presented a trend of progressive increase in the expressions of corticotropin-releasing hormone (crh), pro-opiomelanocortin-A (pomca), proenkephalin-B (penkb), cholecystokinin (cck), oxytocin (oxt), angiotensinogen (agt), and (less pronounced) somatostatin-1B (sst1b). By contrast, fish with a dorsal lesion registered no significant increase or biological trend for the genes evaluated at the different sampling times. Collectively, the results show a rapid and more robust response of neuro-endocrine and tissue repair markers in the injuries below than above the lateral line, which could be attributable to their proximity to vital organs.