GRASS CARP REOVIRUS INFECTION ON PHENOTYPIC MORPHOLOGY AND IMMUNE GENE EXPRESSION IN JUVENILE CTENOPHARYNGODON IDELLA

This study aimed to investigate the tissue distribution of grass carp reovirus (GCRV), external morphological alterations, and immune responses in juvenile grass carp (Ctenopharyngodon idella) following experimental injection. Fishes were challenged with GCRV type II strain YX-246 by intraperitoneal injection. The relative expression of viral and relevant immune genes in diseased fish during infection were systematically analyzed using qRT-PCR, and image processing techniques were employed to quantify the external morphological changes. Spearman correlation analysis was performed to assess the relationships between morphological indicators and viral/immune gene expression. Temporal synchrony and lead–lag relationships were further evaluated by cross-correlation analysis following first-order differencing of the time series. Mortality in the GCRV group occurred on day 4 post-infection (dpi), with a cumulative mortality rate of 100%, while there were no deaths in the PBS group. Viral load demonstrated significant tissue tropism, with the liver and brain being the main sites of replication. Immune gene expression patterns demonstrated pronounced tissue specificity. IFN1 exhibited persistent upregulation in the liver, reaching its maximum amplitude at 7 dpi, whereas in the kidney, heart, and brain, it followed an “upregulation then downregulation” trajectory, peaking at 4 dpi. Mx2 was significantly induced across all tissues, with the strongest response observed in the brain during the viral load peak period. IgM expression was notably upregulated in the liver and brain during late infection, achieving peak expression at 5 dpi and 7 dpi, respectively. MyD88 dynamics in the kidney and brain aligned with viral replication patterns, yet it was suppressed in the liver and showed no significant variation in the heart. Correlation analysis revealed that viral load was significantly positively correlated with the expression of multiple immune genes. Quantification of external morphology revealed that the percentage of bleeding area (P_bs) peaked at 4 dpi; the percentage of darkening area (P_ds) reached its peak at 5 dpi; and the dorsal fin spread angle (A_df) began to decrease persistently at 3 dpi and became significantly lower than the control level at 5 dpi (P<0.05). Furthermore, both P_ds and A_df showed significant correlations with viral load and immune gene expression, indicating that external morphological characteristics can effectively reflect viral infection status and immune response intensity at the tissue level. Cross-correlation analysis revealed that in the brain, fluctuations in P_ds and MyD88 expression exhibited a strong synchronous correlation (r=0.80, P<0.05), whereas changes in P_bs (r=0.64) and A_df (r=0.69) preceded changes in GCRV by approximately 2 days. Studies have shown that GCRV infection causes significant and regular changes in the morphological phenotype of juvenile C. idella, which are highly synchronized with key periods of immune response. However, correlations among the indicators varied, necessitating further analysis of their respective weights to construct a comprehensive evaluation model. By integrating standardized image acquisition with quantitative image analysis, this study correlates morphological features with viral load and immune gene expression, providing important data support for the clinical identification of GCHD, the development of rapid detection models, and the formulation of comprehensive prevention and control strategies.