Mitomycin C eliminates cyanobacterial transcription without detectable prophage induction in a <i>Microcystis</i> -dominated harmful algal bloom in Lake Erie

Although evidence indicates that viruses are important in the ecology of <i>Microcystis</i> spp., many questions remain. For example, how does <i>Microcystis</i> exist at high, bloom-associated cell concentrations in the presence of viruses that infect it? The phenomenon of lysogeny and associated homoimmunity offer possible explanations for this question. Virtually nothing is known about lysogeny in <i>Microcystis</i>, but a metatranscriptomic study suggests that widespread, transient lysogeny is active during blooms. These observations lead us to posit that lysogeny is important in modulating <i>Microcystis</i> blooms. Using a classic mitomycin C-based induction study, we tested for lysogeny in a <i>Microcystis</i>-dominated community in Lake Erie in 2019. Treated communities were incubated with 1 mg L^-1 mitomycin C for 48 h alongside unamended controls. We compared direct counts of virus-like particles (VLPs) and examined community transcription for active infection by cyanophage. Mitomycin C treatment did not increase VLP count. Mitomycin C effectively eliminated transcription in the cyanobacterial community, while we detected no evidence of induction. Metatranscriptomic analysis demonstrated that the standard protocol of 1 mg L^-1 was highly toxic to the cyanobacterial population, which likely inhibited induction of any prophage present. Follow-up lab studies indicated that 0.1 mg L^-1 may be more appropriate for use in freshwater cyanobacterial studies. These findings will guide future efforts to detect lysogeny in <i>Microcystis</i> blooms.IMPORTANCEHarmful algal blooms dominated by <i>Microcystis</i> spp. occur throughout the world's freshwater ecosystems, leading to detrimental effects on ecosystem services that are well documented. After decades of research, the scientific community continues to struggle to understand the ecology of <i>Microcystis</i> blooms. The phenomenon of lysogeny offers an attractive potential explanation for several ecological questions surrounding blooms. However, almost nothing is known about lysogeny in <i>Microcystis</i>. We attempted to investigate lysogeny in a <i>Microcystis</i> bloom in Lake Erie and found that the standard protocols used to study lysogeny in aquatic communities are inappropriate for use in <i>Microcystis</i> studies, and perhaps freshwater cyanobacterial studies more broadly. This work can be used to design better methods to study the viral ecology of <i>Microcystis</i> blooms.