Evaluation of the effects of different formulations of protectants on the preservation of the microbiota in fecal microbiota transplantation.

Abstract

Background: With the increasing indications for fecal microbiota transplantation for the treatment of diseases, there is a growing demand for the preparation of frozen or lyophilized fecal microbiota products that are viable and can stably colonize the recipient. The addition of protective agents plays an important role in the preparation. However, there has been no systematic evaluation of the protective agents used in fecal microbiota sample transplantation preparation for transplantation.

Methods: We were used the donor bacterial flora containing 10 different formulations of protective agents were frozen, lyophilized, and stored. Plate counting, CCK8 assay, flow cytometry after LIVE/DEAD staining, and fluorescence intensity were used to assess viable bacteria in vitro. In addition, the donor bacterial flora samples containing different formulations protective agents were transplanted into antibiotic-treated SPF mice, with 3 mice in each group and a total of 5 groups. Fecal samples were collected for metagenomic sequencing to observe the colonization of the bacterial flora in the recipient mice.

Results: The preliminary screening results showed that the survival rate of bacteria in the 5% trehalose (T) groups, and 5% sucrose, 5% inulin, and 1% cysteine hydrochloride (SI) groups was slightly higher than that in the other groups. SI groups tended to be more protective against anaerobes than T groups. The donor gut microbiota containing the SI groups protective agent exhibited the best colonization of the recipient mice. The protective effects of different formulations of protective agents on the colonized probiotic strains and the metabolic function of the bacterial flora in recipient mice were found to be species specific.

Conclusions: SI groups can not only better protect the activity of anaerobic bacteria in the intestine, but also effectively promote the effective colonization of donor intestinal bacteria in the recipient mice, and the effect of frozen storage method is less, and can be used at the same time as frozen and freeze-dried preparation. It can be used as a reference for the selection of protective agents in the preparation of fecal microbiota transplantation samples.