Impact of optimised sample preparation method on DNA recovery from subgingival plaque in a population with periodontitis

Abstract

Objective(s)

This study aimed to evaluate whether an alternate sample preparation method could enhance DNA yield from subgingival plaque samples collected from patients with untreated periodontitis.

Design

Fifty-six consecutive participants with untreated periodontitis each provided two subgingival plaque samples from periodontal sites with probing pocket depth (PPD) > 5 mm. In the alternate method (AM), 1.4 mm ceramic beads were added to the sample prior to supernatant removal to protect the microbial pellet, after which samples were processed using a standardized extraction protocol and subjected to 16S rRNA gene sequencing (V3–V4 region).

Results

A total of 112 samples were analysed. DNA concentration (ng/µl) was significantly higher in the AM group (23.82 ± 23.31) compared to the standard method (SM) group (13.6 ± 17.07; p < 0.001). The DNA input in 2.5 µl was also significantly greater in the AM group (1156 ± 1139 ng vs. 277.96 ± 273.12 ng; p < 0.001). Shannon diversity was significantly higher in AM than SM (p = 6e–04). No significant differences in DNA yield or microbial diversity were observed between the two periodontal sites within each group.

Conclusion

The alternate sample preparation method improved DNA concentration compared to standard sample preparation method of sub-gingival plaque analysis collected from patients with untreated periodontitis.