Biophysical processes affecting the transit of a genetically-modified Pseudomonas fluorescens through the gut of the woodlouse Porcellio scaber

Our objective was to determine the effect of gut transit retention time of genetically-modified bacteria ingested by the woodlouse Porcellio scaber. The experimental animals were supplied ash leaf litter inoculated with the genetically-modified bacterium Pseudomonas fluorescens KTG and bacteria in food and faeces were counted using selective plating and immunofluorescent techniques. The bacteria were also detected using the polymerase chain reaction (PCR). It was found that plate counts of P. fluorescens KTG in fresh faeces were lower than those found in the litter when the GEMMO was supplied to animals at five different population densities, suggesting that a proportion of the GEMMO population was lost during gut transit. There was no significant difference in the survival of freshly cultured and starved cultures of P. fluorescens KTG on gut transit through P. scaber as determined by plate counts in fresh faeces. Retention time of P. fluorescens KTG in the woodlouse gut was found to be longer than that of the food bolus. The passage of bacteria through the gut was modelled and tracked using microbeads of a size similar to bacteria. Fluorescent microbeads added to food litter were detected within the anterior chamber, papillate region and rectum of the woodlouse for at least 17 days after ingestion. Scanning electron microscopy revealed that beads were retained within the cuticular structure of the digestive tract and also within mucopolysaccharide produced within the gut. Immunofluorescent observations of washed hindgut samples provided little evidence to suggest P. fluorescens KTG had become attached to the hindgut wall during transit. Very few colonies of the GEMMO and indigenous bacteria were detected from homogenised hepatopancreas samples. P. fluorescens KTG was however detected in the hepatopancreas of P. scaber using PCR. It is suggested that the retention of bacteria within the guts of woodlice is by physical rather than biological mechanisms such as growth or attachment.