Electronic Thesis and Dissertation Repository

Thesis Format

Integrated Article

Degree

Doctor of Philosophy

Program

Microbiology and Immunology

Supervisor

Reid, Gregor

Abstract

Arguably the most important insect pollinator, honey bees (Apis mellifera) are threatened by infectious disease, pesticide exposure, and nutritional deficits resulting from habitat loss.

The major goals of this thesis were to advance our understanding of the immune and microbiota factors underpinning synergistic interactions between the threats, and to develop a honey bee-specific probiotic with strain-level functions to alleviate cumulative stress burden.

Drosophila melanogaster was used as a high-throughput discovery platform in molecularly characterizing the effects of candidate probiotic lactobacilli and neonicotinoid-neuroimmune-microbiota mechanisms prior to experimentation in honey bees. Imd pathway activation by lactobacilli was identified to be crucial for mitigating neonicotinoid-induced suppression of Relish/NF-κB signalling, although constitutive overexpression of Rel-68 (active cleavage product) showed no benefit to survival using GAL4/UAS genetic techniques. NOX/DUOX oxidative burst pathways were implicated for their co-modulatory roles but were not directly involved in lactobacilli-mediated resolution of infection susceptibility and/or microbiota dysbiosis.

Next, a pollen patty-based delivery system (BioPatty) was developed to enable honey bee supplementation with three identified probiotic candidates: Lactiplantibacillus plantarum Lp39, Lacticaseibacillus rhamnosus GR-1, and Apilactobacillus kunkeei BR-1 (LX3). The strains were confirmed to be highly viable for a minimum of six days in the BioPatty matrix and could reach their intended hive targets (adult and larval intestinal tracts), with nutrient analyses (AOAC.994.12) demonstrating a near-optimal essential amino acid profile for supporting honey bee health.

Importantly, based on multi-study field trial data and in-vitro rearing experiments, two probiotic functions were validated for the BioPatty including improved pathogen resistance against Paenibacillus larvae (deadly agent of American foulbrood) and microbiota recovery following antibiotic exposure.

Taking a systems-level approach to understanding honey bee microbial ecological networks, a metataxonomic database tool (BEExact) was also developed and validated on ∼234 million short-read 16S rRNA sequences derived from 32 microbiota studies encompassing 50 bee species. In short, BEExact offers a field-wide resource enabling study of unculturable, or yet-to-be cultured, microbial ‘dark matter’ relevant to disease spread amongst the pollinator assemblage.

The collective scope of this work is expansive, and the basic approaches used, in theory, have multitudinous applications broadly relevant to microbial management of terrestrial and aquatic ecosystems.

Summary for Lay Audience

Managed populations of honey bees are utilized in agriculture to ensure adequate pollination and maximal crop yields. However, over the past decade or more, honey bees have been facing unsustainability high levels of colony loss – a well-known societal quandary with infectious disease, pesticide exposure, and habitat loss representing the causal factors involved.

Despite the common preference of grouping these as separate issues, in reality the stress factors are not mutually exclusive and can synergistically influence the health of honey bees. For example, exposure to certain pesticides like neonicotinoids can suppress the immune system of honey bees and thereby increase susceptibility to lethal infections. Similar parallels can be drawn for nutritional deficits induced by a lack of floral diversity, weakened immunity, and infection susceptibility. Current disease prevention strategies address the latter (and most severe) factor by administering prophylactic antimicrobial agents to the hive, although mounting evidence suggests this may jeopardize long-term health by damaging host-adapted symbiont communities in the honey bee gut microbiota.

Here, I investigate the underlying host-microbial interactions that can influence honey bee health outcomes during single or combined exposure to pathogens, pesticides, and/or antibiotics. In doing so, two microbial innovations with translational potential were developed: i) a honey bee-specific probiotic (BioPatty) capable of mitigating multifactorial stressors through strengthening the host innate immune response and restoring microbiota homeostasis post-exposure to noxious stimuli, and ii) a microbiota analysis tool (BEExact) capable of enhancing the overall utility and ecological relevance of routine 16S rRNA gene-based sequencing endeavors in honey bees.

Creative Commons License

Creative Commons Attribution 4.0 License
This work is licensed under a Creative Commons Attribution 4.0 License.

Available for download on Wednesday, August 31, 2022

Share

COinS