DETERMINATION OF DOPAMINE AND SEROTONIN WITH IMMUNE STATE CD4 AND CD8 OF THE NEWCASTLE INFECTION IN BROILERS

This study was aimed to describ the course of Newcastle disease at 24 and 72 hr. of infections in broilers flocks within the period (1-December-2021 to 1- Jun-2022) and the study aimed to determine the immunological (CD4, CD8) response and changes in neurotransmitter hormones. The blood samples were collected and prepared then examined by ELISA kits test and were worked under the protocols of companies. The result of immunological response showed the response to cellular immunity (CD8) in all cases increasing within 24hr.with statistically represented levels while, the result of humoral immunity (CD4) increasing in all cases except two cases within 72hr. with statistically significant levels as well as the results of (CD4/ CD8) ratio. while the results neurotransmitter hormones concentration revealed all cases statistically decreased represented levels at 72hr. The study concluded the 1 st immune response within 24hr. showed elevation with cellular immunity then humoral immunity after 48-72hr. Also the ND infections caused down-regulation of neurotransmitter hormones by decreasing to serotonin and dopamine levels during all periods of infections


INTRODUCTION
Newcastle disease (ND) is one of the major viral contagious diseases in poultry, causing heavy economic losses in poultry production and causing retard growth with high morbidity and mortality and a decrease in egg production (10,31). The (NDV) classified to family Paramyxoviridae, type one (PMV-1) negativesense RNA, single-stranded, non-segmented (24). The ND disease remains the main problem in the poultry industry because caused damage to nervous, respiratory, and GIT systems (7). Also caused failure to the immune response to ND vaccine by causing disturbance to humoral and cellular immunity (27,30). The immune response after six hours' post-infection to ND splenic cell proliferation and produce alpha and beta interferon with (IL-6) to activate cell-mediated immunity (CMI) (15,23). Also, the T-lymphocyte through T helper cell type 1 especially Cytotoxic T lymphocytes (CD8) immediately activated after several hours' post-infection with ND (18). Then activation became more advance through stimulation other of cytokines and T helper cell type 2 (CD4) within 2-3 days of ND infections (29). The infection with ND will cause a challenge to the immune response and activation of the (hypothalamic-pituitaryadrenocortical) center, as well as the concentrations of corticosterone and hypothalamic will increase after exposure to lento-mesogenic (NDV) that will cause down release to serotonin and dopamine levels to change neurochemical responses (1,2,9). Dopamine and serotonin (neurotransmitter hormones) in the brain are responsible for several critical functions (19). These hormones act on the regulation immune system in addition to CNS processes, gastrointestinal motility, blood pressure, and neuroimmune communication (8,23).

MATERIALS AND METHODS Sampling and Preparation:
Thirty samples were collected from Broilers infected flocks during the course of Newcastle disease within 1st and 3rd days of infections between the period (1-December-2021 to 1-Jun-2022) (14). Blood samples were collected in a gel tube and then exposed to a centrifuge (1500/ rpm × 15mint) serum was collected in an Eppendorf

Statistical analysis
The program of Statistical Analysis System, SAS was used different study parameters (Chisquare) to show Least significant difference between transactions and (LSD test) to find significant contrast between variable means (26).

RESULTS AND DISCUSSION
The results show in Table-1 immunological response to humoral immunity (CD4) and cellular immunity (CD8) during the course of Newcastle disease infections result of humoral immunity (CD4) in all cases except two cases revealed increasing levels at 72hr. In comparison with 24hr. with statistically significant at (P≤ 0.05) levels meaning the humoral immunity will be activated after 24-72hr.while the response of cellular immunity (CD8) showed all cases increasing of CD8 levels at 24hr. In comparison with 72hr. with statistically significant at (P≤ 0.05) to ND infection represented the cellular immunity and 2nd response humoral immunity. These results of immune response to cellular and humoral immunity agree with other studies that represented the CMI firstly activated and rapidly expand T-lymphocyte in the thymus gland and spleen especially (CD8) T lymphocytes after mints of exposure to NDV antigen (17,18). Activated T-cell helper (CD4) to activate and differentiate B-lymphocytes into plasma cells to produce antibodies to neutralize specific NDV antigens (16,17). Also, the infection with a lentogenic strain of NDV caused T cells activation and proliferation in vivo and in vitro (12). As well as the immune response to infection with NDV immediately develop T-cell through activation of cytotoxic T cells CD8 a regulatory role and follow by helper T cells to express CD4 on the surface (4,17). The challenge by field strains of NDV caused a significant increase (p ≤ 0.001) in the T-cell population specifically (CD8 and CD4) in the intestinal and respiratory tissue in broilers chicken and in the sera (13).  Table-2 show (CD4/ CD8) ratio between time 24hr. and 72hr. during the course of Newcastle disease infections showed all cases increasing of (CD4/ CD8) ratio levels at 72hr. In compression with 24 hr. with statistically represented at (P≤ 0.05) levels meaning development of both cellular and humoral immune responses. The results of the (CD4/ CD8) ratio also agrees with the study that revealed the administration of the different commercial vaccines to ND days old in broiler flocks increased the concentration of (CD4-CD8) ratio with high significant levels (p≤0.001) levels when detected by ELISA (4,8,17), and immunohistochemistry in the each of intestine, trachea, and serum (3,5). While the Bar-Shira et al., rvealed no significant difference in the average (CD4/CD8) ratio after both 1st and 2nd vaccination with the B1 strain of ND vaccine (6). The result (Table-3) of levels neurotransmitter hormones (Dopamine, Serotonin) during the course of Newcastle disease infections shown are as follows: The result of Dopamine and Serotonin concentration revealed all cases decreased with statistically significant at (P≤ 0.05) levels at 72hr. in compression with 24hr.ND infections. The result of dopamine and serotonin concentration during the course of Newcastle disease infections was compatible that revealed the following. The neurotransmitter hormones especially dopamine and serotonin levels had a relationship with immunological state because play a major role in body activities (appetite, weight gain, energy expenditure, mediatory role of ingestion, and animal behavior) because act a stimulatory, modulatory or inhibitory role in avian species (21,25). The challenge of NDV showed an increase in plasma corticosterone hormone that will lead to a decline in the concentration of dopamine and serotonin levels in mice (1,10). The deficiency in serotonin and dopamine levels caused cannibalism and feather and vent picking and the deficiency of neurotransmitter hormones had a close relationship with neurological signs (4,11). Also, the levels of dopamine and serotonin were significantly decreased (p≤0.05) in broiler chicken feeding a contaminated diet with mycotoxin when compared with groups feeding a diet containing anti-mycotoxin or free mycotoxin (22). In general, the immune and nervous system participate in extensive pathways mediated by a wide of hormones, cytokines, and neurotransmitters (20,23), and elicits immune responses that will lead improve immunity and healthy performance (8,28).

CONCLUSIONS
The conclusion of the study of the Newcastle disease infection course has two immune responses the primary response activation of cellular immunity through increasing of (CD8) T-cell levels during the 1 st 24 hr. of infection then, activation of the secondary immune response by humoral immunity through increasing of (CD4) T-cell levels within 72hr., as well as the ND infections caused downregulation to neurotransmitter hormones through decreasing to serotonin and dopamine levels in all period of infections.