Arantes Julia A, Di Pietro Rebecca, Ratté Mélanie, Arroyo Luis G, Leclère Mathilde, Costa Marcio C
Department of Veterinary Medicine/Faculty of Animal Science and Food Engineering, Universidade de São Paulo, Pirassununga, São Paulo, Brazil.
Department of Veterinary Biomedical Sciences/Faculty of Veterinary Medicine, University of Montreal, Saint Hyacinthe, QC, Canada.
PeerJ. 2025 Feb 20;13:e18860. doi: 10.7717/peerj.18860. eCollection 2025.
Fecal microbiota transplantation (FMT) has been used as a treatment option for horses () with gastrointestinal diseases. Several preparation and conservation protocols to improve bacterial survival have been studied in other species.
This study aimed to evaluate the impact of oxygen exposure and different protectant solutions on bacterial viability before and after freezing using horse feces. Fecal samples from 10 healthy horses were aliquoted and diluted in cryoprotectant solutions containing antioxidants ( = 40) or 10% glycerol ( = 40). Half of the aliquots from each dilution condition were prepared inside an anaerobic chamber, while the other half were prepared under ambient air conditions. Each sample was also analyzed fresh and after freezing at -20 °C for 90 days. Bacterial viability was assessed using flow cytometry. A mixed linear model and the Friedman and Wilcoxon tests were used depending on data distribution.
Freeze-thawing decreased bacterial viability by 47% (mean ± SD: 51 ± 27% before, 27 ± 8% after; < 0.001). Glycerol was superior to the cryoprotectant after freezing (32 ± 8% glycerol, 24 ± 8% cryoprotectant; < 0.001). Oxygen exposure did not affect viability ( = 0.13). There was no statistical difference between protectant solutions in fresh samples ( = 0.16).
Fresh FMT solutions may be better for treating horses with dysbiosis, but if freezing cannot be avoided, glycerol should be used to dilute feces.
粪便微生物群移植（FMT）已被用作治疗马匹胃肠道疾病的一种选择。在其他物种中，已经研究了几种提高细菌存活率的制备和保存方案。
本研究旨在评估氧气暴露和不同保护剂溶液对马粪便冷冻前后细菌活力的影响。将10匹健康马的粪便样本进行分装，并在含有抗氧化剂（n = 40）或10%甘油（n = 40）的冷冻保护剂溶液中稀释。每种稀释条件下的一半分装样本在厌氧箱内制备，另一半在环境空气条件下制备。每个样本还在新鲜时以及在-20°C冷冻90天后进行分析。使用流式细胞术评估细菌活力。根据数据分布使用混合线性模型以及弗里德曼检验和威尔科克森检验。
冻融使细菌活力降低了47%（平均值±标准差：之前为51±27%，之后为27±8%；P < 0.001）。冷冻后甘油优于冷冻保护剂（甘油为32±8%，冷冻保护剂为24±8%；P < 0.001）。氧气暴露不影响活力（P = 0.13）。新鲜样本中保护剂溶液之间无统计学差异（P = 0.16）。
新鲜的FMT溶液可能更适合治疗患有菌群失调的马匹，但如果无法避免冷冻，应使用甘油稀释粪便。
