16S rRNA bioinformatic analysis
The 16S rRNA analysis workflow is shown in Figure 2. The results from
the next-generation sequencing were in the FAST5 format. The FastQ files
were generated from the results in the FAST5 format by using the Guppy
program in MinKNOW (Oxford Nanopore Technologies, UK), in which
real-time quality scores of more than 7 were filtered and demultiplexed.
The FastQ passing quality was aligned to reference sequences in the NCBI
database by using the FASTQ 16S workflow (version 2022.01.07), and the
inclusion criteria were a minimum BLAST e-value of 0.01, minimum
coverage of more than 30%, and a minimum identity of more than 77%.
The data output files were generated as CSV files. Then, the percentage
prevalence of each species and genus were calculated, and species and
genus were filtered at a level of 0.5% for investigation of relative
abundance.
Comparative taxonomy was performed to investigate the relative abundance
of species between neonates delivered vaginally and those delivered by
cesarean delivery.