Eukaryotic Cell Capture by Amplified Magnetic in situ Hybridization Using Yeast as a Model.

Fabiola Bastian,Laurence Fraissinet-Tachet,Pascal Simonet,Mylène Hugoni,Nora M Dempsey,Marie Frenea-Robin,Delphine Melayah

doi:10.3389/fmicb.2021.759478

Abstract

A non-destructive approach based on magnetic in situ hybridization (MISH) and hybridization chain reaction (HCR) for the specific capture of eukaryotic cells has been developed. As a prerequisite, a HCR-MISH procedure initially used for tracking bacterial cells was here adapted for the first time to target eukaryotic cells using a universal eukaryotic probe, Euk-516R. Following labeling with superparamagnetic nanoparticles, cells from the model eukaryotic microorganism Saccharomyces cerevisiae were hybridized and isolated on a micro-magnet array. In addition, the eukaryotic cells were successfully targeted in an artificial mixture comprising bacterial cells, thus providing evidence that HCR-MISH is a promising technology to use for specific microeukaryote capture in complex microbial communities allowing their further morphological characterization. This new study opens great opportunities in ecological sciences, thus allowing the detection of specific cells in more complex cellular mixtures in the near future.

Highlights

In recent years, research in microbial ecology has truly taken off
As a recently developed method, Fluorescent in situ DNAhybridization chain reaction (HCR-Fluorescent In Situ Hybridization (FISH)) may offer the opportunity to overcome the main problem of FISH, i.e., low intensity of the signal, due to low rRNA content found in some environmental microorganisms (Yamaguchi et al, 2015; Jia et al, 2021)
Procedure, is a non-degenerated 16 bp-long eukaryotic universal sequence and corresponds with 100% homology to the antisense of the S. cerevisiae 18S rRNA gene sequence. This sequence has already been used as one of the eukaryotic universal primer pairs in several eukaryotic microorganism diversity studies conducted in different environments, especially in soils (Bailly et al, 2007; Damon et al, 2012; Lehembre et al, 2013)

Summary

Introduction

Research in microbial ecology has truly taken off. This spectacular breakthrough is mainly due to rapid technological advances such as meta-omics, which have significantly increased our ability to study microbial communities from complex environments and their function in various ecosystems (Morris et al, 2002; Hiraoka et al, 2016; Grumaz et al, 2020; Sehnal et al, 2021). As a recently developed method, Fluorescent in situ DNAhybridization chain reaction (HCR-FISH) may offer the opportunity to overcome the main problem of FISH, i.e., low intensity of the signal, due to low rRNA content found in some environmental microorganisms (Yamaguchi et al, 2015; Jia et al, 2021). Unicellular microorganisms cannot be directly isolated from soils or sediments due to the presence of many mineral and calcareous impurities present in these environments

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