Optimization and standardization of the culturomics technique for human microbiome exploration

Abstract

Culturomics is a high-throughput culture approach that has dramatically contributed to the recent renewal of culture. While metagenomics enabled substantial advances in exploring the microbiota, culturomics significantly expanded our knowledge regarding the bacterial gut repertoire through the discovery and the description of hundreds of new taxa. While this approach relies on the variation of culture conditions and media, we have tested so far more than 300 conditions since the beginning of culturomics studies. In this context, we aimed herein to identify the most profitable conditions for optimizing culturomics approach. For this purpose, we have analysed a set of 58 culturomics conditions that were previously applied to 8 faecal specimens, enabling the isolation of 497 bacterial species. As a result, we were able to reduce the number of conditions used to isolate these 497 of more than a half (i.e. to 25 culture conditions). We have also established a list of the 16 conditions that allowed to capture 98% of the total number of species previously isolated. These data constitute a methodological starting point for culture-based microbiota studies by improving the culturomics workflow without any loss of captured bacterial diversity.

Figure 1

Phylogenetic tree based on the available 16S rRNA gene sequences of 480 bacterial species of this study. This image was generated thanks to the itol platform (https://itol.embl.de/).

Figure 2

Comparison of the number of bacterial species isolated by the 58 conditions in blue VS the 25 optimized conditions in yellow.

Figure 3

Comparison of the bacterial species in each culture condition group of this study with the list of species isolated in the human gut by culturomics, established by Lagier et.al. In yellow the group of 22 alcohol conditions, in green the group of 18 new conditions, in blue the group of 18 previously selected conditions and in pink the species isolated in the study of Lagier et.al.

Figure 4

Comparison of bacterial species found in 7 conditions without rumen in blue (Hemoc Ana 37 °C, HS Ae 37 °C, HS Ana 37 °C, HS alcohol Ae 37 °C, HS alcohol Ae 28 °C, HS alcohol Ana 28 °C) against 7 other corresponding conditions with rumen in yellow (HR Ana 37 °C, HRS Ae 37 °C, HRS Ana 37 °C, HRS alcohol Ae 37 °C, HRS alcohol Ana 37 °C, HRS alcohol Ana 37 °C, HRS alcohol Ae 28 °C, HRS alcohol Ana 28 °C).