Evaluation of Bacterial Viability for Fecal Microbiota Transplantation: Impact of Thawing Temperature and Storage Time

Abstract

Fecal Microbiota Transplantation (FMT) represents a promising therapeutic tool under study for several purposes and is currently applied to the treatment of recurrent Clostridioides difficile infection. However, since the use of fresh stool was affected by several issues linked to donor screening, the development of a frozen stool bank is a reliable option to standardize FMT procedures. Nevertheless, different environmental factors impact microbial viability. Herein, we report the effect of different thawing temperatures and storage conditions on bacterial suspensions in the FMT procedure. In total, 20 stool samples were divided into aliquots and tested across a combination of different storing periods (15, 30; 90 days) and thawing procedures (4 °C overnight, room temperature for 1 h; 37 °C for 5 min). Focusing on storage time, our data showed a significant reduction in viability for aerobic and anaerobic bacteria after thawing for 15 days, while no further reductions were observed until after 90 days. Instead, among the different thawing procedures, no significant differences were observed for aerobic bacteria, while for anaerobes, thawing at 37 °C for 5 min was more effective in preserving the bacterial viability. In conclusion, the frozen fecal microbiota remained viable for at least three months, with an excellent recovery rate in all three thawing conditions.

Keywords: Clostridiodes difficile infection; Fecal Microbiota Transplantation (FMT); bacterial viability; stool bank.

Figure 1

Analysis of overall microbial load for the different thawing and storing conditions for aerobic and anaerobic bacteria.