The presence of identical deformed wing virus sequence variants in co-occurring Apis species in Northern Thailand may represent a potential epidemiological threat to native honey bees of Southeast Asia.

Widespread native honey bee species in South and East Asia (Apis cerana, Apis dorsata and Apis florea) and the imported western honey bee (Apis mellifera) share habitats and potentially also share pathogens. Chief among the threats facing A. mellifera in Europe and North America is deformed wing virus (DWV), including its two principal genotypes: A and B (DWV-A and DWV-B respectively). Though DWV-A has been recorded in Asia's native Apis species, it is not known if DWV-B, or both DWV-A and DWV-B, are currently widespread in Asia and, if so, whether viral transmission is primarily intraspecific or interspecific. This study aims to fill these knowledge gaps by (i) determining the DWV genotype in four co-occurring Apis host species using qPCR and (ii) inferring viral transmission between them using nucleotide sequences of DWV from Apis host species collected at three independent localities in Northern Thailand. We found DWV-A and -B in all four Apis species, the exotic A. mellifera and the native A. cerana, A. dorsata and A. florea. That DWV-A sequences were identical across Apis species at the same locality, with a similar pattern for DWV-B sequences, suggests that DWV's epidemiology is largely driven by ongoing interspecific transmission (spillover) of DWV across co-occurring native and exotic Apis species. Both genotypes of DWV represent a serious threat to Asia's exotic and native honey bee species.

Replication of honey bee-associated RNA viruses across multiple bee species in apple orchards of Georgia, Germany and Kyrgyzstan.

The essential ecosystem service of pollination is provided largely by insects, which are considered threatened by diverse biotic and abiotic global change pressures. RNA viruses are one such pressure, and have risen in prominence as a major threat for honey bees (Apis mellifera) and global apiculture, as well as a risk factor for other bee species through pathogen spill-over between managed honey bees and sympatric wild pollinator communities. Yet despite their potential role in global bee decline, the prevalence of honey bee-associated RNA viruses in wild bees is poorly known from both geographic and taxonomic perspectives. We screened members of pollinator communities (honey bees, bumble bees and other wild bees belonging to four families) collected from apple orchards in Georgia, Germany and Kyrgyzstan for six common honey bee-associated RNA virus complexes encompassing nine virus targets. The Deformed wing virus complex (DWV genotypes A and B) had the highest prevalence across all localities and host species and was the only virus complex found in wild bee species belonging to all four studied families. Based on amplification of negative-strand viral RNA, we found evidence for viral replication in wild bee species of DWV-A/DWV-B (hosts: Andrena haemorrhoa and several Bombus spp.) and Black queen cell virus (hosts: Anthophora plumipes, several Bombus spp., Osmia bicornis and Xylocopa spp.). Viral amplicon sequences revealed that DWV-A and DWV-B are regionally distinct but identical in two or more bee species at any one site, suggesting virus is shared amongst sympatric bee taxa. This study demonstrates that honey bee associated RNA viruses are geographically and taxonomically widespread, likely infective in wild bee species, and shared across bee taxa.

Characterization of a Molecular Clone of Deformed Wing Virus B.

Honey bees (Apis mellifera) play a crucial role in agriculture through their pollination activities. However, they have faced significant health challenges over the past decades that can limit colony performance and even lead to collapse. A primary culprit is the parasitic mite Varroa destructor, known for transmitting harmful bee viruses. Among these viruses is deformed wing virus (DWV), which impacts bee pupae during their development, resulting in either pupal demise or in the emergence of crippled adult bees. In this study, we focused on DWV master variant B. DWV-B prevalence has risen sharply in recent decades and appears to be outcompeting variant A of DWV. We generated a molecular clone of a typical DWV-B strain to compare it with our established DWV-A clone, examining RNA replication, protein expression, and virulence. Initially, we analyzed the genome using RACE-PCR and RT-PCR techniques. Subsequently, we conducted full-genome RT-PCR and inserted the complete viral cDNA into a bacterial plasmid backbone. Phylogenetic comparisons with available full-length sequences were performed, followed by functional analyses using a live bee pupae model. Upon the transfection of in vitro-transcribed RNA, bee pupae exhibited symptoms of DWV infection, with detectable viral protein expression and stable RNA replication observed in subsequent virus passages. The DWV-B clone displayed a lower virulence compared to the DWV-A clone after the transfection of synthetic RNA, as evidenced by a reduced pupal mortality rate of only 20% compared to 80% in the case of DWV-A and a lack of malformations in 50% of the emerging bees. Comparable results were observed in experiments with low infection doses of the passaged virus clones. In these tests, 90% of bees infected with DWV-B showed no clinical symptoms, while 100% of pupae infected with DWV-A died. However, at high infection doses, both DWV-A and DWV-B caused mortality rates exceeding 90%. Taken together, we have generated an authentic virus clone of DWV-B and characterized it in animal experiments.

The Gut Microbiota Can Provide Viral Tolerance in the Honey Bee.

Adult honey bees host a remarkably consistent gut microbial community that is thought to benefit host health and provide protection against parasites and pathogens. Currently, however, we lack experimental evidence for the causal role of the gut microbiota in protecting the Western honey bees (Apis mellifera) against their viral pathogens. Here we set out to fill this knowledge gap by investigating how the gut microbiota modulates the virulence of a major honey bee viral pathogen, deformed wing virus (DWV). We found that, upon oral virus exposure, honey bee survival was significantly increased in bees with an experimentally established normal gut microbiota compared to control bees with a perturbed (dysbiotic) gut microbiota. Interestingly, viral titers were similar in bees with normal gut microbiota and dysbiotic bees, pointing to higher viral tolerance in bees with normal gut microbiota. Taken together, our results provide evidence for a positive role of the gut microbiota for honey bee fitness upon viral infection. We hypothesize that environmental stressors altering honey bee gut microbiota composition, e.g., antibiotics in beekeeping or pesticides in modern agriculture, could interact synergistically with pathogens, leading to negative effects on honey bee health and the epidemiology and impact of their viruses.