eDNA- and eRNA-Based Detection of 2-Methylisoborneol-Producing Cyanobacteria and Intracellular Synthesis Dynamics in Freshwater Ecosystem

Taste and odor (T&O) compounds in freshwater are frequently produced by certain cyanobacteria; however, their occurrence remains difficult to predict. This study examined the temporal and spatial variations in the <i>mibC</i> gene, which encodes a critical enzyme in the biosynthesis of 2-methylisoborneol (2-MIB), by analyzing environmental DNA (eDNA) and RNA (eRNA) in the North Han River, Republic of Korea, from July 2019 to October 2021. Surface water was sampled at twelve sites and analyzed for <i>mibC</i> DNA copy number, RNA expression, cyanobacterial cell density, and 2-MIB concentration using quantitative PCR (qPCR), microscopy, and gas chromatography-mass spectrometry (GC-MS). The <i>mibC</i> gene was present throughout the year, exhibiting peaks from late summer to early winter; higher concentrations typically initiated upstream and subsequently moved downstream. RNA expression was elevated from summer to autumn, rapidly declined following heavy rainfall, and reliably preceded increases in 2-MIB concentrations by 2-4 weeks. RNA levels were strongly correlated with 2-MIB concentrations (r = 0.879, <i>p</i> < 0.001) but showed only a moderate association with <i>Pseudanabaena</i> cell density, whereas DNA demonstrated weaker correlations. More than 95% of total 2-MIB was dissolved, limiting the ability to directly estimate concentrations from eRNA data alone. The results indicate that eRNA monitoring is an effective early warning tool for T&O events. In addition, combining eDNA and eRNA analyses enables a more accurate evaluation of T&O-producing cyanobacteria, presenting practical benefits for proactive management of drinking water.