Drosophila melanogaster as a model host for studying Pseudomonas aeruginosa feeding infection

Drosophila is an important model organism for studying epithelial barrier pathogenesis by Pseudomonas aeruginosa because it allows clinically important infectivity to be mimicked. Here, we present a Protocol Extension to introduce P. aeruginosa and other bacteria into the adult Drosophila intestine by feeding, further to our previously published protocol that used the needle-pricking method to impose a wound infection and the injector-pumping method of bacterial delivery directly into the fly hemocoel. We start with Drosophila preparation and priming for feeding, in parallel with bacterial growth and infection mix preparation. We continue with the feeding setup followed by the output measurements, which include, but are not limited to, fly survival, bacterial load and systemic spread, intestinal regeneration and gene expression. The protocol can be used to infect Drosophila with tens of different clinically important and insect-related bacterial species, one at a time or in combination; modify the inoculum consistency to control and even eliminate P. aeruginosa virulence; measure bacterial load in the hemolymph; assess intestinal histopathology of tumor-prone flies; and assess the host transcriptome. Preparation of adult flies for feeding lasts ≤1 week, and infection mixes can be prepared in 2 d, which require a minimum level of expertise in fly handling and microbiological techniques, respectively. All output measurements usually take ≤10 d or until all flies die from a virulent infection. Fly survival and bacterial load assessment require a short training, while intestinal histopathology and host gene expression assessment need up to a month of systematic training. Assessing bacterial load and gut measurements require 2 d of work per infection time point considered.