Isolation and Characterization of the Natural Microbiota of the Model Nematode Caenorhabditis elegans

Summary

Caenorhabditis elegans is one of the main model species in biology, yet almost all research is performed in the absence of its naturally associated microbes. The methods described here will help to improve our understanding of the diversity of associated microbes as a basis for future functional C. elegans research.

Abstract

The nematode Caenorhabditis elegans interacts with a large diversity of microorganisms in nature. In general, C. elegans is commonly found in rotten plant matter, especially rotten fruits like apples or on compost heaps. It is also associated with certain invertebrate hosts such as slugs and woodlice. These habitats are rich in microbes, which serve as food for C. elegans and which can also persistently colonize the nematode gut. To date, the exact diversity and consistency of the native C. elegans microbiota across habitats and geographic locations is not fully understood. Here, we describe a suitable approach for isolating C. elegans from nature and characterizing the microbiota of worms. Nematodes can be easily isolated from compost material, rotting apples, slugs, or attracted by placing apples on compost heaps. The prime time for finding C. elegans in the Northern Hemisphere is from September until November. Worms can be washed out of collected substrate material by immersing the substrate in buffer solution, followed by the collection of nematodes and their transfer onto nematode growth medium or PCR buffer for subsequent analysis. We further illustrate how the samples can be used to isolate and purify the worm-associated microorganisms and to process worms for 16S ribosomal RNA analysis of microbiota community composition. Overall, the described methods may stimulate new research on the characterization of the C. elegans microbiota across habitats and geographic locations, thereby helping to obtain a comprehensive understanding of the diversity and stability of the nematode's microbiota as a basis for future functional research.

Introduction

In nature, C. elegans is commonly found in rotten plant matter, especially rotten fruits like apples or on compost heaps¹. It is also associated with certain invertebrate hosts such as slugs and woodlice^2,3. These habitats are rich in microbes, which not only serve as food for the worm, but may also form stable associations with it. Information on the diversity of naturally associated microorganisms was only published in 2016^4,5,6. Since then, these and only a few more recent studies h....

Protocol

1. Preparation of buffers and media

1. Prepare S-buffer by adding 5.85 g of NaCl, 1.123 g of K₂HPO₄, 5.926 g of KH₂PO₄, and 1 L of deionized H₂O to a flask and autoclave for 20 min at 121 °C.
2. Prepare a viscous medium by adding S-buffer containing 1.2% (w/v) hydroxymethylcellulose (the substance causing viscosity of the medium), 5 mg/mL cholesterol, 1 mM MgSO₄, 1 mM CaCl₂, and 0.1% (v/v) acetone. Autoclave and stir the viscous medium until it is completely homogeneous.
   NOTE: This can take multiple hours. Also, S-buffer can be directly prepared and added....

Results

The nematode C. elegans is frequently found in decomposing fruits, such as apples, and also compost samples. In Northern Germany, C. elegans as well as congeneric species (particularly C. remanei but also C. briggsae) are mainly found from September until November². The nematodes are most commonly found in decomposing plant matter, especially rotting fruits such as apples or pears, and also compost, particularly material that shows a high grade of decomposition........

Discussion

The nematode Caenorhabditis elegans is one of the most intensively studied model organisms in biological research. It was introduced by Sydney Brenner in the 1960s, originally for understanding the development and the function of the nervous system²⁹. Since then, C. elegans has become a powerful model for studying fundamental processes across all biological disciplines, including behavioral biology, neurobiology, aging, evolutionary biology, cell biology, developmental biology, a.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
COMSOL	COMSOL		multiphysics simulation software