Development of qPCR assays for bacterial nitrification and denitrification genes in catfish aquaculture ponds.

Nitrogenous waste products are toxic to fish, and their removal is a critical process in aquaculture production. In earthen pond production systems, such as those used in the catfish industry, phytoplankton act as the dominant sink for ammonia; however, bacteria can also play roles in denitrification and nitrification, particularly when excess ammonia exists. As the US catfish industry continues to intensify, the bacterial communities relevant in the removal of nitrogenous waste will become more integral to efficient production and necessitate further research. Here, quantitative PCR (qPCR) assays targeting four genes covering the denitrification and nitrification pathways present in catfish aquaculture ponds were developed. Twenty-four existing primer sets were used to amplify relevant genes in samples obtained from catfish pond water and sediment and then subjected to high-throughput sequencing to identify the sequence variants present in the environment. Five conventional PCR assays yielded sequencing results conducive to qPCR primer design. Quantitative PCR assays were successfully developed for four targets: