الفهرس 
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Abstract
Spring viremia of carp (SVC) is a serious viral disease, caused by a member of the genus Sprivivirus of the family Rhabdoviridae. SVC outbreaks can cause significant economic losses. The immune responses to Spring viremia of carp virus and the exact mechanisms by which the virus induces pathogenesis are poorly understood. This study aimed to investigate the modulatory effects of SVCV infection on immune- related genes.
Common carp were immersed in SVCV suspension (3×104 TCID50/ml) for 4 hours at 17±1°C. Nine fish (3 fish from each of triplicates) were randomly selected from different aquaria of the treated and control groups at each time point (1st, 3rd, 5th, 7th, 9th, 11th, 14th, and 20th) days post infection (dpi). Fish were euthanized and gills, brain, intestine, and visceral organs (liver, spleen, and kidney) were sampled.
Expression of the target genes (IL-1β, IL-10, IL-12, IFNγ-1, and IFNγ-2) were estimated using the carp actin and elongation factor as internal controls.
In gills, IL-1β was upregulated at the 1st dpi eliciting 3.7-fold as the control and showed the highest expression at the 3rd dpi, where it reached 12.4-fold as control and still upregulated at 5th, 7th, 9th, and 11th dpi but with less levels recording 4.1, 4,
3.6, and 1.9-folds as that of control, respectively. IL-10 displayed an expression of 5.7-fold as the control at the 3rd and still upregulated at the 5th, 7th, and 9th dpi, exhibiting 2.1, 2.2, and 2.4-folds as the control, respectively. While the expression of IL-12 recorded highest expression of 2.3-fold at the 3rd dpi as control and continued in its upregulation at the 5th dpi with 1.8-fold as the corresponding control. IFN γ-1 reached the highest expression at the 5th dpi (5.3 -fold) as that of control and still upregulated at the 7th, 9th dpi showing expression levels of 3, 3.1-folds than control. While IFNγ-2 gene was significantly upregulated reaching 2-fold at the 3rd dpi and 1.7-fold at the 5th dpi in comparison to control. Thus, in gills, the most prominent cytokine at the 1st dpi and 3rd dpi was IL-1β. Among the examined genes both IL-1β and IFNγ-1were the highest expressed ones at the 5th ,7th, and 9th dpi. Starting from the day 11th post infection, insignificant differences were recorded among all examined genes and their expression levels were similar to that of control. In visceral organs, all examined genes exhibited significant upregulations only at the 3rd dpi. IL-1β displayed an upregulation of 34.2-fold as that of control, IL-10 was expressed 12-fold higher than of its control, IL-12 recorded 10.3-fold as the control, IFN γ-1 elicited 4.6-fold as the control, and IFN γ-2 showed 8.5-fold as that of control. When compared altogether at different time points, there were insignificant differences among all examined genes except at the 3rd dpi, where IL1β was significantly upregulated in comparison with the other genes. In the intestine, IL-1β was upregulated at the 7th dpi exhibiting 4.8 -fold as that of control and continued its upregulation eliciting 5.8-fold at the 9th dpi, and at the 11th dpi it reached its maximum expression with 6.3 -fold as the control. On the other hand, IL-10 was significantly upregulated later than IL-1β and reached 2.3-fold at the 9th dpi and the maximum upregulation at the 11th dpi with 4.1 -fold higher than control. Similarly, the expression of IL-12 showed a significant increase of 3.2-fold at the 9th dpi and displayed the maximum expression with 7.6 -fold at the 11th dpi than its control. The upregulation of IFNγ-1 gene was noticed also starting from the 9th dpi and continued till the 11th dpi recording 17-fold and 21-fold as the control, respectively. IFN γ-2 showed an upregulation only at the 11th dpi with 7.5 -fold as the control. There were insignificant differences among genes at different time points except for IFNγ-1 which displayed significant differences in comparison with the other genes at the 9th and 11th dpi. In brain, the IL-1β was upregulated at the 9th dpi displaying 3.3-fold as control and reached its peak (9.1-fold) at the 11th dpi, then it downregulated but still significantly higher than control at the 14th dpi eliciting 3.6-fold than its control.
While IL-10 increased at the 9th dpi recording 2.7-fold than its control and showed the highest expression at the 11th dpi eliciting 4.1 -fold as that of its control. from the day 14th post infection, its expression was similar to the control level. Similarly, IL-12 was upregulated at the 9th dpi giving 4.3-fold and showed the maximum expression at the 11th dpi eliciting 8.8-fold than that of control. IFN γ-1 and IFN γ-2 genes exhibited significant upregulation only at the 11th dpi eliciting 6 -fold and 5.7 -fold, respectively than their controls. Conclusively, in brain, the most prominent cytokines at the 9th dpi were IL-1β and IL-12 in comparison with the others, and were significantly upregulated at the 11th dpi, when compared to IL-10, IFNγ-1, and IFNγ-2. At the 14th dpi, only IL-1β showed a significant upregulation in comparison with the other genes. In gills and visceral organs, the highest fold change was IL-1β at the 3rd dpi. In the intestine, IFN γ-1 was the dominant gene at the 11th dpi. In the brain, IL-1β, IL12, and IFNγ-1 dpi showed higher expressions among other genes at the 11th dpi.The fold changes of IL-1β and IL-10 were significantly higher in visceral organs and gills at the 3rd dpi than other genes. The highest level of IL-12 was found in visceral organs at the 3rd dpi, in intestine, and brain at the 11th dpi. The highest levels of IFN γ-1 were found in the intestine at the 9th and 11th dpi with the maximum fold change at the 11th dpi. The visceral organs at the 3rd dpi had the highest level of IFN γ-2 followed by intestine and brain at the 11th dpi.
For determination of SVCV copy number in selected organs, the TaqMan realtime PCR assay was carried out by using QuantiTect Probe PCR kit. In gills, the viral copies were the highest at the 1st dpi and the 3rd dpi recording (3.08×109) and (2.78×109), respectively. While in the visceral organs reached its peak at the 3rd dpi (6.67×108). In intestine, viral copies were significantly increased to reach its maximum numbers at the 7th dpi (9.0×108). In brain, the viral copies increased to reach the maximum at the 9th dpi (1.03×109).