Dataset Information

Metagenomic detection of viral pathogens in Spanish honeybees: co-infection by Aphid Lethal Paralysis, Israel Acute Paralysis and Lake Sinai Viruses.

ABSTRACT: The situation in Europe concerning honeybees has in recent years become increasingly aggravated with steady decline in populations and/or catastrophic winter losses. This has largely been attributed to the occurrence of a variety of known and "unknown", emerging novel diseases. Previous studies have demonstrated that colonies often can harbour more than one pathogen, making identification of etiological agents with classical methods difficult. By employing an unbiased metagenomic approach, which allows the detection of both unexpected and previously unknown infectious agents, the detection of three viruses, Aphid Lethal Paralysis Virus (ALPV), Israel Acute Paralysis Virus (IAPV), and Lake Sinai Virus (LSV), in honeybees from Spain is reported in this article. The existence of a subgroup of ALPV with the ability to infect bees was only recently reported and this is the first identification of such a strain in Europe. Similarly, LSV appear to be a still unclassified group of viruses with unclear impact on colony health and these viruses have not previously been identified outside of the United States. Furthermore, our study also reveals that these bees carried a plant virus, Turnip Ringspot Virus (TuRSV), potentially serving as important vector organisms. Taken together, these results demonstrate the new possibilities opened up by high-throughput sequencing and metagenomic analysis to study emerging new diseases in domestic and wild animal populations, including honeybees.

SUBMITTER: Granberg F

PROVIDER: S-EPMC3583878 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Metagenomic detection of viral pathogens in Spanish honeybees: co-infection by Aphid Lethal Paralysis, Israel Acute Paralysis and Lake Sinai Viruses.

Granberg Fredrik F Vicente-Rubiano Marina M Rubio-Guerri Consuelo C Karlsson Oskar E OE Kukielka Deborah D Belák Sándor S Sánchez-Vizcaíno José Manuel JM

PloS one 20130227 2

The situation in Europe concerning honeybees has in recent years become increasingly aggravated with steady decline in populations and/or catastrophic winter losses. This has largely been attributed to the occurrence of a variety of known and "unknown", emerging novel diseases. Previous studies have demonstrated that colonies often can harbour more than one pathogen, making identification of etiological agents with classical methods difficult. By employing an unbiased metagenomic approach, which ...[more]

PMID: 23460860

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Project description:Lake Sinai Viruses (Sinaivirus) are commonly detected in honey bees (Apis mellifera) but no disease phenotypes or fitness consequences have yet been demonstrated. This viral group is genetically diverse, lacks obvious geographic structure, and multiple lineages can co-infect individual bees. While phylogenetic analyses have been performed, the molecular evolution of LSV has not been studied extensively. Here, I use LSV isolates from GenBank as well as contigs assembled from honey bee Sequence Read Archive (SRA) accessions to better understand the evolutionary history of these viruses. For each ORF, substitution rate variation, codon usage, and tests of positive selection were evaluated. Outlier regions of high or low diversity were sought with sliding window analysis and the role of recombination in creating LSV diversity was explored. Phylogenetic analysis consistently identified two large clusters of sequences that correspond to the current LSV1 and LSV2 nomenclature, however lineages sister to LSV1 were the most frequently detected in honey bee SRA accessions. Different expression levels among ORFs suggested the occurrence of subgenomic transcripts. ORF1 and RNA-dependent RNA polymerase had higher evolutionary rates than the capsid and ORF4. A hypervariable region of the ORF1 protein-coding sequence was identified that had reduced selective constraint, but a site-based model of positive selection was not significantly more likely than a neutral model for any ORF. The only significant recombination signals detected between LSV1 and LSV2 initiated within this hypervariable region, but assumptions of the test (single-frame coding and independence of substitution rate by site) were violated. LSV codon usage differed strikingly from that of honey bees and other common honey-bee viruses, suggesting LSV is not strongly co-evolved with that host. LSV codon usage was significantly correlated with that of Varroa destructor, however, despite the relatively weak codon bias exhibited by the latter. While codon usage between the LSV1 and LSV2 clusters was similar for three ORFs, ORF4 codon usage was uncorrelated between these clades, implying rapid divergence of codon use for this ORF only. Phylogenetic placement and relative abundance of LSV isolates reconstructed from SRA accessions suggest that detection biases may be over-representing LSV1 and LSV2 in public databases relative to their sister lineages.

Project description: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.

Dataset Information

Metagenomic detection of viral pathogens in Spanish honeybees: co-infection by Aphid Lethal Paralysis, Israel Acute Paralysis and Lake Sinai Viruses.

Publications

Metagenomic detection of viral pathogens in Spanish honeybees: co-infection by Aphid Lethal Paralysis, Israel Acute Paralysis and Lake Sinai Viruses.

Similar Datasets

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