Antagonistic interactions between honey bee bacterial symbionts and implications for disease

BACKGROUND: Honey bees, Apis mellifera, face many parasites and pathogens and consequently rely on a diverse set of individual and group-level defenses to prevent disease. One route by which honey bees and other insects might combat disease is through the shielding effects of their microbial symbionts. Bees carry a diverse assemblage of bacteria, very few of which appear to be pathogenic. Here we explore the inhibitory effects of these resident bacteria against the primary bacterial pathogen of honey bees, Paenibacillus larvae. RESULTS: Here we isolate, culture, and describe by 16S rRNA and protein-coding gene sequences 61 bacterial isolates from honey bee larvae, reflecting a total of 43 distinct bacterial taxa. We culture these bacteria alongside the primary larval pathogen of honey bees, Paenibacillus larvae, and show that many of these isolates severely inhibit the growth of this pathogen. Accordingly, symbiotic bacteria including those described here are plausible natural antagonists toward this widespread pathogen. CONCLUSION: The results suggest a tradeoff in social insect colonies between the maintenance of potentially beneficial bacterial symbionts and deterrence at the individual and colony level of pathogenic species. They also provide a novel mechanism for recently described social components behind disease resistance in insect colonies, and point toward a potential control strategy for an important bee disease

Differential gene expression in Varroa jacobsoni mites following a host shift to European honey bees (Apis mellifera)

Background: Varroa mites are widely considered the biggest honey bee health problem worldwide. Until recently, Varroa jacobsoni has been found to live and reproduce only in Asian honey bee (Apis cerana) colonies, while V. destructor successfully reproduces in both A. cerana and A. mellifera colonies. However, we have identified an island population of V. jacobsoni that is highly destructive to A. mellifera, the primary species used for pollination and honey production. The ability of these populations of mites to cross the host species boundary potentially represents an enormous threat to apiculture, and is presumably due to genetic variation that exists among populations of V. jacobsoni that influences gene expression and reproductive status. In this work, we investigate differences in gene expression between populations of V. jacobsoni reproducing on A. cerana and those either reproducing or not capable of reproducing on A. mellifera, in order to gain insight into differences that allow V. jacobsoni to overcome its normal species tropism. Results: We sequenced and assembled a de novo transcriptome of V. jacobsoni. We also performed a differential gene expression analysis contrasting biological replicates of V. jacobsoni populations that differ in their ability to reproduce on A. mellifera. Using the edgeR, EBSeq and DESeq R packages for differential gene expression analysis, we found 287 differentially expressed genes (FDR ≤ 0.05), of which 91% were up regulated in mites reproducing on A. mellifera. In addition, mites found reproducing on A. mellifera showed substantially more variation in expression among replicates. We searched for orthologous genes in public databases and were able to associate 100 of these 287 differentially expressed genes with a functional description. Conclusions: There is differential gene expression between the two mite groups, with more variation in gene expression among mites that were able to reproduce on A. mellifera. A small set of genes showed reduced expression in mites on the A. mellifera host, including putative transcription factors and digestive tract developmental genes. The vast majority of differentially expressed genes were up-regulated in this host. This gene set showed enrichment for genes associated with mitochondrial respiratory function and apoptosis, suggesting that mites on this host may be experiencing higher stress, and may be less optimally adapted to parasitize it. Some genes involved in reproduction and oogenesis were also overexpressed, which should be further studied in regards to this host shift. © 2016 The Author(s)

Varroa destructor is an effective vector of Israeli acute paralysis virus in the honeybee, Apis mellifera.

The Israeli acute paralysis virus (IAPV) is a significant marker of honeybee colony collapse disorder (CCD). In the present work, we provide the first evidence that Varroa destructor is IAPV replication-competent and capable of vectoring IAPV in honeybees. The honeybees became infected with IAPV after exposure to Varroa mites that carried the virus. The copy number of IAPV in bees was positively correlated with the density of Varroa mites and time period of exposure to Varroa mites. Further, we showed that the mite-virus association could possibly reduce host immunity and therefore promote elevated levels of virus replication. This study defines an active role of Varroa mites in IAPV transmission and sheds light on the epidemiology of IAPV infection in honeybees

Species-specific diagnostics of Apis mellifera trypanosomatids: A nine-year survey (2007-2015) for trypanosomatids and microsporidians in Serbian honey bees

In this study, honey bees collected in Serbia over 9 consecutive years (2007-2015) were retrospectively surveyed to determine the prevalence of eukaryotic gut parasites by molecular screening of archival DNA samples. We developed species-specific primers for PCR to detect the two known honey bee trypanosomatid species, Crithidia mellificae and the recently described Lotmaria passim. These primers were validated for target specificity under single and mixed-species conditions as well as against the bumblebee trypanosomatid Crithidia bombi. Infections by Nosema apis and Nosema ceranae (Microsporidia) were also determined using PCR. Samples from 162 colonies (18 from each year) originating from 57 different localities were surveyed. L. passim was detected in every year with an overall frequency of 62.3% and annual frequencies ranging from 38.9% to 83.3%. This provides the earliest confirmed record to date for L. passim and the first report of this species in Serbia. N. ceranae was ubiquitous, occurring in every year and at 95.7% overall frequency, ranging annually from 83.3% to 100%. The majority of colonies (60.5%) were co-infected with L. passim and N. ceranae, but colony infections by each species were statistically independent of one another over the nine years. Although C. mellificae and N. apis have both been reported recently at low frequency in Europe, neither of these species was detected in Serbia. These results support the hypothesis that L. passim has predominated over C mellificae in A. mellifera during the past decade

Genome Characterization, Prevalence and Distribution of a Macula-Like Virus from Apis mellifera and Varroa destructor

Around 14 distinct virus species-complexes have been detected in honeybees, each with one or more strains or sub-species. Here we present the initial characterization of an entirely new virus species-complex discovered in honeybee (Apis mellifera L.) and varroa mite (Varroa destructor) samples from Europe and the USA. The virus has a naturally poly-adenylated RNA genome of about 6500 nucleotides with a genome organization and sequence similar to the Tymoviridae (Tymovirales; Tymoviridae), a predominantly plant-infecting virus family. Literature and laboratory analyses indicated that the virus had not previously been described. The virus is very common in French apiaries, mirroring the results from an extensive Belgian survey, but could not be detected in equally-extensive Swedish and Norwegian bee disease surveys. The virus appears to be closely linked to varroa, with the highest prevalence found in varroa samples and a clear seasonal distribution peaking in autumn, coinciding with the natural varroa population development. Sub-genomic RNA analyses show that bees are definite hosts, while varroa is a possible host and likely vector. The tentative name of Bee Macula-like virus (BeeMLV) is therefore proposed. A second, distantly related Tymoviridae-like virus was also discovered in varroa transcriptomes, tentatively named Varroa Tymo-like virus (VTLV)

An Accurate, Easy to Use Abundace Scale for Globular Clusters Based on 2.2um Spectra of Giant Stars

We present a new method for the determination of [Fe/H] for globular clusters. This new method is based on moderate resolution (R~1500) near-IR spectroscopy in the K-band of 6 to 10 of the brightest giants in a cluster. Our calibration is derived from spectra of 105 stars in 15 globular clusters. From measurements of the equivalent widths of three features in these spectra, Na, Ca, and CO, we are able to reproduce the Zinn & West (1984) abundance scale as updated by Harris (1996) to better than 0.10 dex for clusters with near solar [Fe/H] down to an [Fe/H] of -1.8. Three advantages of this method are that it can be used for metal rich, heavily reddened globulars in crowded fields, it does not require any knowledge of any other cluster or stellar parameters such as reddening, distance, or luminosity, and it requires only minimal telescope time. If stellar (J-K)0 and MK values are available as well, the accuracy of the [Fe/H] estimate is further improved. Observations of as few as three stars per cluster still gives an [Fe/H] estimate wich is nearly as reliable as that based on two to three times as many stars. The accuracy of an [Fe/H] value based on observations of CO absorption alone is significantly less than that which results from the three spectroscopic indices. However, we predict that space-based observations of this feature in the integrated light of stellar systems will prove to be of great value for abundance determinations at distances as far as the Coma cluster of galaxies.Comment: 53 pages, 16 Postscript figures. Submitted to the A

An updated genetic marker for detection of Lake Sinai Virus and metagenetic applications

Background. Lake Sinai Viruses (LSV) are common RNA viruses of honey bees (Apis mellifera) that frequently reach high abundance but are not linked to overt disease. LSVs are genetically heterogeneous and collectively widespread, but despite frequent detection in surveys, the ecological and geographic factors structuring their distribution in A. mellifera are not understood. Even less is known about their distribution in other species. Better understanding of LSV prevalence and ecology have been hampered by high sequence diversity within the LSV clade. Methods. Here we report a new polymerase chain reaction (PCR) assay that is compatible with currently known lineages with minimal primer degeneracy, producing an expected 365 bp amplicon suitable for end-point PCR and metagenetic sequencing. Using the Illumina MiSeq platform, we performed pilot metagenetic assessments of three sample sets, each representing a distinct variable that might structure LSV diversity (geography, tissue, and species). Results. The first sample set in our pilot assessment compared cDNA pools from managed A. mellifera hives in California (n = 8) and Maryland (n = 6) that had previously been evaluated for LSV2, confirming that the primers co-amplify divergent lineages in real-world samples. The second sample set included cDNA pools derived from different tissues (thorax vs. abdomen, n = 24 paired samples), collected from managed A. mellifera hives in North Dakota. End-point detection of LSV frequently differed between the two tissue types; LSV metagenetic composition was similar in one pair of sequenced samples but divergent in a second pair. Overall, LSV1 and intermediate lineages were common in these samples whereas variants clustering with LSV2 were rare. The third sample set included cDNA from individual pollinator specimens collected from diverse landscapes in the vicinity of Lincoln, Nebraska. We detected LSV in the bee Halictus ligatus (four of 63 specimens tested, 6.3%) at a similar rate as A. mellifera (nine of 115 specimens, 7.8%), but only one H. ligatus sequencing library yielded sufficient data for compositional analysis. Sequenced samples often contained multiple divergent LSV lineages, including individual specimens. While these studies were exploratory rather than statistically powerful tests of hypotheses, they illustrate the utility of high-throughput sequencing for understanding LSV transmission within and among species