PMA-qPCR: ACCELERATING THE MARKET RELEASE OF HIGH-QUALITY BRADYRHIZOBIUM DIAZOEFFICIENS INOCULANTS

Abstract

The quantification of Bradyrhizobium diazoefficiens in inoculants traditionally relies on culture-based assays, which are labor-intensive and time-consuming. To address this limitation, we validated a PMA-qPCR assay as a rapid and reliable alternative for estimating viable Bradyrhizobium counts. The assay demonstrated strong performance, achieving approximately 95% efficiency, a standard deviation of 0.3 log CFU/ml, and an intra-assay reproducibility with a coefficient of variation less than 10%. Key experiments optimized PMA concentration to ensure selective exclusion of non-viable cells without compromising viable cell quantification. Discrimination threshold assessment confirmed the assays ability to differentiate quarter-strength dilutions. Final validation against plate counting revealed an 82% correlation, significantly reducing processing time from 120 hours to just 5 hours. This is the first study to apply PMA-qPCR specifically for Bradyrhizobium diazoefficiens quantification in inoculants. The results highlight its potential as a high-throughput tool for microbial viability assessment, offering improvements in efficiency and precision for batch-to-batch quality control in industrial applications.