Enhanced sialic acid engagement at physiological temperatures by reovirus σ1 mutants facilitates infection of breast cancer cells with low levels of high-affinity receptors.

Reovirus serotype 3 (T3D IMPORTANCE: Reoviruses are promising oncolytic agents, yet clinical efficacy can be hindered by heterogeneous receptor expression in tumors. This study demonstrates that reovirus can bypass high-affinity receptor requirements by optimizing sialic acid interactions or incorporating RGD motifs. Crucially, the data reveal that wild-type reovirus attachment to sialic acids is thermally unstable at physiological temperature (37°C), a restriction masked by traditional 4°C assays. Specific mutations were found to stabilize these interactions at 37°C, providing a mechanistic basis for viral adaptation to receptor-deficient environments. These findings establish new experimental approaches to study attachment at 37°C, which can be applied broadly to discover unanticipated mutational effects on viral entry. Ultimately, evaluating virus-cell attachment under physiological conditions is essential for accurately predicting viral tropism and facilitates the design of next-generation oncolytic therapies better equipped to overcome receptor scarcity and thermal barriers in complex tumor environments.