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Influence of DNA extraction methods and 16S rRNA targeted hypervariable regions on microbial diversity profiling of dental plaque and saliva

Sree Sankar Darveekaran Nair^a,b, Pengfei Zhu^a, Yogendra Bhaskar^a,b, Fei Teng^a,c,*

 
ABSTRACT:     The application of high-throughput sequencing has revolutionized our ability to investigate the microbiota composition of complex oral ecosystems. One such approach is through 16S rRNA sequencing. Deciphering this diverse microbial community using such approach is more accurate than traditional culture-based methods. Numerous factors such as primer selection, DNA extraction methods, and sequencing platforms may compromise the results. Therefore, these factors need to be considered carefully for studies such as microbial diversity analysis and microbial abundance determination. Here, we evaluated the effect of two DNA extraction methods (soil and swab DNA extraction kits) and targeted 16S rRNA hypervariable regions (V1–V3 and V3–V4 dog pads) on plaque and saliva sample types to investigate the diverse microbes in oral samples of five Chinese citizens. Both of the extraction methods were successful in determining the relative abundance of microbes; however, relative abundance percentage of the species reported by the soil extraction method with the hypervariable region V1–V3 is higher in both sample types. The relative taxonomical abundance of microbes at OTU level of species represented by the V3–V4 region is lower; nonetheless, the number of unshared species identified is higher using this region. Both of the hypervariable regions identified unshared species in plaque and saliva sample types. Thus, choice of DNA extraction methods and hypervariable regions greatly influence the representation of oral microbial diversity in plaque and saliva. This study provides a guide to future studies to portray and quantify the human oral microbiota.

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* Corresponding author, E-mail: tengfei@qibebt.ac.cn

Received 7 Jun 2020, Accepted 3 May 2021