Background: Rotavirus infection is one of the most common gastroenteritis in the world, and a million cases are registered to enter hospital every year. Promyelocytic leukemia proteins (PMLs) are IFN-up-regulated proteins, and one of their critical functions is working as antiviral proteins. Recently, promyelocytic leukemia isoform II (PML-II) has been depicted as an isoform responsible for the antiviral function. Methods: Rotavirus prevalence determination was achieved by PCR and Rapid Adeno/Rota Virus test, while the relative expression assay was carried out by real-time PCR technique. Blood and stool samples were collected from 34 children under five years admitted to the hospital with acute gastroenteritis showing signs of dehydration. RNA samples were extracted from blood specimens and converted to cDNA to be used in gene expression analysis of PML, PML-II, and IFN-γ in rotavirus positive or negative samples. Results: Rapid Adeno/Rota Virus Antigen Combo Test and PCR assay could detect the virus in stool samples in 45% and 17.6% of cases, respectively. PML in positive samples decreased to 104fold less than the level in negative ones. The same trend was noticed in the level of IFN-γ and PML-II expression as their expression reduced to 104 or 13fold in rotavirus-infected samples compared to the control, respectively. Conclusion: Altogether, our data showed that the gene expression of PML, PML-II, and type II IFN considerably diminished in rotavirus-infected samples compared to the negative control.