Apparent cooperativity between human CMV virions introduces errors in conventional methods of calculating multiplicity of infection

Human cytomegalovirus (HCMV) virions may display apparent cooperativity when infecting cells, a phenomenon that challenges conventional methods of calculating the multiplicity of infection. To investigate this, researchers examined the infection process for three HCMV strains on fibroblasts and epithelial cells, titrating viral inoculum concentrations in precise steps.

Using flow cytometry to measure infected cell frequency, they discovered that cell infection rates climbed faster than linearly with rising inoculum concentrations, indicating cooperation between individual infecting virions. Mathematical modeling failed to find explanations rooted in viral or cellular heterogeneity, or simple aggregation/clumping of viral particles.

Instead, stochastic simulations of alternative models, suggesting cell resistance reduction upon multiple virion exposure or compensatory effects in infectivity of less infectious virions, yielded similar apparent cooperativity results. Further analysis of existing datasets revealed similar patterns for HIV and vaccinia virus infecting CRFK or HeLa cells, but not for tobacco mosaic virus on plant leaves.

The study suggests the need for a rigorous evaluation methodology to distinguish true cooperativity from statistical artifacts, emphasizing the importance of viral cooperativity knowledge for accurate multiplicity of infection quantification.