# Development of flow cytometry assay to quantify packaging of C. jejuni by Tetrahymena species

## TL;DR

A new flow cytometry method was developed to quickly measure how the waterborne pathogen Campylobacter jejuni is packaged by Tetrahymena ciliates, which may help in understanding its transmission.

## Contribution

The study introduces a novel flow cytometry approach for quantifying C. jejuni packaging by Tetrahymena species, offering a faster alternative to traditional microscopy.

## Key findings

- Cocultures of Tetrahymena with C. jejuni showed species-specific and strain-specific differences in pellet formation.
- Fluorescence intensity correlated with the number of bacteria per pellet, confirmed by TEM.
- The flow cytometry method enables efficient screening of C. jejuni strains and conditions.

## Abstract

The human pathogen Campylobacter jejuni can be packaged within multilamellar bodies (MLB), also called fecal pellets, produced by ciliates such as Tetrahymena pyriformis when these microorganisms are cocultivated. This packaging increases the survival of C. jejuni in oxygenic conditions and potentially protects it against other stressors. Traditional methods for detecting and quantifying these pellets, such as transmission electron microscopy (TEM) and fluorescence microscopy, are time-consuming and labor intensive. In this study, we devised an approach for utilizing flow cytometry to distinguish and quantify C. jejuni-containing pellets produced by both T. pyriformis and T. thermophila. Cocultures of each Tetrahymena species with four different C. jejuni strains, along with monoculture controls, were incubated for 24 h, stained with SYTO9, and analysed using flow cytometry. The results revealed ciliate species-specific and bacterial strain-specific differences in the number of pellets and their fluorescence intensity. TEM confirmed that this variability in fluorescence corresponds to differences in the number of bacteria per pellet. Our method provides a rapid and efficient means of quantifying bacteria-containing MLBs, which would facilitate the screening and comparison of a large quantity of C. jejuni strains and different conditions for studying the packaging of C. jejuni by ciliates.

A flow cytometry method was developed to measure more accurately the packaging of the water-borne pathogen Campylobacter by the ciliate Tetrahymena, which might promote its transmission by water.