Performing Department
(N/A)
Non Technical Summary
Bees are routinely transported for crop pollination services, and their associated parasites and pathogens are consequently co- introduced into novel environments. Importantly, pathogens have been associated with losses of several agriculturally important wild and managed pollinators. Agriculturally important pollinators including honey bees, leafcutter bees, and mason bees can suffer from chalkbrood disease, caused by species of fungi within the genus Ascosphaera. Co-occurrence of Ascosphaera species is presumed common in both managed and natural pollination systems, and recently introduced non-native mason bees (Osmia) from Asia have also brought along novel species of Ascosphaera in the US, which have been detected in native Osmia. This proposed Research project seeks to identify co-occurring Ascosphaera species in managed and wild Osmia, determine the pathogenicity of Ascosphaera species and their interactions inside bee nests, and evaluate consequences of Ascosphaera co-infection for native and introduced Osmia. Osmia nests will be collected in both native and non-native managed systems, and employ high-throughput sequencing, and analyze larval bioassay data to evaluate the outcomes of Ascosphaera co-infection on native and introduced mason bees. Single-species pathogen studies have been crucial for generating best management practices (BMPs) for the movement and trade of pollinators that safeguard pollinator health. However, a better understanding of novel co-infection dynamics and disease in bees could improve BMPs.?
Animal Health Component
100%
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Goals / Objectives
The overall goal of this project has been to support PD K. LeCroy in understanding how Ascosphaera species interact and shape disease outcomes in wild and managed mason bee species. This goal has been supported by developing collaborations within and across disciplines, mentoring undergraduate student researchers, and advancing the PD's skills in high-throughput sequencing, fungal culturing, and manipulative bioassays with mason bees. The following objectives pertain to this plan:Objective 1: Characterize Ascosphaera taxonomic diversity and co-occurrence in wild & managed mason bee species.Objective 2: Assess the ultimate consequences of Ascosphaera co-infections on pollinator health in vivo.
Project Methods
Objective 1: Characterize Ascosphaera taxonomic diversity and co-occurrence in wild & managed mason bee species. DNA has been extracted for some samples in the original planned study as a pilot test of the DNA extraction protocol (approx. 15% of specimens, n=250), but approximately 1,500 specimens remain to undergo high-quality DNA extraction with the improved DNA extraction protocol for specimens. All 250 specimens with previously extracted DNA also have ample leftover specimen tissue that will be used in the new high-yield DNA extraction protocol. After DNA extraction, molecular detection via PCR will be performed using Ascosphaera-specific primers followed by validation using gel electrophoresis. The resulting positive amplicon samples will be used for Illumina sequencing and analyzed with standard community profiling methods.Data Analyses & Interpretation: To test for patterns of co-occurrence of Ascosphaera among Osmia species, I will perform statistical tests using R. I will use a generalized linear model with binomial error distribution to evaluate differences in the detection of Ascosphaera species among each of the Osmia species found in nests. The predictor variables will be the Ascosphaera species, the Osmia species, and their two-way interaction. Significance of model terms will be evaluated by comparison to a model excluding interaction terms via the 'anova()' function and pairwise differences were evaluated using post-hoc Tukey tests via the 'emmeans()' function. I will inspect for residual model spatial autocorrelation using correlograms, bubble plots, variograms, and estimating Moran's I.I will additionally run a generalized linear model with a binomial distribution to evaluate the effect of multiple infections (which may be 4 levels: Ascosphaera sp. 1, Ascosphaera sp. 2, Ascosphaera sp. 3, or mixed infections) on Osmia individual mortality (defined as chalky cadavers and other strict diagnostic features).Expected Outcomes & Use of Results: This will be the first study to document and publish rates of co-infection by Ascosphaera species in mason bees. Based on preliminary findings, I predict that native Osmia will show higher prevalence of Ascosphaera compared to non-native Osmia, and the majority of infections will be from non-native Ascosphaera. Illumina sequencing will allow for detection of Ascosphaera species co-occurrence. If non-native Ascosphaera naganensis and A. fusiformis are strongly competitive as hypothesized, we may observe few instances of co-occurrence with these species and other Ascosphaera spp. I will publish the results of this study in a peer-reviewed journal, such as in Biological Conservation.Pitfalls, Limitations, & Their Resolution: Rates of Ascosphaera detection range between 14-47% in this study system, and therefore statistical power for planned tests may be limited if enough bees are not collected. Data from preliminary studies show an overall Ascosphaera detection rate of approximately 25%, and the sample size we have will provide adequate statistical power. With the planned analyses, a power analysis shows we must screen approximately 960 bees, and we have at minimum 1,400 bees to screen.Objective 2: Assess the ultimate consequences of Ascosphaera co-infections on pollinator health in vivo. Although the PD has completed alarval bioassay, the PD still needs to complete data analysis, confirm via molecular detection any background Ascosphaera presence along with treatment efficacy for control and experimental treatments. After completing the analyses, the PD will write and publish findings in a peer-reviewed journal such as Nature Microbiology.