Mimicking early Batrachochytrium dendrobatidis-host interactions using A6 cells

Scope of the method

Alternative method relates to
  • Animal health
Alternative method is situated in
  • Basic Research
Type of alternative method
  • In vitro - Ex vivo
This method makes use of
  • Animal derived cells / tissues / organs
Species from which cells/tissues/organs are derived
Xenopus laevis
Type of cells/tissues/organs
Kidney

Description

Method keywords
  • A6 cells
  • Immunofluorescence
  • in vitro model
  • chytrid
Scientific area keywords
  • host-pathogen interaction
  • adhesion
  • invasion
  • intracellular maturation
  • fungus
  • amphibian
Method description

We describe a fluorescent cell-based in vitro infection model that reproduces host-Batrachochytrium dendrobatidis (Bd) interactions. Using the epithelial cell line A6 from Xenopus laevis, we reproduced different stages of host cell infection and intracellular growth of Bd, resulting in host cell death, a key event in chytridiomycosis. The presented in vitro models may facilitate future mechanistic studies of host susceptibility and pathogen virulence.

Lab equipment
  • Biosafety cabinet ;
  • CO2 incubator at 26°C ;
  • Fluorescent microscope.
Method status
  • Published in peer reviewed journal

Pros, cons & Future potential

Advantages

To date, infectivity and the pathogenicity of Bd have mostly been studied using light microscopy (LM), scanning electron microscopy (SEM) and transmission electron microscopy (TEM) on in vivo-infected skin tissues or ex vivo-infected skin explants. We now established a cell-based assay that mimics the colonization stages of Bd in vitro (adhesion, germ tube development, penetration into skin cells, invasive growth and the induction of host cell death), allowing rapid and efficient screening of host-Bd interactions and reducing the number of animals used in infection trials.

Challenges

This is and stays an in vitro model that mimics the in vivo situation. Caution should always be exercised when extrapolating in vitro data to the in vivo situation, but in vitro cell culture models allow an experimental flexibility making them highly suitable to study host-pathogen interactions.

Modifications

The method is optimized for the fungal pathogen Batrachochytrium dendrobatidis. Expanding this to other pathogens needs further optimization.

Future & Other applications

The method can be used for a wide range of applications. The availability of an in vitro model using a continuous cell line may, for example, be used to analyze the differences in host-pathogen interactions between different Bd strains. Also, expression patterns during different infection steps can be examined.

References, associated documents and other information

References

Elin Verbrugghe, Pascale Van Rooij, Herman Favoreel, An Martel, Frank Pasmans (2019) In vitro modeling of Batrachochytrium dendrobatidis infection of the amphibian skin. PLoS ONE 14(11): e0225224. https://doi.org/10.1371/journal.pone.0225224

Associated documents
journal.pone.0225224.pdf

Contact person

Elin Verbrugghe

Organisations

Ghent University
Department of Pathology, Bacteriology and avian diseases
Belgium
Flemish Region