Efficacy of a multistrain synbiotic treatment in post-acute COVID-19 patients: A double-blind, Placebo-Controlled Randomized trial
Fecal samples from study participants were collected through fecal swabs. DNA samples isolated from patients' fecal swabs were obtained with Nimbus automatic extractor (Seegene Technologies, Korea) starting from 500 µl of sample following the manufacturer's protocol after treatment with 500 µl of Lysis Buffer ATL (QIAGEN, Hilden, Germany) at 56°C for 10 min with 20 µl Proteinase K (Darmstadt, Germany).   The V3–V4 hypervariable region of the bacterial 16S  rRNA gene was amplified from total DNA according to the Illumina 16S Metagenomic Sequencing Library Preparation instructions, as previously described in Diamanti-Picchianti A, et al (Int J Mol Med ). In summary, the V3-V4 amplicon was obtained by PCR with universal primers, followed by purification. A second PCR was then performed to barcode the libraries using the Illumina dual-index system. Following a second step of purification, libraries were diluted to 4nM and pooled.