The microbiome has a profound impact on host fitness. pH, oxygen, nutrients, or other factors such as food or pharmaceuticals, subject the microbiome to variations in the gastrointestinal tract. This variation is a cause for concern given dysbiosis of the microbiome is correlated with various disease states. Currently, much research relies on model organisms to study microbial communities since intact microbiomes are challenging to utilize. The objective of this study is to culture an explanted colon microbiome of 4 Balb/c mice to develop an in vitro tool for future microbiome studies. We cultured homogenates of the distal colons of 4 mice in trans-well culture dishes. These dishes were incubated for 24 hours in two different oxygen concentration levels and the pH was compared before and after incubation of the cultures. To analyze the integrity of the microbiome, we utilized massively paralleled DNA sequencing with 16S metagenomics to characterize fecal and colon samples to speculate whether future studies may utilize feces in constructing an in vitro microbial community to spare animal lives. We found that pH and familial relationships had a profound impact on community structure while oxygen did not have a significant influence. The feces and the colon were similar in community profiles, which lends credence to utilizing feces in future studies. The gut microbiome is of great interest and great importance for studies in a variety of different diseases. Many laboratories do not have access to germ-free mice, which is one optimal way to study mammalian microbiomes, but this technique allowed for the in vitro culturing of a majority of the prokaryotes isolated from the colons of mice. This may allow an alternative to study the interactions of this very diverse population of microorganisms without the need for germ-free conditions.
Fluctuations in oxygen, pH, nutrients, or other factors such as food or pharmaceuticals, may perturb the microbiota of the gastrointestinal (GI) tract. This environmental variation is a cause for concern given dysbiosis of the microbiome is correlated with disease states; thereby, model organisms are utilized to study microbial communities during, after, or before shifts in microbes since intact ex vivo microbiomes have historically been challenging to utilize. The objective of this study is to culture an explant microbiome of 4 Balb/c, laboratory bred mice to develop an ex vivo tool for future microbiome studies. We cultured homogenates of the distal colon of 4 mice in three dimensional, 24 well plate culture dishes. These dishes were incubated for 24 hours in two different oxygen concentration levels, 0% and 20%. The pH of the plate was tested before and after incubation. To analyze the integrity of the microbiome, we utilized 16S sequencing. Further, we utilized 16S metagenomics to characterize fecal samples and colon samples to speculate whether future studies may utilize feces in constructing an explant microbiome to spare animal lives. We found that pH and familial relationship had a profound impact on community structure while oxygen did not have a significant influence. The feces and the colon were similar in community profiles, which lends credence to utilizing feces in future studies. In addition, our efforts successfully cultured archaea, which included difficult to culture strains such as Miscellaneous Crenarchaeota group (MCG) and Methanobacteria. Ultimately, further attempts to culture and preserve an animal’s microbiome needs to control for and maintain stable pH.
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