This study evaluated the pathophysiological, biochemical, and immunological status of olive flounder (Paralichthys olivaceus) infected with the myxosporean parasite Kudoa septempunctata. Flounder fish collected from Kudoa-infected and uninfected farms were confirmed by microscopic and TaqMan probe-based quantitative PCR screening. Morphological, biochemical, histological, and immune gene expression analyses were performed on uninfected and infected hosts to assess the effect of K. septempunctata. Histological studies confirmed the presence of Kudoa myxospores in the trunk muscles of infected flounder fish. Serum biochemical parameters, including the levels of myeloperoxidase activity, superoxide dismutase activity, alanine aminotransferase, alkaline phosphatase, amylase, bilirubin, total protein, cholesterol, calcium, potassium, sodium, phosphorus, glucose, and galactose, were found to exhibit no significant variations (p > 0.05) between uninfected and infected flounder fish. However, immune-related genes such as Mx, lysozyme, signal transducer and activator of transcription 1, interferon-γ, interferon regulatory factor, and tumour necrosis factor showed significantly elevated expression (p < 0.05) in the trunk muscles of infected flounder fish while no significant differences were noted in uninfected fish trunk muscle and head-kidney of infected and uninfected flounder fish.