Abstract
Honeybee (Apis mellifera L.) workers act as passive vectors of Paenibacillus larvae spores, which cause the quarantine disease American foulbrood (AFB). We assessed the relative proportions of P. larvae within the honeybee microbiome using metabarcoding analysis of the 16 S rRNA gene. The microbiome was analyzed in workers outside of the AFB zone (control - AFB0), in workers from asymptomatic colonies in an AFB apiary (AFB1), and in workers from colonies exhibiting clinical AFB symptoms (AFB2). The microbiome was processed for the entire community and for a cut-off microbiome comprising pathogenic/environmental bacteria following the removal of core bacterial sequences; varroosis levels were considered in the statistical analysis. No correlation was observed between AFB status and varroosis level, but AFB influenced the worker bee bacterial community, primarily the pathogenic/environmental bacteria. There was no significant difference in the relative abundance of P. larvae between the AFB1 and AFB0 colonies, but we did observe a 9-fold increase in P. larvae abundance in AFB2 relative to the abundance in AFB1. The relative sequence numbers of Citrobacter freundii and Hafnia alvei were higher in AFB2 and AFB1 than in AFB0, whereas Enterococcus faecalis, Klebsiella oxytoca, Spiroplasma melliferum and Morganella morganii were more abundant in AFB0 and AFB1 than in AFB2.
Highlights
Honeybee (Apis mellifera L.) workers act as passive vectors of Paenibacillus larvae spores, which cause the quarantine disease American foulbrood (AFB)
We analyzed the occurrence of P. larvae in AFB-affected apiaries, including colonies exhibiting clinical signs, and in asymptomatic colonies, by employing a high-throughput sequencing (HTS) approach and comparing the obtained data with those from control samples
Our samples consisted of worker honeybees, which are suitable for AFB diagnostics[6, 20, 21] and facilitate both horizontal and vertical bacterial transmission[22]
Summary
Honeybee (Apis mellifera L.) workers act as passive vectors of Paenibacillus larvae spores, which cause the quarantine disease American foulbrood (AFB). In a study investigating three honeybee pathogens in Spain, the detected P. larvae (as well as Melissococcus plutonius) prevalence was two-fold greater in adult bees than in brood samples[8]. In the case of P. larvae, even a tolerant honeybee colony typically does not exceed 2.5 × 105 spores per g of honey[5] If such a concentration of spores is present in hive food stores, sequencing analysis of the adults feeding on the contaminated honey or using it to feed the brood will be successful, and the total microbiome will be influenced by the number of pathogen sequences. HTS was employed to compare the relative proportions of P. larvae in the microbiomes of honeybee workers obtained from colonies with and without clinical symptoms as well as control samples from outside of the AFB zone. The presence of pathogenic P. larvae detected in commonly investigated sample types is demonstrated from a novel perspective in this study and the impact of P. larvae on the whole microbial community structure is assessed
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