Preservation of Human Gut Microbiota Inoculums for In Vitro Fermentations Studies

Carvalho, Nelson Mota de; Oliveira, Diana; Saleh, Mayra Anton Dib; Pintado, Manuela; Madureira, Ana Raquel

doi:10.3390/fermentation7010014

Cited by 19 publications

(16 citation statements)

References 53 publications

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“…In this study, the aerobic as well as anaerobic cell count reached the same level as before the freeze-drying preservation step. This may be due to different cultivation conditions in the study of Carvalho et al (2021) and this study that resulted in lower cell counts. Further, during freezing, intracellular ice can be formed and the increase in solute concentration imposes osmotic stress to the cell (Santivarangkna et al, 2008).…”

Section: Re-establishment Of An In Vitro System After Dryingmentioning

confidence: 80%

“…Compared with other studies, when stool samples were preserved by freezing, the cell count irreversibly dropped about one log CFU ml −1 and could not reach the same level as before preservation (Carvalho et al ., 2021 ). In this study, the aerobic as well as anaerobic cell count reached the same level as before the freeze–drying preservation step.…”

Section: Resultsmentioning

confidence: 99%

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Preservation by lyophilization of a human intestinal microbiota: influence of the cultivation pH on the drying outcome and re‐establishment ability

Haindl

Totzauer

Kulozik

2022

Microbial Biotechnology

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Faecal microbiota transplantation is an emerging medical concept for the treatment of gastrointestinal diseases. This concept, however, has disadvantages as low storability of stool and intensive donor screening. A solution to overcome these problems would be the preservation of an in vitro microbiota through freeze-drying. However, the influence of the entire preservation process, including cultivation and lyophilization, has not been assessed so far. In this study, the influences of the process steps cultivation, drying and re-cultivation were determined with cell count, production of metabolites, microbial composition and diversity in the system as evaluation criteria. All pH conditions resulted in stable, culturable communities after re-cultivation. Cell count, richness, diversity and microbial composition were affected by freeze-drying, but these effects were reversible and vanished during re-cultivation. Hence, the re-cultivated system did not differ from the system before drying. The metabolism, measured by short-chain fatty acids as indicators, showed slight changes due to natural dynamics. Consequently, the cultivation prior to drying was identified to have more influence than the drying itself on the preservation process and therefore the biggest potential for optimization. Hence, the highest similarity with the initial stool sample was obtained with pH 6.0 -6.5 during cultivation.

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Section: Re-establishment Of An In Vitro System After Dryingmentioning

confidence: 80%

Section: Resultsmentioning

confidence: 99%

Preservation by lyophilization of a human intestinal microbiota: influence of the cultivation pH on the drying outcome and re‐establishment ability

Haindl

Totzauer

Kulozik

2022

Microbial Biotechnology

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“…Biobanked stools stored in the presence of preservation chemicals, such as glycerol, are commonly used for either microbiome assay, fermentation or FMT applications. 13 , 17 , 18 Using the bacterial cell counting approach, Guerin-Danan et al . demonstrated that freezing in glycerol buffer had no effect on aerobes in fecal samples, while decreased the survival of anaerobes, but did not exceed the inter-individual variations.…”

Section: Introductionmentioning

confidence: 99%

Evaluating live microbiota biobanking using an ex vivo microbiome assay and metaproteomics

Zhang

Mayne

et al. 2022

Gut Microbes

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Biobanking of live microbiota is becoming indispensable for mechanistic and clinical investigations of drug–microbiome interactions and fecal microbiota transplantation. However, there is a lack of methods to rapidly and systematically evaluate whether the biobanked microbiota maintains their cultivability and functional activity. In this study, we use a rapid ex vivo microbiome assay and metaproteomics to evaluate the cultivability and the functional responses of biobanked microbiota to treatment with a prebiotic (fructo-oligosaccharide, FOS). Our results indicate that the microbiota cultivability and their functional responses to FOS treatment were well maintained by freezing in a deoxygenated glycerol buffer at −80°C for 12 months. We also demonstrate that the fecal microbiota is functionally stable for 48 hours on ice in a deoxygenated glycerol buffer, allowing off-site fecal sample collection and shipping to laboratory for live microbiota biobanking. This study provides a method for rapid evaluation of the cultivability of biobanked live microbiota. Our results show minimal detrimental influences of long-term freezing in deoxygenated glycerol buffer on the cultivability of fecal microbiota.

show abstract

Section: Introductionmentioning

confidence: 99%

“…Biobanked stools stored in the presence of preservation chemicals, such as glycerol, are commonly used for either microbiome assay, fermentation or FMT applications 13,17,18 . Using the bacterial cell counting approach, Guerin-Danan et al demonstrated that freezing in glycerol buffer had no effect on aerobes in fecal samples, while decreased the survival of anaerobes, but did not exceed the inter-individual variations 19 .…”

Section: Introductionmentioning

confidence: 99%

Evaluating live microbiota biobanking using an ex vivo microbiome assay and metaproteomics

Zhang¹,

Mayne²,

L³

et al. 2021

Preprint

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Biobanking of live microbiota is becoming indispensable for mechanistic and clinical investigations of drug-microbiome interactions and fecal microbiota transplantation. However, there is a lack of methods to rapidly and systematically evaluate whether the biobanked microbiota maintains their cultivability and functional activity. In this study, we use a rapid ex vivo microbiome assay and metaproteomics to evaluate the cultivability and the functional responses of biobanked microbiota to treatment with a prebiotic (fructo-oligosaccharide, FOS). Our results indicate that the microbiota cultivability and their functional responses to FOS treatment were well maintained by freezing in a deoxygenated glycerol buffer at -80°C for 12 months. We also demonstrate that the fecal microbiota is functionally stable for 48 hours on ice in a deoxygenated glycerol buffer, allowing off-site fecal sample collection and shipping to laboratory for live microbiota biobanking. This study provides a method for rapid evaluation of the cultivability of biobanked live microbiota. Our results show minimal detrimental influences of long-term freezing in deoxygenated glycerol buffer on the cultivability of fecal microbiota.

show abstract

Preservation of Human Gut Microbiota Inoculums for In Vitro Fermentations Studies

Cited by 19 publications

References 53 publications

Preservation by lyophilization of a human intestinal microbiota: influence of the cultivation pH on the drying outcome and re‐establishment ability

Preservation by lyophilization of a human intestinal microbiota: influence of the cultivation pH on the drying outcome and re‐establishment ability

Evaluating live microbiota biobanking using an ex vivo microbiome assay and metaproteomics

Evaluating live microbiota biobanking using an ex vivo microbiome assay and metaproteomics

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