Evaluation of the MIB-producing potential based on real-time qPCR in drinking water reservoirs

Abstract

Cyanobacteria release 2-methylisoborneol (MIB) as a secondary metabolite. Here, we propose a reverse tran- scription quantitative real-time PCR (RT-qPCR) based method to evaluate the MIB-producing potential in source water by detecting the MIB-synthesis gene (mic). A MIBQSF/R primer set was designed based on 35 mic gene sequences obtained from 12 pure-cultured MIB-producing strains and 23 sequences from the NCBI database. This primer set successfully identified all known 43 MIB-producing cyanobacterial strains (12 from this study and 31 from the NCBI database), belonging to different genera, showing a wider coverage than previous primer sets. The efficiency of the method was proved by the amplification efficiency (E = 91.23%), $R^2$ of the standard curve (0.999), the limit of detection (LOD, 5.7 fg μL$^{−1}$), and the limit of quantification (LOQ, $1.86\times 10^4$ gene copies μL$^{−1}$). Further, the method was verified by the correlation between the mic gene abundance and MIB concen- tration 50 field samples from different reservoirs ($R^2 = 0.614$, p < 0.001) and one reservoir ($R^2 = 0.752$, p < 0.001), suggesting its potential as an alternative warning tool to evaluate the risk of MIB problems in source water.