DOI: 10.5061/dryad.crjdfn3df
title: Primers, site data, and virus abundance in honey bees and Andrena spp.

description: Bees are focal pollinators, essential for maintaining biodiversity and
 crop production. Thus, reports of high annual honey bee colony losses and
 population declines among many wild bees in different parts of the world
 are of major concern. The spread of viruses is highlighted as a potential
 threat to bee communities. Viruses infect a wide range of bee species, and
 can be transmitted interspecifically through shared floral resources.
 Therefore, the role of flowers as hubs of bee virus transmission requires
 a community ecology perspective. Here, we investigate local and
 landscape-scale characteristics of floral communities potentially
 associated with the spread of viruses in the solitary Andrena spp. (mining
 bees). We surveyed 14 sites in a Mediterranean agroecosystem with varying
 local densities of honey bee (Apis mellifera) foragers and diversity of
 flowering species, and assessed the prevalence of four common
 Hymenoptera-associated viruses [deformed wing virus (DWV), black queen
 cell virus (BQCV), sacbrood virus (SBV) and Lake Sinai virus-2 (LSV2)] in
 co-foraging honey bees and mining bees. We found that the probability of
 virus presence in mining bees was generally associated with the diversity
 and composition of the local (site level) floral community, and with
 floral resource availability at the landscape scale (up to 1000 m range).
 In addition, SBV and DWV prevalence in mining bees was positively related
 to the density of SBV-infected, and total honey bee foragers,
 respectively. These findings demonstrate the focal role that the floral
 community at multiple spatial scales, and co-foraging pollinator species,
 may play in virus spread and, potentially, pollinator health.
description: We conducted a field survey during spring 2018, which included a
 total of fourteen sites. Each site comprised a 625
 m<sup>2</sup> plot located at the center of a larger wild
 bloom patch (mostly &gt;3000m<sup>2</sup>). Field work was
 conducted in mid-March, corresponding to peak spring bloom, and lasted two
 weeks. Each site was sampled once between 8:00 and 16:00 when weather
 conditions were favorable for bee activity (temperature &gt;17 °C,
 wind velocity &lt;3 m/s, and clear or partially clear skies). Foraging
 bee sampling was carried out for a total of 40 person-minutes (excluding
 handling time of captured bees) and conducted by slowly walking throughout
 the plot and hand-netting any bee that was observed landing on a flower,
 while keeping records of the visited flower species. Each captured bee was
 identified in-situ to the lowest taxonomic level possible, immediately
 placed in a vial, and kept on dry ice. We collected additional foraging
 bees as needed to attain a sample of at least 11 <em>Andrena
 </em>and 11 <em>A. mellifera</em> individuals per site
 for viral screening. At the end of each sampling day, all the collected
 specimens were brought to the lab and stored at –80 °C.  The sample of
 mining bees that was used for viral screening consisted of the three most
 dominant <em>Andrena</em> species (collectively comprising
 &gt; 70% of the observed <em>Andrena</em> individuals in
 the study): <em>A. ocraceohirta, A. aerinifrons
 levantina</em>, <em>A. urfanella</em>, and a closely
 related morphospecies from the subgenus <em>Truncandrena</em>.
 From each site, virus identification efforts were focused on individual
 <em>A. mellifera</em> and<em> Andrena</em>
 (median=11). High throughput sequencing identified regional virus genome
 sequence variations for BQCV and LSV-2, which required utilization of new
 primer sequences (Daughenbaugh et al., 2021). The presence and abundance
 of four common hymenopteran viruses, namely BQCV, DWV, SBV, and LSV-2, was
 assessed using the procedure of RNA extraction followed by cDNA
 preparation, polymerase chain reaction (PCR) and qPCR, as described in
 Daughenbaugh et al. (2021).
description: # Primers, site data, and virus abundance in honey bees and Andrena spp.
 [https://doi.org/10.5061/dryad.crjdfn3df](https://doi.org/10.5061/dryad.crjdfn3df) ## Description of the data and files structure #### The dataset includes the following four tables: **T1. Primers** Lists the primers used in the qPCR assay for viral screening of the sampled bees. For each primer, the following attributes are specified: purpose, primer name, sequence (5’-3’), product size (bp), and reference. **T2. Sites** Contains detailed information on each surveyed site, including the following attributes: floral species richness (flowering plants visited by bees) as recorded during the survey, floral diversity (Shannon index), Floral Resource Availability (FRA) index computed based on land cover data, coordinates (UTM) of each site, forager density of *Apis mellifera* and *Andrena* (bees per 40 minutes), analysis sample size for each *Apis mellifera* and *Andrena* forager for each of the four viruses (BQCV, DWV, LSV, SBV), and the prevalence of each virus in *Apis mellifera* and *Andrena*. The site coordinates are reported at full collected precision to enable spatial analyses. The focal *Andrena* species studied are common and widespread in the region, thrive in disturbed habitats, and are not listed as threatened, endangered, or vulnerable in global or regional assessments. Therefore, publishing precise coordinates is not expected to pose a conservation risk. **T3. qPCR-** ***Andrena*** **sample** Presents data on virus abundance in individual *Andrena* samples for viruses identified in the preceding Next Generation Sequencing (NGS) assay conducted on pooled bee sub-samples (Daughenbaugh et al. 2021). This includes the four focal viruses - BQCV, DWV, SBV, LSV - and an additional, less prevalent virus, AmFV. Virus abundance in individual bee samples was assessed using qPCR, with viral RNA copy numbers calculated from a standard curve and expressed as the number of viral RNA copies per 100 ng of RNA. For statistical analyses, quantitative data were reduced to presence (1) or absence (0). In the table, "specimen.number" identifies each bee in the sample, "qPCR.batch" refers to the qPCR run (1–3), and "Genus" and "Species" indicate taxonomic classification. The standard curves for each of the five viruses are presented in separate files. **T4. qPCR-** ***Apis mellifera*** **sample** Virus abundance in individual honey bee (*Apis mellifera*) samples. Column names and descriptions are as described under "T3. qPCR- *Andrena* sample". NAs (not available) indicate cases where there was insufficient RNA left to generate cDNA for viral RNA quantification by qPCR. The standard curves for each of the five viruses are presented in separate files. ### Contact  In case of questions regarding this data file, users may contact Idan Kahnonitch at [idan.kanonitch@mail.huji.ac.il](mailto:idan.kanonitch@mail.huji.ac.il). ### Files and variables #### Files: **T1._Primers.xlsx** **T2._Sites.xlsx**  **T3._qPCR-_Andrena_sample.csv** **T4._qPCR-_Apis_mellifera_sample.csv** **Standard_curve_BQCV.jpeg** **Standard_curve_DWV.jpeg** **Standard_curve_SBV.jpeg** **Standard_curve_LSV2.jpeg** **Standard_curve_AmFV.jpeg** **Description:**  This dataset consists of four tables, each presented in a separate Excel file, and five image plots. It contains the following information: 1. Details of primers used in qPCR assays (**T1. Primers**). 2. Surveyed sites with key attributes (**T2. Sites**). 3. Virus abundance data for *Andrena* (**T3. qPCR – Andrena sample**). 4. Virus abundance data for *Apis mellifera* (**T4. qPCR – Apis mellifera sample**) 5. qPCR standard curve plot for BQCV (Black queen cell virus), used to calculate viral RNA copy numbers in individual bee samples (**Standard_curve_BQCV**) 6. qPCR standard curve plot for DWV (Deformed wing virus), used to calculate viral RNA copy numbers in individual bee samples (**Standard_curve_DWV**) 7. qPCR standard curve plot for LSV2 (Lake Sinai virus 2), used to calculate viral RNA copy numbers in individual bee samples (**Standard_curve_LSV2**) 8. qPCR standard curve plot for SBV (Sacbrood virus), used to calculate viral RNA copy numbers in individual bee samples (**Standard_curve_SBV**) 9. qPCR standard curve plot for AmFV (Apis mellifera filamentous virus), used to calculate viral RNA copy numbers in individual bee samples (**Standard_curve_AmFV**) ## Code/software The dataset tables were created in Microsoft Excel (XLSX and CSV) and can be viewed using many free or widely available programs, including: 1. Google Sheets 2. Chrome extension “Office Editing” 3. LibreOffice Calc 4. Apache OpenOffice 5. Aspose Excel Viewer 6. Zoho Sheet The standard curve plots are provided as image files (JPEG) and can be viewed with common free programs, such as: 1. Windows Photos (Windows) 2. Preview (Mac) 3. ImageJ (cross-platform, open source) 4. GIMP (cross-platform, open source)