Biology subjects

Ryabov, E. V.

Publications and source records attributed to Ryabov, E. V..

13 recordsLinked to original sources

bioRxiv2025

Emergence of a novel potato leafroll virus variant in Scotland and its genetic modulation by aphid and tuber transmission.

Potato leafroll virus (PLRV) is a major pathogen affecting potatoes worldwide. Since 2018, PLRV incidence increased in Scottish potato crops. Deep sequencing of PLRV in Scottish potato plants revealed the prevalence of a novel PLRV type which became predominant in 2023, displacing the phylogenetically distinct variants that have been present in the region since at least 1989. Analysis of the infection dynamics of the cDNA clone-derived PLRV isolates in potato plants indicated that the novel PLRV may accumulate to higher levels compared to the historic one. Analysis of the genetic diversity of PLRV in early and late field generations (FGs) of seed potatoes showed a significantly reduced genetic diversity of the PLRV structural genes in the early FGs, compared to the late FGs, while divergency of the non-structural genes remained similar across all FGs. Considering that late FGs are more likely to be infected with PLRV via tuber transmission, and early FGs via aphid transmission, these findings suggest that aphid transmission imposes a genetic bottleneck on the structural genes of PLRV, but not on its non-structural genes.

microbiology↗

bioRxiv2024

Nano Colonies: Rearing honey bee queens and their offspring in small laboratory arenas

Honey bees create complex societies of self-organized individuals in intricate colonies. Studies of honey bees are carried out in both the field and the laboratory. However, field research is encumbered by the difficulties of making reliable observations and environmental confounders. Meanwhile, laboratory trials produce data that are not field realistic as they lack key characteristics of a natural colony. Additionally, advances in honey bee research have been hindered without reliable methodology to rear queens in the laboratory. Here we provide a new system to reliably produce queens and worker brood in the laboratory and describe how this system fits with artificial insemination of queens as a step towards a continuous self-contained source of bees. The process creates a bridge between field research and laboratory trials and provides a secure system for contagious or regulated elements while maintaining many of the intrinsic characteristics of a honey bee colony.

zoology↗

bioRxiv2022

Host-driven temperature dependence of viral infection in honey bees

The temperature dependence of infection reflects changes in the performance of parasites and hosts. High temperatures (i.e., fever) often mitigate infection by favoring heat-tolerant hosts over heat-sensitive parasites. Honey bees exhibit an endothermic, colony-level temperature regulation that is exceptional among insects and favors resistance to several parasites. However, proliferation of viruses is heavily host-dependent, suggesting that viral infection could be linked to--not threatened by--optimum host function. To understand how temperature-driven changes in performance of viruses and hosts shape virus proliferation, we compared the temperature dependence of isolated viral enzyme activity, three honey bee traits, and infection of honey bee pupae. Viral enzyme activity varied by <2-fold over a >30 {degrees}C interval that spanned the temperatures typical of ectothermic insects and honey bees. In contrast, metrics of honey bee performance peaked at high ([≥] 35 {degrees}C) temperatures and were highly temperature-sensitive, with respiration varying 8-fold over a 20 {degrees}C interval and successful development requiring a narrow 8 {degrees}C temperature range. Although these results suggested that hosts would gain a relative advantage over viruses with increasing temperature, the temperature dependence of pupal infection matched that of pupal development, falling only near pupaes upper thermal limits. Our results reflect the host-dependent nature of virus proliferation, suggesting that infection is accelerated--not curtailed--by optimum host function, contradicting predictions of infection based on the relative performance of parasites and hosts, and suggesting tradeoffs between infection resistance and host survival. Despite a 98% reduction in infection at the upper end of the colony temperature range, the narrow thermal safety margin for honey bee development might preclude the effectiveness of fever for controlling viruses.

ecology↗

bioRxiv2022

Oversharing by honey bees and the spread of viruses

Deformed wing virus (DWV) is an emerging insect pathogen efficiently transmitted through communicable and vector-borne routes with Apis mellifera. Continual transmission of DWV between hosts and vectors is required for maintenance of the pathogen within the population, and this vector-host-pathogen system offers unique disease transmission dynamics for pathogen maintenance between vector and a social host. In a series of experiments, we study how vector-vector, host-host and host-vector transmission routes maintain DWV in a honey bee population. We found co-infestations on shared hosts allowed for movement of DWV from mite to mite. Additionally, two social behaviors of the honeybee, trophallaxis and cannibalization of pupae, provide routes for communicable transmission from bee to bee. Communicable circulation of the virus solely amongst hosts was then shown to act as a reservoir of DWV for naive Varroa to acquire and subsequently vector the pathogen. Our findings illustrate communicable transmission between hosts can maintain and influence the future acquisition and vectoring of a pathogen by a vector. There are a handful of other infectious diseases, both zoonotic and which impact human health, that have potentially similar transmission dynamics.

pathology↗

bioRxiv2022

The vectoring competence of the mite Varroa destructor for Deformed wing virus of honey bees is highly dynamic and affects survival of the mite.

The ectoparasitic mite, Varroa destructor and the viruses it vectors, including types A and B of Deformed wing virus (DWV), pose a major threat to honey bees, Apis mellifera. Analysis of 256 mites collected from the same set of colonies on five occasions from May to October 2021 showed that less than a half of them, 39.8% (95% confidence interval (CI): 34.0 - 46.0%), were able to induce an overt-level DWV infection with more than 109 viral genomes per bee in the pupa after 6 days of feeding, with both DWV-A and DWV-B being vectored at similar rates. To investigate the effect of the phoretic stage on the mites ability to vector DWV, the mites from two field collection events were divided into two groups, one of which was tested immediately for their infectiveness, and the other was kept with adult worker bees in cages for 12 days prior to testing their infectiveness. We found that while 39.2 % (95% CI: 30.0 - 49.1%) of the immediately tested mites induced overt-level DWV infections, 12-day phoretic passage significantly increased the infectiousness of mites to 89.8% (95% CI: 79.2 - 95.6%). It is likely that Varroa mites that survive brood interruptions in field colonies are increasingly infectious. We found that mite lifespan was significantly affected by the type of DWV it transmitted to pupae. The mites, which induced overt DWV-B but not DWV-A infection had an average lifespan of 15.5 days (95% CI: 11.8 - 19.2 days), which was significantly shorter than those of the mites which induced overt DWV-A but not DWV-B infection, with an average lifespan of 24.3 days (95% CI: 20.2 - 28.5), or the mites which did not induce high levels of DWV-A or DWV-B, with an average survival of 21.2 days (95% CI: 19.0 - 23.5 days). The mites which transmitted high levels of both DWV-A and DWV-B had an intermediate average survival of 20.5 days (95% CI: 15.1 - 25.9 days). The negative impact of DWV-B on mite survival could be a consequence of the ability of DWV-B, but not DWV-A to replicate in Varroa mites.

microbiology↗

bioRxiv2022

Promiscuous feeding across multiple honey bee hosts amplifies the vectorial capacity of Varroa destructor

Varroa destructor is a cosmopolitan pest and leading cause of colony loss of the European honey bee. Historically described as a competent vector of honey bee viruses, this arthropod vector is cause for the global pandemic of Deformed wing virus, now endemic in honeybee populations. Our work shows viral spread is driven by Varroa actively switching from one adult bee to another as they feed. Assays using fluorescent microspheres were used to show the movement of fluids in both directions between host and vector when Varroa feed. Therefore, Varroa could be in either an infectious or naive state dependent upon the disease status of their host. We tested this and confirm that the relative risk of a Varroa feeding was dependent on the infectiousness of their previous host. Varroa exhibit remarkable heterogeneity in their host switching behavior, with some Varroa switching infrequently while others switch at least daily. As a result, relatively few of the most active Varroa parasitize the majority of bees. This multiple feeding behavior has analogs in vectorial capacity models of other systems, where promiscuous feeding by individual vectors is a leading driver of vectorial capacity. We propose that the honeybee-Varroa relationship offers a unique opportunity to apply principles of vectorial capacity to a social organism, as virus transmission is both vectored and occurs through multiple host-to-host routes common to a crowded society.

animal behavior and cognition↗

bioRxiv2021

Beeporter: a high-throughput tool for non-invasive analysis of honey bee virus infection

Honey bees face numerous pests and pathogens but arguably none are as devastating as Deformed wing virus (DWV). Development of antiviral therapeutics and virus-resistant honey bee lines to control DWV in honey bees is slowed by the lack of a cost-effective high-throughput screening of DWV infection. Currently, analysis of virus infection and screening for antiviral treatments in bees and their colonies is tedious, requiring a well-equipped molecular biology laboratory and the use of hazardous chemicals. Here we utilize a cDNA clone of DWV tagged with green fluorescent protein (GFP) to develop the Beeporter assay, a method for detection and quantification of DWV infection in live honey bees. The assay involves infection of honey bee pupae by injecting a standardized DWV-GFP inoculum, followed by incubation for up to 44 hours. GFP fluorescence is recorded at intervals via commonly available long-wave UV light sources and a smartphone camera or a standard ultraviolet transilluminator gel imaging system. Nonlethal DWV monitoring allows high-throughput screening of antiviral candidates and a direct breeding tool for identifying honey bee parents with increased antivirus resistance. For even more rapid drug screening, we also describe a method for screening bees using 96-well trays and a spectrophotometer.

microbiology↗

bioRxiv2020

Pupal cannibalism by worker honey bees contributes to the spread of Deformed wing virus.

Transmission routes impact pathogen virulence and genetics, therefore comprehensive knowledge of these routes and their contribution to pathogen circulation is essential for understanding host-pathogen interactions and designing control strategies. Deformed wing virus (DWV), a principal viral pathogen of honey bees associated with increased honey bee mortality and colony losses, became highly virulent with the spread of its vector, the ectoparasitic mite Varroa destructor. Reproduction of Varroa mites occurs in capped brood cells and mite-infested pupae from these cells usually have high levels of DWV. The removal of mite-infested pupae by worker bees, Varroa Sensitive Hygiene (VSH), leads to cannibalization of pupae with high DWV loads, thereby offering an alternative route for virus transmission. We used genetically tagged DWV to investigate virus transmission to and between worker bees following pupal cannibalisation under experimental conditions. We demonstrated that cannibalization of DWV-infected pupae resulted in high levels of this virus in worker bees and that the acquired virus was then transmitted between bees via trophallaxis, allowing circulation of Varroa-vectored DWV variants without the mites. Despite the known benefits of hygienic behaviour, it is possible that higher levels of VSH activity may result in increased transmission of DWV via cannibalism and trophallaxis.

microbiology↗

bioRxiv2020

Development of a honey bee RNA virus vector based on the genome of Deformed wing virus.

We developed a honey bee RNA-virus vector based on the genome of a picorna-like Deformed wing virus (DWV), the main viral pathogen of the honey bee (Apis mellifera). To test the potential of DWV to be utilized as a vector, the 717 nt sequence coding for the enhanced green fluorescent protein (eGFP), flanked by the peptides targeted by viral protease, was inserted into an infectious cDNA clone of DWV in-frame between the leader protein and the virus structural protein VP2 genes. The in vitro RNA transcripts from egfp-tagged DWV cDNA clones were infectious when injected into honey bee pupae. Stable DWV particles containing genomic RNA of the recovered DWV with egfp inserts were produced, as evidenced by cesium chloride density gradient centrifugation. These particles were infectious to honey bee pupae when injected intra-abdominally. Fluorescent microscopy showed GFP expression in the infected cells and Western blot analysis demonstrated accumulation of free eGFP rather than its fusions with DWV LP and/or VP2 proteins. Analysis of the progeny egfp-tagged DWV showed gradual accumulation of genome deletions for egfp, providing estimates for the rate of loss of a non-essential gene an insect RNA virus genome during natural infection.

microbiology↗

bioRxiv2019

Deformed Wing Virus spillover from honey bees to bumble bees: a reverse genetic study.

Deformed wing virus (DWV) is a persistent pathogen of European honey bees and the major contributor to overwintering colony losses. The prevalence of DWV in honey bees has led to significant concerns about spillover of the virus to other pollinating species. Bumble bees are both a major group of wild and commercially-reared pollinators. Several studies have reported pathogen spillover of DWV from honey bees to bumble bees, but evidence of a true sustained viral infection has yet to be demonstrated. Here we investigate the infectivity and transmission of DWV in bumble bees using the buff-tailed bumble bee Bombus terrestris as a model. We apply a reverse genetics approach combined with controlled laboratory conditions to detect and monitor DWV infection. A novel reverse genetics system for three representative DWV variants, including the two master variants of DWV - type A and B - was used. Our results directly confirm DWV replication in bumble bees but also demonstrate striking resistance to infection by certain routes. Bumble bees may support DWV replication but it is not clear how infection could occur under natural environmental conditions.

microbiology↗

bioRxiv2019

A novel system for maintaining Varroa destructor mites on artificial diets and its application for studying mites as a vector for honey bee viruses.

The mite Varroa destructor is one of the most destructive parasites of the honey bee (Apis mellifera) and the primary cause of colony collapse in most regions of the world. These mites cause serious injury to their hosts, especially during the larval and pupal stages, and serve as the vector for several viruses, which affect honey bee health causing colony death. Attempts by beekeepers to control these mites have yielded limited success. The inability to rear populations of mites in vitro that excludes contact with their honey bee hosts has stymied research of Varroa biology. Previous attempts to rear and/or maintain Varroa mites in vitro by feeding them on artificial diets have had limited success. Several methods were plagued by mechanical failures including leaking membranes and, thus far, none have been widely adopted. Here we report a robust system for maintaining Varroa mites that includes an artificial diet, which does not contain honey bee tissue-derived components, thus making it particularly valuable in studying mite vectoring of honey bee viruses. With our system we demonstrated for the first time that Varroa mites maintained on an artificial diet supplemented with the particles of honey bee viruses, cDNA clone-derived genetically tagged Varroa destructor virus-1 and wild-type Deformed wing virus, can acquire and later transmit these viruses to recipient honey bee pupae. Along with providing an opportunity to study parasites and pathogens in the absence of honey bee hosts, this in vitro system for Varroa mite maintenance is both scalable and consistent. These features can be used to better understand mite nutritional needs, metabolic activity, responses to chemicals and other biological functions.

zoology↗

bioRxiv2019

Deformed wing virus type A, a major honey bee pathogen, is vectored by the mite Varroa destructor in a non-propagative manner.

Honey bees, the primary managed insect pollinator, suffer considerable losses due to Deformed wing virus (DWV), an RNA virus vectored by the mite Varroa destructor. Mite vectoring has resulted in the emergence of virulent DWV variants. The basis for such changes in DWV is poorly understood. Most importantly, it remains unclear whether replication of DWV occurs in the mite. In this study, we exposed Varroa mites to DWV type A via feeding on artificially infected honey bees. A significant, 357-fold increase in DWV load was observed in these mites after 2 days. However, after 8 additional days of passage on honey bee pupae with low viral loads, the DWV load dropped by 29-fold. This decrease significantly reduced the mites ability to transmit DWV to honey bees. Notably, negative-strand DWV RNA, which could indicate viral replication, was detected only in mites collected from pupae with high DWV levels but not in the passaged mites. We also found that Varroa mites contain honey bee mRNAs, consistent with the acquisition of honey bee cells which would additionally contain DWV replication complexes with negative-strand DWV RNA. We propose that transmission of DWV type A by Varroa mites occurs in a non-propagative manner.

microbiology↗

bioRxiv2019

Dynamic evolution in the key honey bee pathogen Deformed wing virus

The impacts of invertebrate RNA virus population dynamics on virulence and infection outcomes are poorly understood. Deformed wing virus (DWV), the main viral pathogen of honey bees, negatively impacts bee health which can lead to colony death. Despite previous reports on the reduction of DWV diversity following the arrival of the parasitic mite Varroa destructor, the key DWV vector, we found high genetic diversity of DWV in infested United States (US) honey bee colonies. Phylogenetic analysis showed that the divergent US DWV genotypes are of monophyletic origin, which were likely generated as a result of diversification after a genetic bottleneck. To investigate the population dynamics of this divergent DWV, we designed a series of novel infectious cDNA clones corresponding to co-existing DWV genotypes, thereby devising a reverse genetic system for an invertebrate RNA virus quasispecies. Equal replication rates were observed for all clone-derived DWV variants in single infections. Surprisingly, individual clones replicated to the same high levels as their mixtures and even the parental highly diverse natural DWV population, suggesting that complementation between genotypes was not required to replicate to high levels. Mixed clone-derived infections showed a lack of strong competitive exclusion, suggesting that the DWV genotypes were adapted to co-exist. Mutational and recombination events were observed across clone progeny providing new insights into the forces that drive and constrain virus diversification. Accordingly, herein we propose a new model of Varroa-induced DWV dynamics whereby an initial selective sweep is followed by virus diversification fueled by negative frequency-dependent selection for new genotypes. This selection likely reflects the ability of rare lineages to evade host defenses, specifically antiviral RNA interference (RNAi). In support of this, we show that RNAi induced against one DWV strain is less effective against an alternate strain from the same population.\n\nAuthor SummaryVirulence of Deformed wing virus (DWV), a major pathogen of honey bees, showed a sharp and significant increase following the introduction of its vector, the mite Varroa destructor. Varroa vectoring resulted in genetic changes of DWV, including reduction of DWV diversity to nearly clonal levels in the UK and Hawaii. Contrary to the previous reports, we discovered that virulent DWV populations circulating across the Varroa-infested United States included many divergent genotypes generated following a strong bottleneck event. We designed a series of the full-length infectious cDNA clones that captured the diversity of a typical virulent DWV population from a declining Varroa-infested colony, effectively establishing first reverse genetic system for an invertebrate RNA virus quasispecies, in order to investigate interactions between the virus genotypes. We demonstrated that individual co-existing DWV genotypes and diverse natural DWV populations replicated equally well indicating that complementation between isolates was not required to enable DWV replication to high levels. Also, no obvious competitive exclusion was detected between genotypes in mixed infections suggesting DWV genotypes are adapted to co-exist to maintain overall population diversity. We suggest that introduction of Varroa resulted in an initial selective sweep of DWV diversity which was followed by DWV diversification driven by selection for new genotypes capable of evading host defenses, specifically antiviral RNA interference.

microbiology↗