Evaluation of an optimal preparation of human standardized fecal inocula for in vitro fermentation studies

M. Aguirre, A. Eck, M.E. Koenen, P.H.M. Savelkoul, A.E. Budding, K. Venema

Research output: Contribution to journalArticleAcademicpeer-review

6 Citations (Scopus)

Abstract

This study investigated the optimal preservation approach to prepare human feces as inoculum for in vitro fermentations as an alternative to the use of fresh feces. The four treatments studied were: Treatment 1) fresh feces resuspended in dialysate solution + glycerol; Treatment 2) fresh feces resuspended in dialysate solution + glycerol and then stored at -80 degrees C; Treatment 3) fecal sample frozen with 15 g glycerol; and Treatment 4) fecal sample frozen. All the treatments contained 8.75 g of feces, 3.5 ml dialysate and 4.9 ml glycerol when inoculated in TIM-2 in vitro system. Treatment 1 (fresh fecal preparation) was used as a reference. The effects were evaluated in terms of i) metabolic activity and ii) composition of the microbiota using fermentation experiments in the TIM-2 in vitro system. In all treatments, high levels of acetate were produced followed by n-butyrate and propionate. However, the metabolic activity of the bacteria, in terms of short-chain fatty acid production, was affected by the different treatments. Microbiota composition was analyzed using the IS-pro profiling technique. Diversity in Actinobacteria, Firmicutes, Fusobacteria and Verrucomicrobia and Proteobacteria groups seemed to be preserved in all treatments whereas. it was observed to decline in the Bacteroidetes group. Preparing a human fecal inoculum resuspended in dialysate solution with glycerol and then stored at 80 degrees C showed high similarities to the results obtained with fresh feces, and is proposed as the optimal way to freeze fecal material as an alternative to fresh feces for in vitro fermentation studies. 

Original languageEnglish
Pages (from-to)78-84
Number of pages7
JournalJournal of Microbiological Methods
Volume117
DOIs
Publication statusPublished - Oct 2015

Keywords

  • Cryopreservation
  • Gut microbiota
  • In vitro model
  • Inoculum preparation
  • LARGE-INTESTINE
  • MICROBIAL DIVERSITY
  • GUT MICROBIOTA
  • FATTY-ACIDS
  • STORAGE
  • BACTERIA
  • PRODUCTS
  • MODEL
  • COLON
  • FECES

Cite this

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title = "Evaluation of an optimal preparation of human standardized fecal inocula for in vitro fermentation studies",
abstract = "This study investigated the optimal preservation approach to prepare human feces as inoculum for in vitro fermentations as an alternative to the use of fresh feces. The four treatments studied were: Treatment 1) fresh feces resuspended in dialysate solution + glycerol; Treatment 2) fresh feces resuspended in dialysate solution + glycerol and then stored at -80 degrees C; Treatment 3) fecal sample frozen with 15 g glycerol; and Treatment 4) fecal sample frozen. All the treatments contained 8.75 g of feces, 3.5 ml dialysate and 4.9 ml glycerol when inoculated in TIM-2 in vitro system. Treatment 1 (fresh fecal preparation) was used as a reference. The effects were evaluated in terms of i) metabolic activity and ii) composition of the microbiota using fermentation experiments in the TIM-2 in vitro system. In all treatments, high levels of acetate were produced followed by n-butyrate and propionate. However, the metabolic activity of the bacteria, in terms of short-chain fatty acid production, was affected by the different treatments. Microbiota composition was analyzed using the IS-pro profiling technique. Diversity in Actinobacteria, Firmicutes, Fusobacteria and Verrucomicrobia and Proteobacteria groups seemed to be preserved in all treatments whereas. it was observed to decline in the Bacteroidetes group. Preparing a human fecal inoculum resuspended in dialysate solution with glycerol and then stored at 80 degrees C showed high similarities to the results obtained with fresh feces, and is proposed as the optimal way to freeze fecal material as an alternative to fresh feces for in vitro fermentation studies. ",
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author = "M. Aguirre and A. Eck and M.E. Koenen and P.H.M. Savelkoul and A.E. Budding and K. Venema",
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pages = "78--84",
journal = "Journal of Microbiological Methods",
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Evaluation of an optimal preparation of human standardized fecal inocula for in vitro fermentation studies. / Aguirre, M.; Eck, A.; Koenen, M.E.; Savelkoul, P.H.M.; Budding, A.E.; Venema, K.

In: Journal of Microbiological Methods, Vol. 117, 10.2015, p. 78-84.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Evaluation of an optimal preparation of human standardized fecal inocula for in vitro fermentation studies

AU - Aguirre, M.

AU - Eck, A.

AU - Koenen, M.E.

AU - Savelkoul, P.H.M.

AU - Budding, A.E.

AU - Venema, K.

PY - 2015/10

Y1 - 2015/10

N2 - This study investigated the optimal preservation approach to prepare human feces as inoculum for in vitro fermentations as an alternative to the use of fresh feces. The four treatments studied were: Treatment 1) fresh feces resuspended in dialysate solution + glycerol; Treatment 2) fresh feces resuspended in dialysate solution + glycerol and then stored at -80 degrees C; Treatment 3) fecal sample frozen with 15 g glycerol; and Treatment 4) fecal sample frozen. All the treatments contained 8.75 g of feces, 3.5 ml dialysate and 4.9 ml glycerol when inoculated in TIM-2 in vitro system. Treatment 1 (fresh fecal preparation) was used as a reference. The effects were evaluated in terms of i) metabolic activity and ii) composition of the microbiota using fermentation experiments in the TIM-2 in vitro system. In all treatments, high levels of acetate were produced followed by n-butyrate and propionate. However, the metabolic activity of the bacteria, in terms of short-chain fatty acid production, was affected by the different treatments. Microbiota composition was analyzed using the IS-pro profiling technique. Diversity in Actinobacteria, Firmicutes, Fusobacteria and Verrucomicrobia and Proteobacteria groups seemed to be preserved in all treatments whereas. it was observed to decline in the Bacteroidetes group. Preparing a human fecal inoculum resuspended in dialysate solution with glycerol and then stored at 80 degrees C showed high similarities to the results obtained with fresh feces, and is proposed as the optimal way to freeze fecal material as an alternative to fresh feces for in vitro fermentation studies. 

AB - This study investigated the optimal preservation approach to prepare human feces as inoculum for in vitro fermentations as an alternative to the use of fresh feces. The four treatments studied were: Treatment 1) fresh feces resuspended in dialysate solution + glycerol; Treatment 2) fresh feces resuspended in dialysate solution + glycerol and then stored at -80 degrees C; Treatment 3) fecal sample frozen with 15 g glycerol; and Treatment 4) fecal sample frozen. All the treatments contained 8.75 g of feces, 3.5 ml dialysate and 4.9 ml glycerol when inoculated in TIM-2 in vitro system. Treatment 1 (fresh fecal preparation) was used as a reference. The effects were evaluated in terms of i) metabolic activity and ii) composition of the microbiota using fermentation experiments in the TIM-2 in vitro system. In all treatments, high levels of acetate were produced followed by n-butyrate and propionate. However, the metabolic activity of the bacteria, in terms of short-chain fatty acid production, was affected by the different treatments. Microbiota composition was analyzed using the IS-pro profiling technique. Diversity in Actinobacteria, Firmicutes, Fusobacteria and Verrucomicrobia and Proteobacteria groups seemed to be preserved in all treatments whereas. it was observed to decline in the Bacteroidetes group. Preparing a human fecal inoculum resuspended in dialysate solution with glycerol and then stored at 80 degrees C showed high similarities to the results obtained with fresh feces, and is proposed as the optimal way to freeze fecal material as an alternative to fresh feces for in vitro fermentation studies. 

KW - Cryopreservation

KW - Gut microbiota

KW - In vitro model

KW - Inoculum preparation

KW - LARGE-INTESTINE

KW - MICROBIAL DIVERSITY

KW - GUT MICROBIOTA

KW - FATTY-ACIDS

KW - STORAGE

KW - BACTERIA

KW - PRODUCTS

KW - MODEL

KW - COLON

KW - FECES

U2 - 10.1016/j.mimet.2015.07.019

DO - 10.1016/j.mimet.2015.07.019

M3 - Article

VL - 117

SP - 78

EP - 84

JO - Journal of Microbiological Methods

JF - Journal of Microbiological Methods

SN - 0167-7012

ER -