Assessment of 16S rRNA gene primers for studying bacterial community structure and function of aging flue-cured tobaccos

Selection of optimal primer pairs in 16S rRNA gene sequencing is a pivotal issue in microorganism diversity analysis. However, limited effort has been put into investigation of specific primer sets for analysis of the bacterial diversity of aging flue-cured tobaccos (AFTs), as well as prediction of the function of the bacterial community. In this study, the performance of four primer pairs in determining bacterial community structure based on 16S rRNA gene sequences in AFTs was assessed, and the functions of genes were predicted using Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt). Results revealed that the primer set 799F-1193R covering the amplification region V5V6V7 gave a more accurate picture of the bacterial community structure of AFTs, with lower co-amplification levels of chloroplast and mitochondrial genes, and more genera covered than when using the other primers. In addition, functional gene prediction suggested that the microbiome of AFTs was involved in kinds of interested pathways. A high abundance of functional genes involved in nitrogen metabolism was detected in AFTs, reflecting a high level of bacteria involved in degrading harmful nitrogen compounds and generating nitrogenous nutrients for others. Additionally, the functional genes involved in biosynthesis of valuable metabolites and degradation of toxic compounds provided information that the AFTs possess a huge library of microorganisms and genes that could be applied to further studies. All of these findings provide a significance reference for researchers working on the bacterial diversity assessment of tobacco-related samples.