De Leva, Giovanna (2024) Stress, Immunity and Honeybee Health: New Strategies to Limit Colony Losses. [Tesi di dottorato]

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Abstract

Honey bee health decline poses a significant global issue due to the profound impact of these pollinators on the environment and the human economy. Reduced bee survival is the ultimate outcome of a multifactorial syndrome triggered by various stressors that interact synergistically. A common factor among all collapsing colonies is the high load of parasites and associated pathogens, such as Deformed Wing Virus (DWV). DWV is an endemic immunosuppressive virus that causes asymptomatic covert infections, usually kept under control by the bees’ immune system unless exposed to stressors that weaken antiviral defenses. DWV-induced immunosuppression not only affects individual defense against pathogens but may also disrupt the regulation of the gut bacterial community, which plays a crucial role in gut physiology and immunity. The limited understanding of the molecular mechanisms underlying DWV infection hampers the development of new management strategies and therapeutic approaches aimed at mitigating the negative impact of pathogens on honey bees. In the first chapter of the present thesis, we explored the effects of DWV infection on the modulation of bee gut microbiota. We analyzed bacterial gut community via 16S rRNA amplicon sequencing under natural and artificial infection conditions. Both approaches indicated the occurrence of gut dysbiosis in infected bees, characterized by a depletion of Bacilli and an increase of Alphaproteobacteria and Clostridia. A preliminary experiment demonstrated that DWV infection can be limited by supplementing honey bees with Lactobacillus species, a taxon which is negatively impacted by DWV infection. The second chapter focuses on the identification of protein interactors between the virus and host involved in DWV pathogenesis. Our proteomic approach, consisting in co-immunoprecipitation and mass spectrometry analysis of bee protein extracts, identified 142 interactor proteins. Among these, arginine kinase, was selected for functional studies, to assess its role in DWV infection. Arginine kinase expression correlated positively with DWV load and its knockdown in honey bee-infected cells reduced DWV load after artificial infection. These results make it a potential candidate for developing new therapeutic strategies based on gene knockdown aimed at reducing DWV replication. Collectively, these results represent a novel contribution to the fast-growing research area of probiotics and therapeutics aimed at preserving pollinators' health against pathogens.

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