Search

Climate change is predicted to affect host-parasite interactions, and for some hosts, parasite infection is expected to increase with rising temperatures. Global population declines of important pollinators already have been attributed to climate change and parasitism. However, the role of climate in driving parasite infection and the genetic basis for pollinator hosts to respond often remain obscure. Based on decade-long field data, we investigated the association between climate and Nosema bombi (Microsporidia) infection of buffed-tailed bumblebees (Bombus terrestris), and whether host genotypes play a role. For this, we genotyped 876 wild bumblebee queens and screened for N. bombi infection of those queens between 2000 and 2010. We recorded seven climate parameters during those 11 years and tested for correlations between climate and infection prevalence. Here we show that climatic factors drive N. bombi infection and that the impact of climate depends on mitochondrial DNA cytochrome oxidase I (COI) haplotypes of the host. Infection prevalence was correlated with climatic variables during the time when queens emerge from hibernation. Remarkably, COI haplotypes best predict this association between climatic factors and infection. In particular, two host haplotypes ("A" and "B") displayed phenotypic plasticity in response to climatic variation: Temperature was positively correlated with infection of host haplotype B, but not haplotype A. The likelihood of infection of haplotype A was associated with moisture, conferring greater resistance to parasite infection during wetter years. In contrast, infection of haplotype B was unrelated to moisture. To the best of our knowledge, this is the first study that identifies specific host genotypes that confer differential parasite resistance under variable climatic conditions. Our results underscore the importance of mitochondrial haplotypes to ward off parasites in a changing climate. More broadly, this also suggests that COI may play a pertinent role in climate change adaptations of insect pollinators.

The antagonistic relationship between parasites and their hosts is strongly influenced by genotype-by-genotype interactions. Defense against parasitism is commonly studied in the context of immune system-based mechanisms and, thus, the focus in the search for candidate genes in host-parasite interactions is often on immune genes. In this study, we investigated the association between prevalence of parasite infection and host mitochondrial DNA (mtDNA) haplotypes in two natural populations of bumblebees (Bombus terrestris). The two most common haplotypes of the host populations, termed A and B, differ by a single nonsynonymous nucleotide substitution within the coding region of cytochrome oxidase I, an important player in metabolic pathways. We screened infection by Nosema bombi, a common endoparasite of bumblebees, and the corresponding host mtDNA-haplotype frequencies in over 1400 bumblebees between 2000 and 2010. The island population of Gotland showed lower mtDNA diversity compared to the mainland population in Switzerland. Over time, we observed large fluctuations in infection prevalence, as well as variation in host haplotype frequencies in both populations. Our long-term observation revealed that N. bombi infection of specific host genotypes is transient: We found that with increasing infection prevalence, proportionally more individuals with haplotype B, but fewer individuals with haplotype A were infected. This suggests that the presence of N. bombi in specific host genotypes relates to infection prevalence. This may be a result of parasite competition, or differential resilience of host types to ward off infections. The findings highlight the important role of host mtDNA haplotypes in the interaction with parasites.

A total of 1940 isolates from gut samples of 60 bumblebees representing Bombus pascuorum, Bombus terrestris, Bombus lucorum and Bombus lapidarius was collected and identified through state-of the-art taxonomic methods. The bacterial species diversity in these Bombus species exceeded that suggested by phylotype analysis through 16S rRNA amplicon sequencing, and revealed that B. pascuorum and B. terrestris had a unique microbiota composition, each. Representatives of most phylotypes reported earlier and detected in the present study were effectively isolated, and included several novel bacterial taxa and species reported for the first time in the bumblebee gut. Isolates were screened in pectin degradation assays and growth inhibition assays against the honeybee pathogens Paenibacillus larvae, Melissococcus plutonius and Ascosphaera apis and the bumblebee parasite Crithidia bombi. While inhibitory activity against each of these pathogens was observed, only one single culture was able to degrade pectin and polygalacturonic acid in vitro. The availability of accurately identified microbial isolates will facilitate future evaluation of the functional potential of the bumblebee gut microbiota.

As pollinators, bees are cornerstones for terrestrial ecosystem stability and key components in agricultural productivity. All animals, including bees, are associated with a diverse community of microbes, commonly referred to as the micro biome. The bee micro biome is likely to be a crucial factor affecting host health. However, with the exception of a few pathogens, the impacts of most members of the bee microbiome on host health are poorly understood. Further, the evolutionary and ecological forces that shape and change the microbiome are unclear. Here, we discuss recent progress in our understanding of the bee microbiome, and we present challenges associated with its investigation. We conclude that global coordination of research efforts is needed to fully understand the complex and highly dynamic nature of the interplay between the bee micro biome, its host, and the environment. High-throughput sequencing technologies are ideal for exploring complex biological systems, including host-microbe interactions. To maximize their value and to improve assessment of the factors affecting bee health, sequence data should be archived, curated, and analyzed in ways that promote the synthesis of different studies. To this end, the BeeBiome consortium aims to develop an online database which would provide reference sequences, archive metadata, and host analytical resources. The goal would be to support applied and fundamental research on bees and their associated microbes and to provide a collaborative framework for sharing primary data from different research programs, thus furthering our understanding of the bee microbiome and its impact on pollinator health. Funding Agencies|National Evolutionary Synthesis Center (NESCent), NSF [EF-0905606]

This study provides, for the first time, sequence data for the protozoan flagellates Crithidia bombi and Crithidia mellificae (Kinetoplastea: Trypanosomatidae). We amplified the partial sequences of the small subunit ribosomal RNA (SSU rRNA), glycosomal glyceraldehyde phosphate dehydrogenase (gGAPDH), cytochrome b (Cyt b), and the complete internal transcribed spacer region 1 (ITS1) of the ribosomal RNA gene region for 66 clones of C. bombi from Switzerland and Alaska. Furthermore, we sequenced the same stretch of SSU rRNA and gGAPDH for one isolate of C. mellificae from Switzerland. All four molecular markers classified the C. bombi samples into two distinct lineages A and B. Both lineages were found in the two sampling locations. Variation within lineages was small or non-existing. Sequence differences between lineages were 1.64% for SSU rRNA, 4.36% for gGAPDH, and 12.02% for Cyt b. The ITS1-sequences of lineages A and B have diverged so much that no alignment was possible. With regard to ITS1, we additionally found fragment length polymorphism (variation in microsatellite repeat numbers) as well as nucleotide diversity within each lineage. Furthermore, the sequences of SSU rRNA and gGAPDH of C. mellificae were different from both lineages of C. bombi. The separation of lineages A and B, based on sequence differences and phylogenetic reconstruction, is so pronounced as to characterize two species of "C. bombi." We propose to retain C. bombi for the more common lineage A and suggest the name Crithidia expoeki n. sp. for lineage B.

We present details and characteristics of 123 novel polymorphic microsatellite DNA loci for Bombus terrestris. Thirty-four of these loci have been tested in nine other Bombus species and 25 of them showed polymorphisms in at least one species. These microsatellite DNA loci together with the already established 60 loci will be useful for characterizing wild and managed populations of B. terrestris and other Bombus species as well as for detailed genetic studies in including mapping studies and genome annotations.

The primitively social bumblebee Bombus terrestris is an ecological model species as well as an important agricultural pollinator. As part of the ongoing development of genomic resources for this model organism, we have constructed a publicly available bacterial artificial chromosome (BAC) library from males of a field-derived colony. We have shown that this library has a high coverage, which allows any particular sequence to be retrieved from at least one clone with a probability of 99.7%. We have further demonstrated the library’s usefulness by successfully screening it with probes derived both from previously described B. terrestris genes and candidate genes from another bumblebee species and the honeybee. This library will facilitate genomic studies in B. terrestris and will allow for novel comparative studies in the social Hymenoptera.

In early 1992, the European bumblebee, Bombus terrestris, was first seen in Tasmania and currently has spread to most of the island. Here, we report on the genetic structure, using micro-satellites, of the invading population from samples collected in the years 1998-2000, a few years after the first sighting of the species in its new area. The data show that the Tasmanian population has a very low genetic diversity, with less than half of the allelic richness (Richness=2.89 alleles; H(exp)=0.591) and lower levels of heterozygosity as compared to populations in New Zealand (4.24 alleles; H(exp)=0.729) and Europe (5.08 alleles; H(exp)=0.826). In addition, the genetic data suggest that the invasion must have happened once, probably around late 1991, and was the result of very few, perhaps only two, individuals arriving in Tasmania. Furthermore, these founders came from the New Zealand population. Today, the population in the south of Tasmania seems to act as a source population from which individuals migrate into other parts of the state. A similar source-sink structure seems also the case for New Zealand. The data show that B. terrestris is a highly invasive species capable of establishing itself even after a dramatic genetic bottleneck. B. terrestris may be an invasive species due to the haplo-diploid sex determination system, which exposes recessive, deleterious mutations to selection. Offspring of such purged lines may then be able to tolerate high levels of inbreeding.

Bumblebees are highly valued for their pollination services in natural ecosystems as well as for agricultural crops. These precious pollinators are known to be declining worldwide, and one major factor contributing to this decline are infections by parasites. Knowledge about parasites in wild bumblebee populations is thus of paramount importance for conservation purposes. We here report the geographical distribution of Crithidia and Nosema, two common parasites of bumblebees, in a yet poorly investigated country: Mexico. Based on sequence divergence of the Cytochrome b and Glycosomal glyceraldehyde phosphate deshydrogenase (gGPDAH) genes, we discovered the presence of a new Crithidia species, which is mainly distributed in the southern half of the country. It is placed by Bayesian inference as a sister species to C. bombi. We suggest the name Crithidia mexicana for this newly discovered organism. A population of C. expoeki was encountered concentrated on the flanks of the dormant volcanic mountain, Iztaccihuatl, and microsatellite data showed evidence of a bottleneck in this population. This study is the first to provide a large-scale insight into the health status of endemic bumblebees in Mexico, based on a large sample size (n=3,285 bees examined) over a variety of host species and habitats.

Made available in DSpace on 2015-10-21T20:31:01Z (GMT). No. of bitstreams: 0 Previous issue date: 2015-04-24. Added 1 bitstream(s) on 2015-10-22T09:48:01Z : No. of bitstreams: 1 WOS000353676700001.pdf: 2518339 bytes, checksum: d1f8d18bd059b76504f3fcd9aa639a74 (MD5) National Institutes of Health (NIH) Agriculture and Food Research Initiative Competitive grant from the USDA National Institute of Food and Agriculture Research Council of Norway (NFR) PEW Charitable Trust University of East Anglia, UK Israel Science Foundation (ISF) Biotechnology and Biological Sciences Research Council, UK University of East Anglia University of Alabama College of Arts and Sciences Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Marie Curie International Outgoing Fellowship Swiss National Science Foundation New Faculty Initiative Grant (NFIG) from Illinois State University College of Arts and Sciences DFG Instituto Nacional de Bioinformatica (INB) from ISCIII in Spain Background: The shift from solitary to social behavior is one of the major evolutionary transitions. Primitively eusocial bumblebees are uniquely placed to illuminate the evolution of highly eusocial insect societies. Bumblebees are also invaluable natural and agricultural pollinators, and there is widespread concern over recent population declines in some species. High-quality genomic data will inform key aspects of bumblebee biology, including susceptibility to implicated population viability threats.Results: We report the high quality draft genome sequences of Bombus terrestris and Bombus impatiens, two ecologically dominant bumblebees and widely utilized study species. Comparing these new genomes to those of the highly eusocial honeybee Apis mellifera and other Hymenoptera, we identify deeply conserved similarities, as well as novelties key to the biology of these organisms. Some honeybee genome features thought to underpin advanced eusociality are also present in bumblebees, indicating an earlier evolution in the bee lineage. Xenobiotic detoxification and immune genes are similarly depauperate in bumblebees and honeybees, and multiple categories of genes linked to social organization, including development and behavior, show high conservation. Key differences identified include a bias in bumblebee chemoreception towards gustation from olfaction, and striking differences in microRNAs, potentially responsible for gene regulation underlying social and other traits.Conclusions: These two bumblebee genomes provide a foundation for post-genomic research on these key pollinators and insect societies. Overall, gene repertoires suggest that the route to advanced eusociality in bees was mediated by many small changes in many genes and processes, and not by notable expansion or depauperation. Illinois State Univ, Sch Biol Sci, Normal, IL 61790 USA ETH, Inst Integrat Biol, Expt Ecol, CH-8092 Zurich, Switzerland E Carolina Univ, Dept Biol, Greenville, NC 27858 USA Hebrew Univ Jerusalem, Alexander Silberman Inst Life Sci, Dept Ecol Evolut &Behav, Jerusalem, Israel Univ Ghent, Fac Sci, Lab Zoophysiol, B-9000 Ghent, Belgium Univ Otago, Dept Biochem, Labo Evolut &Dev Genet, Dunedin 9054, New Zealand Univ Otago, Dept Biochem, Natl Res Ctr Growth &Dev, Dunedin 9054, New Zealand Univ Missouri, Div Plant Sci, Div Anim Sci, Columbia, MO 65211 USA Univ Missouri, MU Informat Inst, Columbia, MO 65211 USA Georgetown Univ, Dept Biol, Washington, DC 20057 USA Arizona State Univ, Sch Life Sci, Tempe, AZ 85287 USA Univ Copenhagen, Dept Biol, Ctr Funct &Comparat Insect Gen, Copenhagen, Denmark Univ Hohenheim, Inst Anim Sci, D-70599 Stuttgart, Germany Univ Alabama, Dept Biol Sci, Tuscaloosa, AL 35487 USA Univ Illinois, Dept Entomol, Urbana, IL 61801 USA Univ Ghent, Fac Biosci Engn, Dept Crop Protect, Lab Agrozool, B-9000 Ghent, Belgium Univ Halle Wittenberg, Inst Biol, Wittenberg, Germany Univ Geneva, Sch Med, Dept Genet Med &Dev, CH-1211 Geneva, Switzerland Swiss Inst Bioinformat, CH-1211 Geneva, Switzerland MIT, Comp Sci &Artificial Intelligence Lab, Cambridge, MA 02139 USA MIT &Harvard, Broad Inst, Cambridge Ctr 7, Cambridge, MA 02142 USA Univ Munster, Inst Evolut &Biodivers, D-48149 Munster, Germany Ctr Genom Regulat CRG, Barcelona 08003, Spain UPF, Barcelona, Spain Ernst Moritz Arndt Univ Greifswald, Inst Math &Comp Sci, D-17487 Greifswald, Germany Univ Calif Berkeley, Lawrence Berkeley Natl Lab, Genom Div, Berkeley, CA 94720 USA Natl Lib Med, Natl Ctr Biotechnol Informat, Bethesda, MD 20894 USA Norwegian Univ Food Sci, Dept Chem Biotechnol &Food Sci, N-1432 As, Norway Univ E Anglia, Sch Comp Sci, Norwich NR4 7TJ, Norfolk, England Univ Dusseldorf, Inst Evolut Genet, D-40225 Dusseldorf, Germany Univ Cologne, Inst Genet, Cologne, Germany Univ Sao Paulo, Dept Biol, Fac Filosofia Ciencias &Letras Ribeirao Preto, BR-14040901 Ribeirao Preto, Brazil Univ Edinburgh, Ashworth Labs, Inst Evolutionary Biol &Edinburgh Gen, Edinburgh EH9 3FL, Midlothian, Scotland Univ E Anglia, Sch Biol Sci, Norwich NR4 7TJ, Norfolk, England Royal Holloway Univ London, Sch Biol Sci, London, England Maynooth Univ, Dept Biol, Co, Kildare, Ireland Univ Bristol, Sch Biol Sci, Bristol BS8 1TQ, Avon, England CSIRO, Land &Water Flagship, Canberra, ACT, Australia Trin Coll Dublin, Sch Nat Sci, Dept Zool, Dublin, Ireland Zool Soc London, Inst Zool, London NW1 4RY, England ETH, Inst Integrat Biol, Theoret Biol, CH-8092 Zurich, Switzerland Swiss Inst Bioinformat, Lausanne, Switzerland Swiss Fed Inst Technol, Dept Biosyst Sci &Engn, Computat Evolut, Basel, Switzerland USDA ARS, Bee Res Lab, Washington, DC 20250 USA Univ Sao Paulo, Dept Genet, Fac Med Ribeirao Preto, BR-14040900 Ribeirao Preto, Brazil N Carolina State Univ, Ctr Res Sci Computat, Raleigh, NC 27695 USA Kyoto Univ, Grad Sch Agr, Lab Insect Ecol, Kyoto, Japan Univ Sao Paulo, Dept Biol Celular &Mol &Bioagentes Patogen, Fac Med Ribeirao Preto, BR-14040900 Ribeirao Preto, Brazil Inst Fed Educ Ciencia &Tecnol Sao Paulo, BR-15991502 Matao, Brazil Ohio State Univ, Dept Entomol, Wooster, OH 44791 USA Oxford Brookes Univ, Fac Hlth &Life Sci, Dept Biol &Med Sci, Oxford OX3 0BP, England Xian Jiaotong Liverpool Univ, Dept Biol Sci, Suzhou, Peoples R China Univ Hosp Halle Saale, Dept Lab Med, Halle, Germany German Ctr Integrat Biodivers Res iDiv, Leipzig, Germany Univ Southampton, Southampton, Hants, England Univ Leicester, Dept Biol, Leicester, Leics, England Univ Fed Sao Carlos, Ctr Ciencias Biol &Saude, Dept Genet &Evolucao, BR-13565905 Sao Carlos, SP, Brazil Univ Estadual Paulista, Fac Ciencias Agr &Vet, Dept Tecnol, BR-14884900 Jaboticabal, Brazil Univ N Carolina, Dept Biol, Greensboro, NC 27403 USA Natl Ctr Sci Res Demokritos, Inst Biosci &Applicat, Athens, Greece Univ Munich, Munich, Germany Baylor Coll Med, Dept Mol &Human Genet, Human Genome Sequencing Ctr, Houston, TX 77030 USA Univ Illinois, Roy J Carver Biotechnol, Urbana, IL 61801 USA Univ Illinois, Dept Crop Sci, Urbana, IL 61801 USA Univ Illinois, Inst Genom Biol, Urbana, IL 61801 USA MD Anderson Canc Ctr, Sch Hlth Profess, Mol Genet Technol Program, Unit 2, Houston, TX 77025 USA Univ Chicago, Dept Human Genet, Chicago, IL 60637 USA Univ Ghent, Dept Biochem &Microbiol, Lab Prot Biochem &Biomol Engn, B-9000 Ghent, Belgium Queen Mary Univ London, Sch Biol &Chem Sci, Dept Biol &Expt Psychol, London E1 4NS, England Guangzhou Univ, Sch Life Sci, Guangzhou, Guangdong, Peoples R China Johns Hopkins Univ, McKusick Nathans Inst Genet Med, Ctr Computat Biol, Baltimore, MD 21205 USA Univ Illinois, Neurosci Program, Dept Entomol, Carl R Woese Inst Genom Biol, Urbana, IL 61801 USA Departamento de Tecnologia, Faculdade de Ciências Agrárias e Veterinárias, Universidade Estadual Paulista, Jaboticabal, 14884-900, Brazi National Institutes of Health (NIH): DP1 OD006416 National Institutes of Health (NIH): U54 HG003273 Agriculture and Food Research Initiative Competitive grant from the USDA National Institute of Food and Agriculture: 2010-65106-21301 FAPESP: 11/03171-5 Marie Curie International Outgoing Fellowship: PIOF-GA-2011-303312 Swiss National Science Foundation: 31003A-125350 Swiss National Science Foundation: 31003A-143936

Trypanosomatids infecting honey bees have been poorly studied with molecular methods until recently. After the description of Crithidia mellificae (Langridge and McGhee, 1967) it took about forty years until molecular data for honey bee trypanosomatids became available and were used to identify and describe a new trypanosomatid species from honey bees, Lotmaria passim (Evans and Schwarz, 2014). However, an easy method to distinguish them without sequencing is not yet available. Research on the related bumble bee parasites Crithidia bombi and Crithidia expoeki revealed a fragment length polymorphism in the internal transcribed spacer 1 (ITS1), which enabled species discrimination. In search of fragment length polymorphisms for differential diagnostics in honey bee trypanosomatids, we studied honey bee trypanosomatid cell cultures of C. mellificae and L. passim. This research resulted in the identification of fragment length polymorphisms in ITS1 and ITS1-2 markers, which enabled us to develop a diagnostic method to differentiate both honey bee trypanosomatid species without the need for sequencing. However, the amplification success of the ITS1 marker depends probably on the trypanosomatid infection level. Further investigation confirmed that L. passim is the dominant species in Belgium, Japan and Switzerland. We found C. mellificae only rarely in Belgian honey bee samples, but not in honey bee samples from other countries. C. mellificae was also detected in mason bees (Osmia bicornis and Osmia cornuta) besides in honey bees. Further, the characterization and comparison of additional markers from L. passim strain SF (published as C. mellificae strain SF) and a Belgian honey bee sample revealed very low divergence in the 18S rRNA, ITS1-2, 28S rRNA and cytochrome b sequences. Nevertheless, a variable stretch was observed in the gp63 virulence factor.

Sperm length is highly variable, both between and within species, but the evolutionary significance of this variation is poorly understood. Sexual selection on sperm length requires a significant additive genetic variance, but few studies have actually measured this. Here we present the first estimates of narrow sense heritability of sperm length in a social insect, the bumblebee Bombus terrestris. In spite of a balanced and straightforward rearing design of colonies, and the possibility to replicate measurements of sperm within single males nested within colonies, the analysis proved to be complex. Several appropriate statistical models were derived, each depending on different assumptions. The heritability estimates obtained ranged from h (2) = 0.197 +/- 0.091 to h (2) = 0.429 +/- 0.154. All our estimates were substantially lower than previous estimates of sperm length heritability in non-social insects and vertebrates.

We report the details and characteristics of a total of 44 novel microsatelllite loci for Bombus spp. Most of them are highly polymorphic in Bombus terrestris, and a high degree of polymorphism is also found where these primers have been tested in 10 other bumblebee species. These markers will therefore be useful for the genetic study of this group.

The mating systems of social insects range from monandry, through low levels of polyandry to extremely high levels of polyandry. While the distribution of these mating systems across taxa is increasingly well-known, little is known about the proximate reasons behind the number of times social insect queens mate. Here we examined the role of mating duration and mating partner in predicting multiple mating in the naturally polyandrous bumble bee, Bombus hypnorum. We found that, as mating duration decreased, the likelihood of re-mating increased. Weak evidence suggested that male colony-of-origin may also influence re-mating. Dissections showed that males transferred a mating plug to queens. The plug does not last as long as in the congener B. terrestris, and males of different colonies varied in the allocation of resources to plug production. These results provide insight into the proximate factors behind multiple mating in bumble bees.

In this study we describe colony development, larval development, worker reproduction and mating frequency of the queen in laboratory colonies of Bombus impatiens Cresson. Comparison of our observations with data from B. terrestris - the best studied bumblebee - revealed both similarities and dissimilarities between the two. Colonies of B. impatiens have a high number of workers (374.5 ± 108), but relatively low number of young queens (9.4 ± 20) and males (8.9 ± 20). As in B. terrestris, size differences between the castes in B. impatiens were pronounced and became conspicuous as differences in the molting weight of the larvae, starting at the second instar. Workers are able to lay eggs and about 9% of the workers in old colonies had mature oocytes in their ovaries. Despite this, workers were almost never observed to lay eggs in the presence of the queen, and the "competition phase" which is known from B. terrestris was never observed. Mating frequency of the queen was examined using molecular techniques. Two out of ten queens had mated with 2 different males, suggesting some degree of polyandry in the population.

1. The Palaearctic Bombus ruderatus (in 1982/1983) and Bombus terrestris (1998) have both been introduced into South America (Chile) for pollination purposes. We here report on the results of sampling campaigns in 2004, and 2010–2012 showing that both species have established and massively expanded their range. 2. Bombus terrestris, in particular, has spread by some 200 km/year and had reached the Atlantic coast in Argentina by the end of 2011. Both species, and especially B. terrestris, are infected by protozoan parasites that seem to spread along with the imported hosts and spillover to native species. 3. Genetic analyses by polymorphic microsatellite loci suggest that the host population of B. terrestris is genetically diverse, as expected from a large invading founder population, and structured through isolation by distance. Genetically, the populations of the trypanosomatid parasite, Crithidia bombi, sampled in 2004 are less diverse, and distinct from the ones sampled later. Current C. bombi populations are highly heterozygous and also structured through isolation by distance correlating with the genetic distances of B. terrestris, suggesting the latter’s expansion to be a main structuring factor for the parasite. 4. Remarkably, wherever B. terrestris spreads, the native Bombus dahlbomii disappears although the reasons remain unclear. Our ecological and genetic data suggest a major invasion event that is currently unfolding in southern South America with disastrous consequences for the native bumblebee species., Die Eidgenössische Technische Hochschule Zürich, University of Stirling, Centro de Estudios Parasitológicos y de Vectores, Facultad de Ciencias Naturales y Museo, Pontificia Universidad Católica de Valparaiso

The mating frequency of females in social insects is particularly interesting, because polyandry reduces colony relatedness and increases within-colony genetic variance. It thereby affects a complex balance of benefits and costs that determine the degree of reproductive skew, sex allocation to offspring, or the opportunities for nepotism and policing strategies. Few systematic surveys of female mating frequencies exist and many are based on unreliable behavioural observation or sperm counts. Here, we report the results of a survey of mating frequencies in eight European Bombus spp. by means of highly polymorphic microsatellite loci. Only B. hypnorum was found to be multiply mated, while the pattern found in B. terrestris, B. lucorum, B. pratorum, B. lapidarius, B. sicheli, B. hortorum, and B. pascuorum was compatible with single mating. The findings are compatible with recent claims that, with some exceptions, mating frequencies of social insect females are generally low.

1 Whenever parasitism by more than one female occurs, larvae of parasitoids not only have to resist host defence but also face competition with other (unrelated) larvae. Competition is particularly important in solitary parasitoids where only one larva is able to complete its development. Such a situation is found in Conopidae (Diptera) parasitizing adult bumble bees where larvae of two species of conopid flies, Sicus ferrugineus L. and Physocephala rufipes F. often compete within the common host Bombus pascuorum Scopoli. This study analysed the larval development of the two species and asks how competition among larvae may be regulated. 2 Parasitized workers of B.pascuorum were caught in the field and kept according to different experimental schedules in the laboratory. This provided stage-structured data for the temporal course of development of the parasitic larvae. For the analysis, a simulation model was constructed that estimated the duration of all parasitic stages (Manly, 1990, first method). In both species the egg stage was found to be approximately 2 days, first instar 3 days, second instar 4 days, and third instar 3 days. The total development time is an estimated 10.8 days from oviposition in S.ferrugineus and 11.4 days in P.rufipes. S.ferrugineus develops faster in the beginning, probably because of its larger egg size, whereas P.rufipes pupates at larger size. First-instar larvae of both species possess strong, pointed mandibles. 3 The success of conopid larvae seems only marginally affected by host defence, for a single larva per host almost always completes development. Under competition, however, mortality rate increases substantially, and most larvae die in their first instar. Moreover, they show signs of melanization. The estimates for developmental times and the patterns found in this study suggest that conopid larvae seem capable of physical attacks, particularly during the first instar, when elimination of competitors is most common, and that S.ferrugineus has a time advantage because of its faster early development. Because most studies have previously been carried out with hymenopteran parasitoids, this study provides new information about the other large group of parasitoid insects, the Diptera, and demonstrates convergent patterns.

We here present an efficient, precise and reliable method to isolate and cultivate healthy and viable single Crithidia bombi cells from bumblebee faeces using flow cytometry. We report a precision of >99% in obtaining single trypanosomatid cells for further culture and analysis (“cloning”). In the study, we have investigated the use of different liquid media to cultivate C. bombi and present an optimal medium for obtaining viable clones from all tested, infected host donors. We show that this method can be applied to genotype a collection of clones from natural infections. Furthermore, we show how to cryo-preserve C. bombi cells to be revived to become infective clones after at least 4 years of storage. Considering the high prevalence of infections in natural populations, our method provides a powerful tool in studying the level and diversity of these infections, and thus enriches the current methodology for the studies of complex host-parasite interactions. ISSN:1932-6203

Mixed-genotype infections have major consequences for many essential elements of host-parasite interactions. With genetic exchange between co-infecting parasite genotypes increased diversity among parasite offspring and the emergence of novel genotypes from infected hosts is possible. We here investigated mixed- genotype infections using the host, Bombus spp. and its trypanosome parasite Crithidia bombi as our study case. The natural infections of C. bombi were genotyped with a novel method for a representative sample of workers and spring queens in Switzerland. We found that around 60% of all infected hosts showed mixed-genotype infections with an average of 2.47±0.22 (S.E.) and 3.65±1.02 genotypes per worker or queen, respectively. Queens, however, harboured up to 29 different genotypes. Based on the genotypes of co-infecting strains, these could be putatively assigned to either ‘primary’ and ‘derived’ genotypes - the latter resulting from genetic exchange among the primary genotypes. High genetic relatedness among co-infecting derived but not primary genotypes supported this scenario. Co-infection in queens seems to be a major driver for the diversity of genotypes circulating in host populations. ISSN:1932-6203

Behavioral Ecology, 5 (1), ISSN:1045-2249, ISSN:1465-7279

The breeding structure of protozoan infections, i.e. whether and how frequently parasites exchange genes (“sexual reproduction”), is a crucially important parameter for many important questions; it also matters for how new virulent strains might emerge. Whether protozoan parasites are clonal or sexual is therefore a hotly debated issue. For trypanosomatids, few experimental tests of breeding structure exist to date and are limited to the vector-borne human diseases Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major. We infected the natural host (Bombus terrestris) of the monoxenous parasite Crithidia bombi (Trypanosomatida) either with a single strain of the parasite or in mixed infections and tested for genetic exchange among co-infecting strains using microsatellite markers. We show that strains regularly exchange genetic material, with occasional self-crossing during mixed infections. Most offspring clones fit the expected allelic pattern from a standard Mendelian segregation. In some cases, alleles are lost or gained, leading to an entirely new genotype different from either parent. Genetic exchange in C. bombi therefore does occur and the process also leads to allelic loss or gain that could result from slippage during recombination. The majority of novel offspring types correspond to a recombination of parental alleles. The case of C. bombi demonstrates that directly transmitted, monoxenic trypanosomatids can also exchange genes. Sex therefore seems to be found in very different lineages of the trypanosomatids. Furthermore, the data allowed estimating a frequency at which C. bombi shows genetic exchange in populations.

Conopid flies (Conopidae, Diptera) are common larval parasites of bumblebees. The larva develops inside the abdomen of workers, queens and males. Development is completed within 10–12 days after oviposition when the host is killed and the parasite pupates in situ. Development results in parasitised bees becoming unable to carry large loads of nectar, as the conopid larvae reside where the honey crop is normally located. Furthermore, an addition to the bee's unloaden body mass is likely (average larval weight reached at pupation by the common parasite species Sicus ferrugineus: ±SD 36.3±12.3 mg, n=59; by Physocephala rufipes: 55.8±16.9 mg, n=108). We here asked whether the propensity of workers of the bumblebee Bombus pascuorum to collect nectar rather than pollen is related to the presence of conopid larvae. For samples of bees (n=2254 workers) collected over 3 years of field studies in northwestern Switzerland, there was no difference in the frequency of bees caught as pollen collectors among parasitised (38.1% of cases, n=210) as compared to non-parastised bees (43.9%, n=360) (χ2=1.83, n.s.). However, compared to the non-parasitised bees (n=360), those hosts containing a third (last) instar larva (n=9) were less likely to collect pollen than expected by chance χ2=6.91, P=0.003. Similarly, hosts with short survival time between capture and being killed by the developing larva (which hence must have harboured a late instar parasite at time of capture) were less likely to collect pollen (8%, n=25) than those found not parasitised (37.6%, n=891 χ2=9.16, P

We present field data on the ecology of a host-parasite system, consisting of several species of parasitoid flies (Conopidae, Diptera) and their bumblebee hosts (Bombini, Apoidea, Hymenoptera). Host animals were systematically sampled from different study sites throughout a season and checked for successful infestation in the form of puparia of these endoparasitic flies. Such dissection of the bees revealed that infestation occurs primarily during the summer months (June to September), with an observed maximum frequency of parasitization of 46.7% in workers in one of our study sites. On average, 13.2% of all workers (range 0–46.7%) and 7.1% of all males (range 0–28.6%) contained the puparium of a conopid. Two conopid generaSicus (64% of cases) andPhysocephala (36%) accounted for the infestation, with the latter being more abundant later in the year. A multivariate analysis identified host species, sex (male or worker), and study area as the most important factors accounting for the observed variance in the probability of being parasitized during the summer months. On average, males were less affected than workers. The marked seasonal appearance of conopids seems to account for differences among species, in particular for low levels of infestation among species completing their life cycles early (e.g.B. pratorum) and among the early flying, hibernated quens ofBombus andPsithyrus species. The results are discussd with respect to the impact of conopids on host ecology and evolution. Additional observations on the occurrence of further endoparasites (Sphaerularia bombi andSyntretus sp.) are reported.

Detecting functional homology between invertebrate and vertebrate immunity is of interest in terms of understanding the dynamics and evolution of immune systems. Trans-generational effects on immunity are well known from vertebrates, but their existence in invertebrates remains controversial. Earlier work on invertebrates has interpreted increased offspring resistance to pathogens as trans-generational immune priming. However, interpretation of these earlier studies involves some caveats and thus full evidence for a direct effect of maternal immune experience on offspring immunity is still lacking in invertebrates. Here we show that induced levels of antibacterial activity are higher in the worker offspring of the bumblebee, Bombus terrestris L., when their mother queen received a corresponding immune challenge prior to colony founding. This shows trans-generational immune priming in an insect, with ramifications for the evolution of sociality.

PUBLISHED, 1. Parasites can have dramatic effects on the ecology of their hosts. Such strong host-parasite interactions are the result of either parasites with generally high virulence, or generally benign parasites that nevertheless express context-dependent virulence. Theoretically, one indication that an apparently benign parasite nevertheless has a large impact on its host should be the existence of strong genotypic interactions between host and parasite. 2. Crithidia bombi (Trypanosomatidae) is a highly prevalent but generally benign gut parasite of the bumble bee Bombus terrestris. The demonstration of strong genotypic interactions between C. bombi and B. terrestris, however, suggests that context-dependent virulence may have a large impact on the host population. We thus investigated the effects of C. bombi across the entire life-cycle of its host, including the stressful times of hibernation and colony-founding. Due to the high prevalence and rates of transmission of the parasite in field populations, we used a large-scale laboratory experiment. 3. Under stressful hibernation, infected queens lost more weight. Infection also significantly reduced colony-founding success, colony size, male production and overall fitness, by up to 40%. These findings show that strong genotypic host-parasite interactions may indeed be a reliable indicator that apparently benign and highly prevalent parasites are nevertheless exerting a dramatic impact on their host populations.

PUBLISHED, Social insect colonies provide model systems for the examination of conflicts among parties with different genetic interests. As such, they have provided the best tests of inclusive fitness theory. However, much remains unknown about in which party?s favour such conflicts are resolved, partly due to the only recent advent of the molecular tools needed to examine the outcome of these conflicts. Two key conflicts in social insect colonies are over control of the reproductive sex ratio and the production of male offspring. Most studies have examined only one of these conflicts, but in reality they occur in tandem and may influence each other. Using microsatellite analyses, we examined the outcome of conflict over sex ratios and male production in the bumble bee, Bombus hypnorum. We determined the genotypes of mother queens, their mates and males for each of ten colonies. In contrast to other reports of mating frequency in this species, all of the queens were singly-mated. The population sex ratio was consistent with queen control, suggesting that queens are winning this conflict. In contrast, workers produced over 20% of all males in queen-right colonies, suggesting that they are more effective in competing over male-production. Combining our results with previous work, we suggest that worker-reproduction is a labile trait that may well impose only small costs on queen fitness.

Transmission to a new host is a critical step in the life cycle of a parasite. Variation in the characteristics of the transmission process, for example, due to host demography, is assumed to select for different variants of the parasite. We have experimentally tested how variation in the time to transmission (early or late after infection) and exposure to adverse conditions outside the host (immediate or delayed contact with new host) interact to determine the success of the infection in the next host, using the trypanosome Crithidia bombi infecting its bumblebee host, Bombus terrestris. These two experimentally manageable steps mimic the processes of within- and among-host selection for the parasite. We found that early transmission led to higher infection success in the next host as did immediate contact with the new host. However, there was no interaction between the two parameters as would be expected if early-transmitted variants, resulting from rapid multiplication within the host, would be less adapted to the conditions encountered during the between-host transfer or infection of the next host. Furthermore, typing the genetic variability of the parasites with microsatellites showed that the four different transmission routes of our experiment selected for different degrees of allelic diversity of the infecting parasite populations. The results support the idea that variation in the transmission process selects for different genotypic variants of the parasite. At the same time, the relationship of allelic diversity with infection intensity suggested that the coinfection model of May and Nowak (1995) may be appropriate, where each parasite is able to infect and multiply independent of others within the same host.

Two parasitoid flies,Physocephala rufipes andSicus ferrugineus (Diptera, Conopidae), and their hosts,Bombus spp., coexist at various locations in northwestern Switzerland. A detailed field study showed that both conopid species use the hostB. pascuorum to a similar degree, while the hostB. terr-luc (a pooled category ofB. terrestris andB. lucorum) is more frequently parasitised than expected byS. ferrugineus. The hostB. lapidarius in turn is exclusively used byP. rufipes. Furthermore, hosts ofB. terr-luc andB. pascuorum parasitised byS. ferrugineus were larger than hosts parasitised byP. rufipes, or than those not parasitised. The findings suggest thatS. ferrugineus selects larger hosts and may displaceP. rufipes. Pupal weight, a predictor of adult body size and parasitoid fecundity, is positively correlated with host size and larger pupae are more likely to emerge, while host species had no effect on the probability of emergence in either conopid species. Host species affected pupal weight inS. ferrugineus, but not inP. rufipes, althoughP. rufipes grew larger in hosts of a given size. Daughters emerged from larger pupae than males, but this did not correlate with larger host sizes. These observations add to the scarce knowledge of dipteran parasitoids.

Background The bumblebee Bombus terrestris is an ecologically and economically important pollinator and has become an important biological model system. To study fundamental evolutionary questions at the genomic level, a high resolution genetic linkage map is an essential tool for analyses ranging from quantitative trait loci (QTL) mapping to genome assembly and comparative genomics. We here present a saturated linkage map and match it with the Apis mellifera genome using homologous markers. This genome-wide comparison allows insights into structural conservations and rearrangements and thus the evolution on a chromosomal level. Results The high density linkage map covers ~ 93% of the B. terrestris genome on 18 linkage groups (LGs) and has a length of 2'047 cM with an average marker distance of 4.02 cM. Based on a genome size of ~ 430 Mb, the recombination rate estimate is 4.76 cM/Mb. Sequence homologies of 242 homologous markers allowed to match 15 B. terrestris with A. mellifera LGs, five of them as composites. Comparing marker orders between both genomes we detect over 14% of the genome to be organized in synteny and 21% in rearranged blocks on the same homologous LG. Conclusions This study demonstrates that, despite the very high recombination rates of both A. mellifera and B. terrestris and a long divergence time of about 100 million years, the genomes' genetic architecture is highly conserved. This reflects a slow genome evolution in these bees. We show that data on genome organization and conserved molecular markers can be used as a powerful tool for comparative genomics and evolutionary studies, opening up new avenues of research in the Apidae., BMC Genomics, 12, ISSN:1471-2164

Parasites of social insect workers can be transmitted within the colony to other, related host individuals or, alternatively, to unrelated workers of other colonies. Division of labour affects the probability of transmission, as young individuals often work inside the nest whereas older ones often leave the nest to forage. Therefore, the relative probabilities of transmission within-vs. between-nests is also affected by the delay between host infection and the shedding of propagules, i.e. the latent period of the parasite strain. We therefore hypothesized that strains of the flagellate parasite Crithidia bombi (Trypanosomatidae, Zoomastigophorea) infecting workers of the bumble bee Bombus terrestris (Hymenoptera, Apidae) could differ in their delays and coexist in a population. This would be the case if strains that are shed after a short time delay were more efficiently transmitted to other colony members, whereas strains with long delays were more efficiently transmitted to non-related workers in the population. We tested this hypothesis by experimentally varying time delay and by allowing transmission to either sister workers from the same nest or unrelated workers from other nests. Transmission of C. bombi was measured as the number of parasitic cells shed by the exposed workers after a standard period. The results showed that relatedness as such had no effect, but that delay and nest identity were highly significant effects to explain variation in transmission success. There was a significant interaction between nest identity and delay, such that bees of some colonies acted as efficient transmitters for C. bombi under short delays and vice versa. We discuss how division of labour may affect parasitism in social insects and, vice versa, how division of labour may be under selection from the effects of parasitism, using available evidence from the literature.