Abstract
American Foulbrood is a worldwide distributed, fatal disease of the brood of the Western honey bee (Apis mellifera). The causative agent of this fatal brood disease is the Gram-positive, spore-forming bacterium Paenibacillus larvae, which can be classified into four different genotypes (ERIC I-IV), with ERIC I and II being the ones isolated from contemporary AFB outbreaks. P. larvae is a peritrichously flagellated bacterium and, hence, we hypothesized that P. larvae is capable of coordinated and cooperative multicellular behaviors like swarming motility and biofilm formation. In order to analyze these behaviors of P. larvae, we firstly established appropriate functional assays. Using these assays we demonstrated that P. larvae ERIC II, but not P. larvae ERIC I, was capable of swarming. Swarming motility was hampered in a P. larvae ERIC II-mutant lacking production of paenilarvin, an iturin-like lipopeptide exclusively expressed by this genotype. Both genotypes were able to form free floating biofilm aggregates loosely attached to the walls of the culture wells. Visualizing the biofilms by Congo red and thioflavin S staining suggested structural differences between the biofilms formed. Biofilm formation was shown to be independent from paenilarvin production because the paenilarvin deficient mutant was comparably able to form a biofilm.
Highlights
The Western honey bee Apis mellifera is a generalist pollinator and managed colonies of A. mellifera are widely used in global agriculture for the pollination of many crops and fruit grown in the open field[1,2]
The surface layer protein SplA is expressed by P. larvae ERIC II and mediates bacterial adhesion to the midgut epithelium, a step that is obviously relevant for the pathogenic strategy of P. larvae ERIC II15,19
To test the hypothesis that P. larvae is able to perform a multicellular flagellum-driven surface movement known as swarming motility, we first established an appropriate experimental procedure based on soft agar plates (0.5%) made from brain heart infusion broth (BHI) and on inoculating 10 μl of a bacterial suspension with an OD600 of 0.1 centrally onto the agar plate
Summary
The Western honey bee Apis mellifera is a generalist pollinator and managed colonies of A. mellifera are widely used in global agriculture for the pollination of many crops and fruit grown in the open field[1,2]. All P. larvae ERIC I strains formed macrocolonies but were not able to swarm while all tested P. larvae ERIC II strains nearly covered the entire surfaces of the plates within two days (Fig. 2). To test the hypothesis that P. larvae is capable of forming multicellular aggregates known as biofilms we established culture conditions that resulted in the formation of free floating bacterial aggregates that were visible to the naked eye and resembled biofilms (Fig. 3A,B).
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