Histo-FISH protocol to detect bacterial compositions and biofilms formation in vivo

Area of Science:

• Microbiology
• Gastroenterology
• Microbial Ecology

Background:

• In vivo studies of gastrointestinal tract (GIT) biofilms are limited, with in vitro methods often failing to capture microbial complexity and yielding unreliable results.
• Understanding in vivo biofilm dynamics is crucial for assessing microbial colonization and function within the gut.
• Existing methods struggle to accurately represent the in vivo environment of GIT biofilms.

Purpose of the Study:

• To develop a simple and reliable in vivo method for detecting bacterial populations within GIT biofilms.
• To standardize a novel Histo-FISH technique for visualizing Lactobacillus and Bifidobacterium species in histological slices of the GIT.
• To enable the analysis of bacterial colonization and biofilm formation in the stomach and caecum of mice.

Main Methods:

• Development and standardization of a Histo-FISH (fluorescent in situ hybridization) method using specific fluorochrome-labeled probes.
• Application of Histo-FISH to histological slices of paraffin-embedded GIT tissues from BALB/c and germ-free mice.
• Visualization of bacterial populations, specifically Lactobacillus spp. and Bifidobacterium spp., within biofilms on mucosal surfaces and in GIT contents.

Main Results:

• The Histo-FISH method was successfully standardized for in vivo detection of Lactobacillus and Bifidobacterium in biofilms.
• The technique allows for the visualization of bacterial populations on GIT mucosal surfaces and within GIT contents.
• The method demonstrated suitability for analyzing bacterial colonization and biofilm formation in the stomach and caecum of mice.

Conclusions:

• Histo-FISH provides an effective approach for studying in vivo biofilm dynamics in the gastrointestinal tract.
• This method offers potential for detecting other probiotic or pathogenic bacteria by adapting labeling probes.
• The Histo-FISH technique facilitates the analysis of microbial colonization processes and biofilm formation in complex gut environments.