Variation in North American bumble bee nest success and colony sizes under captive rearing conditions.

Of the 265 known bumble bee (Bombus) species, knowledge of colony lifecycle is derived from relatively few species. As interest in Bombus commercialization and conservation grows, it is becoming increasingly important to understand colony growth dynamics across a variety of species since variation exists in nest success, colony growth, and reproductive output. In this study, we reported successful nest initiation and establishment rates of colonies and generated a timeline of colony development for 15 western North American Bombus species, which were captively reared from wild-caught gynes from 2009 to 2019. Additionally, we assessed variation in colony size among 5 western North American Bombus species from 2015 to 2018. Nest initiation and establishment rates varied greatly among species, ranging from 5-76.1% and 0-54.6%, respectively. Bombus griseocollis had the highest rates of nest success across the 11-yr period, followed by B. occidentalis, B. vosnesenskii, and B. huntii. Furthermore, days to nest initiation and days to nest establishment varied among species, ranging from 8.4 to 27.7 days and 32.7 to 47 days. Colony size also differed significantly among species with B. huntii and B. vosnesenskii producing more worker/drone cells than B. griseocollis, B. occidentalis, and B. vancouverensis. Additionally, gyne production differed significantly among species with B. huntii colonies producing more gynes than B. vosnesenskii. Results from this study increase knowledge of systematic nesting biology for numerous western North American Bombus species under captive rearing conditions, which can further improve rearing techniques available to conservationists and researchers.

Comparative analysis of 3 pollen sterilization methods for feeding bumble bees.

Pollen is an essential component of bee diets, and rearing bumble bees (Bombus spp.) for commercial use necessitates feeding pollen in mass quantities. This pollen is collected from honey bee (Apis mellifera L.) colonies because neither an artificial diet nor an economical, large-scale pollen collection process from flowers is available. The provenance of honey bee-collected pollen is often unknown, and in some cases has crossed international borders. Both deformed wing virus (DWV) and the fungal pathogen Ascosphaera apis (Claussen) Olive & Spiltoir (cause of chalkbrood disease); occur in honey bee-collected pollen, and infections have been observed in bumble bees. We used these pathogens as general surrogates for viruses and spore-forming fungal diseases to test the efficacy of 3 sterilization methods, and assessed whether treatment altered pollen quality for the bumble bee. Using honey bee-collected pollen spiked with known doses of DWV and A. apis, we compared gamma irradiation (GI), ozone fumigation (OZ), and ethylene oxide fumigation (EO) against an untreated positive control and a negative control. Following sterilization treatments, we tested A. apis spore viability, detected viral presence with PCR, and tested palatability to the bumble bee Bombus impatiens Cresson. We also measured bacterial growth from pollens treated with EO and GI. GI and EO outperformed OZ treatment in pathogen suppression. EO had the highest sterilizing properties under commercial conditions and retained palatability and supported bee development better than other treatments. These results suggest that EO sterilization reduces pathogen risks while retaining pollen quality as a food source for rearing bumble bees.

Bumble bees exhibit body size clines across an urban gradient despite low genetic differentiation.

Environmental heterogeneity resulting from human-modified landscapes can increase intraspecific trait variation. However, less known is whether such phenotypic variation is driven by plastic or adaptive responses to local environments. Here, we study five bumble bee (Apidae: Bombus) species across an urban gradient in the greater Saint Louis, Missouri region in the North American Midwest and ask: (1) Can urban environments induce intraspecific spatial structuring of body size, an ecologically consequential functional trait? And, if so, (2) is this body size structure the result of plasticity or adaptation? We additionally estimate genetic diversity, inbreeding, and colony density of these species-three factors that affect extinction risk. Using ≥ 10 polymorphic microsatellite loci per species and measurements of body size, we find that two of these species (Bombus impatiens, Bombus pensylvanicus) exhibit body size clines across the urban gradient, despite a lack of population genetic structure. We also reaffirm reports of low genetic diversity in B. pensylvanicus and find evidence that Bombus griseocollis, a species thought to be thriving in North America, is inbred in the greater Saint Louis region. Collectively, our results have implications for conservation in urban environments and suggest that plasticity can cause phenotypic clines across human-modified landscapes.

Dominant bee species and floral abundance drive parasite temporal dynamics in plant-pollinator communities.

Pollinator reductions can leave communities less diverse and potentially at increased risk of infectious diseases. Species-rich plant and bee communities have high species turnover, making the study of disease dynamics challenging. To address how temporal dynamics shape parasite prevalence in plant and bee communities, we screened >5,000 bees and flowers over an entire growing season for five common bee microparasites (Nosema ceranae, Nosema bombi, Crithidia bombi, Crithidia expoeki and neogregarines). Over 110 bee species and 89 flower species were screened, revealing that 42% of bee species (12.2% individual bees) and 70% of flower species (8.7% individual flowers) had at least one parasite in or on them, respectively. Some common flowers (for example, Lychnis flos-cuculi) harboured multiple parasite species whilst others (for example, Lythrum salicaria) had few. Significant temporal variation of parasite prevalence in bees was linked to bee diversity, bee and flower abundance and community composition. Specifically, we found that bee communities had the highest prevalence late in the season, when social bees (Bombus spp. and Apis mellifera) were dominant and bee diversity was lowest. Conversely, prevalence on flowers was lowest late in the season when floral abundance was highest. Thus turnover in the bee community impacted community-wide prevalence, and turnover in the plant community impacted when parasite transmission was likely to occur at flowers. These results imply that efforts to improve bee health will benefit from the promotion of high floral numbers to reduce transmission risk, maintaining bee diversity to dilute parasites and monitoring the abundance of dominant competent hosts.

Why Are Queens Broodless? Failed Nest Initiation Not Linked to Parasites, Mating Status, or Ovary Development in Two Bumble Bee Species of Pyrobombus (Hymenoptera: Apidae: Bombus).

Bumble bees (Bombus [Hymenoptera: Apidae]) are important pollinators for agricultural crops, which has led to their commercial domestication. Despite their importance, little is known about the reproductive biology of bumble bees native to North America. The Hunt bumble bee (Bombus huntii Greene [Hymenoptera: Apidae]) and the Vosnesensky bumble bee (Bombus vosnesenskii Radoszkowski [Hymenoptera: Apidae] are native candidates for commercial production in western North America due to their efficacy in providing commercial pollination services. Availability of pollinators native to the region in which services would be provided would minimize the likelihood of introducing exotic species and spreading novel disease. Some parasites are known to affect bumble bee reproduction, but little is known about their prevalence in North America or how they affect queen success. Only 38% of wild-caught B. huntii and 51% wild-caught B. vosnesenskii queens collected between 2015 and 2017 initiated nests in the laboratory. Our objective was to identify causal factors leading to a queen's inability to oviposit. To address this, we dissected each broodless queen and diagnosed diseases, assessed mating status, and characterized ovary development. Nematodes, arthropods, and microorganisms were detected in both species. Overall, 20% of queens were infected by parasites, with higher rates in B. vosnesenskii. Over 95% of both species were mated, and over 88% had developed ovaries. This suggests that parasitism and mating status were not primary causes of broodlessness. Although some failure to nest can be attributed to assessed factors, additional research is needed to fully understand the challenges presented by captive rearing.

Characterizing bumble bee (Bombus) communities in the United States and assessing a conservation monitoring method.

AIM: Bumble bees (Hymenoptera: Apidae: Bombus) are economically and ecologically important pollinators in agroecosystems and wildland habitats. In the Nearctic region, there are approximately 41 species, of which the IUCN lists twelve species as vulnerable, endangered, or critically endangered. We conducted a standardized faunal survey to inform ongoing conservation efforts including petitions under review for the Endangered Species Act. Furthermore, we test the appropriateness of a methodology for accurately sampling bumble bee communities. LOCATION: The United States of America, including 31 sites in 15 states. METHODS: We surveyed 15 states in the summer of 2015 to assess community composition and relative species abundance at agricultural and seminatural sites throughout the United States. We collected approximately 100 bees, using aerial nets, from each of 31 sites and identified specimens to species, totaling 3,252 bees. We assessed our survey methodology to understand whether it accurately sampled the potential community of bumble bees at each site for utility in future monitoring efforts. RESULTS: Average site species richness was 5.1 ± 2.05, and we detected 30 of the 41 species documented historically within the contiguous United States. Sampling a site beyond 100 bees rarely added additional species detections, whereas adding additional sampling sites within an ecoregion frequently increased the species richness for the ecoregion. Thirteen of the 30 species we detected each accounted for <1% of the total fauna, and two species accounted for 49.02% of all bees captured. Species richness and evenness increased with increasing latitude across communities. MAIN CONCLUSIONS: Species diversity and evenness in bumble bees increases in northern latitudes and increasing elevation in the United States; however, a few common species tend to dominate communities while many species occur only in low numbers. The results of this survey effort can inform current conservation evaluations and planning.

A Nonlethal Method to Examine Non-Apis Bees for Mark-Capture Research.

Studies of bee movement and activities across a landscape are important for developing an understanding of their behavior and their ability to withstand environmental stress. Recent research has shown that proteins, such as egg albumin, are effective for mass-marking bees. However, current protein mass-marking techniques require sacrificing individual bees during the data collection process. A nonlethal sampling method for protein mark-capture research is sorely needed, particularly for vulnerable, sensitive, or economically valuable species. This study describes a nonlethal sampling method, in which three non-Apis bee species (Bombus bifarius Cresson [Hymenoptera: Apidae], Osmia lignaria Say [Hymenoptera: Megachilidae], and Megachile rotundata Fabricius [Hymenoptera: Megachilidae]) were tested for a unique protein marker by immersing them momentarily in saline buffer and releasing them. Results showed that an egg albumin-specific enzyme-linked immunosorbent assay was 100% effective at detecting the protein on bees that were sampled nonlethally. Furthermore, this sampling method did not have an impact on bee survivorship, suggesting that immersing bees in buffer is a reliable and valid surrogate to traditional, destructive sampling methods for mark-capture bee studies.

Novel multiplex PCR reveals multiple trypanosomatid species infecting North American bumble bees (Hymenoptera: Apidae: Bombus).

Crithidia bombi and Crithidia expoeki (Trypanosomatidae) are common parasites of bumble bees (Bombus spp.). Crithidia bombi was described in the 1980s, and C. expoeki was recently discovered using molecular tools. Both species have cosmopolitan distributions among their bumble bee hosts, but there have been few bumble bee studies that have identified infections to species since the original description of C. expoeki in 2010. Morphological identification of species is difficult due to variability within each stage of their complex lifecycles, although they can be easily differentiated through DNA sequencing. However, DNA sequencing can be expensive, particularly with many samples to diagnose. In order to reliably and inexpensively distinguish Crithidia species for a large-scale survey, we developed a multiplex PCR protocol using species-specific primers with a universal trypanosomatid primer set to detect unexpected relatives. We applied this method to 356 trypanosomatid-positive bumble bees from North America as a first-look at the distribution and host range of each parasite in the region. Crithidia bombi was more common (90.2%) than C. expoeki (21.3%), with most C. expoeki-positive samples existing as co-infections with C. bombi (13.8%). This two-step detection method also revealed that 2.2% samples were positive for trypanosmatids that were neither C. bombi nor C. expoeki. Sequencing revealed that two individuals were positive for C. mellificae, one for Lotmaria passim, and three for two unclassified trypanosomatids. This two-step method is effective in diagnosing known bumble bee infecting Crithidia species, and allowing for the discovery of unknown potential symbionts.

Rarely reported, widely distributed, and unexpectedly diverse: molecular characterization of mermithid nematodes (Nematoda: Mermithidae) infecting bumble bees (Hymenoptera: Apidae: Bombus) in the USA.

Mermithid nematodes (Nematoda: Mermithida: Mermithidae) parasitize a wide range of both terrestrial and aquatic invertebrate hosts, yet are recorded in bumble bees (Insecta: Hymenoptera: Apidae: Bombus) only six times historically. Little is known about the specific identity of these parasites. In a single-season nationwide survey of internal parasites of 3646 bumble bees, we encountered six additional instances of mermithid parasitism in four bumble bee species and genetically characterized them using two regions of 18S to identify the specific host-parasite relationships. Three samples from the northeastern USA are morphologically and genetically identified as Mermis nigrescens, whereas three specimens collected from a single agricultural locality in the southeast USA fell into a clade with currently undescribed species. Nucleotide sequences of the V2-V6 region of 18S from the southeastern specimens were 2.6-3.0% divergent from one another, and 2.2-4.0% dissimilar to the nearest matches to available data. The dearth of available data prohibits positive identification of this parasite and its affinity for specific bumble bee hosts. By doubling the records of mermithid parasitism of bumble bee hosts and providing genetic data, this work will inform future investigations of this rare phenomenon.

The Giant Resin Bee, Megachile sculpturalis Smith: New Distributional Records for the Mid- and Gulf-south USA.

BACKGROUND: Megachile (Callomegachile) sculpturalis Smith, the giant resin bee, is an adventive species in the United States. First established in the United States during the early 1990s, records currently exist from most states east of the Mississippi River along with Iowa and Kansas. NEW INFORMATION: New distributional records are presented for Megachile (Callomegachile) sculpturalis Smith, an introduced bee. Additional records presented here expand the known distribution southwest through Arkansas, Louisiana, Mississippi, Missouri, and Texas. An updated host plant list containing new records is also presented, expanding the number of known floral associations.

Mitochondrial DNA diversity of honey bees (Apis mellifera) from unmanaged colonies and swarms in the United States.

To study the genetic diversity of honey bees (Apis mellifera L.) from unmanaged colonies in the United States, we sequenced a portion of the mitochondrial DNA COI-COII region. From the 530 to 1,230 bp amplicon, we observed 23 haplotypes from 247 samples collected from 12 states, representing three of the four A. mellifera lineages known to have been imported into the United States (C, M, and O). Six of the 13 C lineage haplotypes were not found in previous queen breeder studies in the United States. The O lineage accounted for 9% of unmanaged colonies which have not yet been reported in queen breeder studies. The M lineage accounted for a larger portion of unmanaged samples (7%) than queen breeder samples (3%). Based on our mitochondrial DNA data, the genetic diversity of unmanaged honey bees in the United States differs significantly from that of queen breeder populations (p < 0.00001). The detection of genetically distinct maternal lineages of unmanaged honey bees suggests that these haplotypes may have existed outside the managed honey bee population for a long period.

Multiplex polymerase chain reaction diagnostics of bed bug (Hemiptera: Cimicidae).

Cimex lectularius L. (Hemiptera: Cimicidae) is a widespread blood feeding pest of humans around the world, including North America, and has recently undergone a resurgence. A molecular diagnostic technique applying multiplex polymerase chain reaction (PCR) was developed to distinguish bed bug eggs, leg fragments, and degraded samples from other arthropods that frequently occur in human dwellings. A 410-428-bp region of the mitochondrial DNA 16S rRNA gene was used. To design C. lectularius-specific PCR primers, DNA sequences of various bed bug samples from the United States, Canada, and Australia, along with sequences of other Cimicidae and arthropods that often occur in dwellings, were considered. Based on DNA sequence variation, one reverse PCR primer specific for C. lectularius was identified. Multiplex PCR using three primers will yield a 417- and 140-bp amplicon for C. lectularius and a single 410-428-bp amplicon for other taxa. This assay was successful in identifying C. lectularius eggs, leg fragments, and degraded samples. This technique should provide a reliable, quick, and economical technique for identifying C. lectularius, when morphological identification is not possible.