The paratransgenic potential of Lactobacillus kunkeei in the honey bee Apis mellifera

The honey bee (Apis mellifera) is a domestic insect of high value to human societies, as a crop pollinator in agriculture and a model animal in scientific research. The honey bee, however, has experienced massive mortality worldwide due to the phenomenon Colony Collapse Disorder (CCD), resulting in alarming prospects for crop failure in Europe and the USA. The reasons for CCD are complex and much debated, but several honey bee pathogens are believed to be involved. Paratransgenesis is a Trojan horse strategy, where endogenous microorganisms are used to express effector molecules that antagonise pathogen development. For use in honey bees, paratransgenesis must rely on a set of criteria that the candidate paratransgenic microorganism must fulfil in order to obtain a successful outcome: (1) the candidate must be genetically modifiable to express effector molecules; (2) the modified organism should have no adverse effects on honey bee health upon reintroduction; and (3) it must survive together with other non-pathogenic bee-associated microorganisms. Lactic acid bacteria (LAB) are common gut bacteria in vertebrates and invertebrates, and some have naturally beneficial properties in their host. In the present work we aimed to find a potential paratransgenic candidate within this bacterial group for use in honey bees. Among isolated LAB associated with bee gut microbiota, we found the fructophilic Lactobacillus kunkeei to be the most predominant species during foraging seasons. Four genetically different strains of L. kunkeei were selected for further assessment. We demonstrated (1) that L. kunkeei is transformable; (2) that the transformed cells had no obvious adverse effect on honey bee survival; and (3) that transformed cells survived well in the gut environment of bees upon reintroduction. Our study demonstrates that L. kunkeei fulfils the three criteria for paratransgenesis and can be a suitable candidate for further research on this strategy in honey bees.