Protecting bees from viruses
Lead Research Organisation:
The Pirbright Institute
Department Name: Arthropod Genetics
Abstract
Honey bees are crucial for pollination of food crops and production of hive products such as honey. In the past two decades, honey bees have suffered from declines across Europe and North America. This is due to a number of incursions of non-native pests and pathogens which damage honey bees, but also due to a change in the prevalence and virulence in viruses posited to be induced by the introduced parasitic mite, Varroa destructor. There are no controls for these viruses and management of Varroa is challenging due to widespread resistance to chemical pesticides and few new products available for this market.
Wolbachia are intracellular bacteria which live inside the cells of a large number of insect species, including species of wasps, bees and a small number of honey bees, and even some spider and worm species. Wolbachia are widespread in the UK. Some strains of this bacteria can make their host resistant to certain pathogenic viruses. This has been exploited by researchers working with the mosquitoes that transmit Dengue, Zika and Chikungunya viruses; in laboratory and field trials, the bacteria made the mosquitoes resistant to these viruses. Importantly, some strains of Wolbachia that make their hosts resistant to viruses do not appear to cause a serious fitness penalty to their insect host.
Our central hypothesis, based on these clear data from other insect species, is that infection with appropriate strains of Wolbachia will make honey bees resistant to important honey bee viruses; these are related to viruses known to be inhibited by Wolbachia. We will test this hypothesis in cultured bee cells, which allows more rapid testing of Wolbachia-virus combinations than using bees. We will also explore effects on the microsporidian parasite, Nosema, as there is some evidence that Wolbachia can also inhibit cellular pathogens. In addition to testing Wolbachia in honey bee cell lines, we can perform direct studies on individual honey bees in laboratory cages. In mosquitoes, adults can be injected with a purified strain of Wolbachia, which will disseminate through the tissues, this is called a 'transient' infection. These insects can then be challenged with viruses to get an indication of the effects of Wolbachia infection. Adult honey bees will be challenged in the same way in a contained environment, as a complementary approach for the potential anti-viral or anti-microsporidian activity of Wolbachia.
We propose to investigate the feasibility of using Wolbachia to inhibit positive sense RNA viruses, using a honey bee cell line as initial proof of concept. We will deliver this through four linked objectives:
Obj1:Set up cell culture and establish Wolbachia infected and Wolbachia-free derivatives
Obj2:Set up viral and Nosema cultures in cell lines
Obj3:Test pathogen infection on Wolbachia infected and Wolbachia free cell lines
Obj4:Validate and test transient Wolbachia infections of worker honey bees
There is a clear need for new methods for control of viral and cellular pathogens in bees. This project aims to explore the potential of a Wolbachia-based, non-chemical approach that, if realised, would potentially fit well into an enhanced, sustainable integrated bee health programme. If Wolbachia can inhibit viruses and/or Nosema spp. in honey bees, this would provide an exciting new approach to control of bee diseases; this project will test the feasibility of this approach. Wolbachia is transmitted through the female germline, so Wolbachia infected queen bees could be used to generate disease resistant colonies. This Wolbachia-based approach does not involve the use of recombinant DNA methods and the resulting strains are not considered to be genetically modified organisms (GMOs), providing lower barriers to any eventual field use than some other potential genetic approaches to pathogen resistance.
Wolbachia are intracellular bacteria which live inside the cells of a large number of insect species, including species of wasps, bees and a small number of honey bees, and even some spider and worm species. Wolbachia are widespread in the UK. Some strains of this bacteria can make their host resistant to certain pathogenic viruses. This has been exploited by researchers working with the mosquitoes that transmit Dengue, Zika and Chikungunya viruses; in laboratory and field trials, the bacteria made the mosquitoes resistant to these viruses. Importantly, some strains of Wolbachia that make their hosts resistant to viruses do not appear to cause a serious fitness penalty to their insect host.
Our central hypothesis, based on these clear data from other insect species, is that infection with appropriate strains of Wolbachia will make honey bees resistant to important honey bee viruses; these are related to viruses known to be inhibited by Wolbachia. We will test this hypothesis in cultured bee cells, which allows more rapid testing of Wolbachia-virus combinations than using bees. We will also explore effects on the microsporidian parasite, Nosema, as there is some evidence that Wolbachia can also inhibit cellular pathogens. In addition to testing Wolbachia in honey bee cell lines, we can perform direct studies on individual honey bees in laboratory cages. In mosquitoes, adults can be injected with a purified strain of Wolbachia, which will disseminate through the tissues, this is called a 'transient' infection. These insects can then be challenged with viruses to get an indication of the effects of Wolbachia infection. Adult honey bees will be challenged in the same way in a contained environment, as a complementary approach for the potential anti-viral or anti-microsporidian activity of Wolbachia.
We propose to investigate the feasibility of using Wolbachia to inhibit positive sense RNA viruses, using a honey bee cell line as initial proof of concept. We will deliver this through four linked objectives:
Obj1:Set up cell culture and establish Wolbachia infected and Wolbachia-free derivatives
Obj2:Set up viral and Nosema cultures in cell lines
Obj3:Test pathogen infection on Wolbachia infected and Wolbachia free cell lines
Obj4:Validate and test transient Wolbachia infections of worker honey bees
There is a clear need for new methods for control of viral and cellular pathogens in bees. This project aims to explore the potential of a Wolbachia-based, non-chemical approach that, if realised, would potentially fit well into an enhanced, sustainable integrated bee health programme. If Wolbachia can inhibit viruses and/or Nosema spp. in honey bees, this would provide an exciting new approach to control of bee diseases; this project will test the feasibility of this approach. Wolbachia is transmitted through the female germline, so Wolbachia infected queen bees could be used to generate disease resistant colonies. This Wolbachia-based approach does not involve the use of recombinant DNA methods and the resulting strains are not considered to be genetically modified organisms (GMOs), providing lower barriers to any eventual field use than some other potential genetic approaches to pathogen resistance.
Technical Summary
The Aim of the project is to explore the use of Wolbachia as a route to developing virus-resistant bees (European honey bee, Apis mellifera). Wolbachia infection has been shown to strongly inhibit many viruses and some cellular pathogens in a range of insects, leading us to hypothesise that protective effects may be elicited in honey bees against important bee pathogens. We will
Obj 1 Develop methods for establishing Wolbachia infections in cultured honey bee cells; We will obtain a stable honey bee cell line (AmE-711) from our collaborator and introduce selected Wolbachia using the shell vial technique. We will use quantitative real-time PCR to track the Wolbachia infection rates over multiple passages and select for lines with stable, high infection rates of Wolbachia.
Obj 2 Develop cell-culture based methods for amplifying key viruses (Deformed Wing Virus, DMV, and Chronic Bee Paralysis Virus (CBPV)) and also the microsporidian Nosema ceranae. Develop real-time PCR-based methods to monitor pathogen replication.
Obj 3 Based on Obj1&2, compare pathogen infection and replication in cells with and without Wolbachia to determine the effect of each of our Wolbachia strains.
Obj4 In parallel we will develop a transient infection method for honey bees, similar to that developed for several mosquito species, which will allow us to investigate host-Wolbachia-virus effects in vivo as well as in cultured cells. Transient infections of Wolbachia will be performed through adult injections and tested for infectivity. If successful pathogens will be administered using established techniques and the effect of Wolbachia infection on pathogen infection replication determined.
Obj 1 Develop methods for establishing Wolbachia infections in cultured honey bee cells; We will obtain a stable honey bee cell line (AmE-711) from our collaborator and introduce selected Wolbachia using the shell vial technique. We will use quantitative real-time PCR to track the Wolbachia infection rates over multiple passages and select for lines with stable, high infection rates of Wolbachia.
Obj 2 Develop cell-culture based methods for amplifying key viruses (Deformed Wing Virus, DMV, and Chronic Bee Paralysis Virus (CBPV)) and also the microsporidian Nosema ceranae. Develop real-time PCR-based methods to monitor pathogen replication.
Obj 3 Based on Obj1&2, compare pathogen infection and replication in cells with and without Wolbachia to determine the effect of each of our Wolbachia strains.
Obj4 In parallel we will develop a transient infection method for honey bees, similar to that developed for several mosquito species, which will allow us to investigate host-Wolbachia-virus effects in vivo as well as in cultured cells. Transient infections of Wolbachia will be performed through adult injections and tested for infectivity. If successful pathogens will be administered using established techniques and the effect of Wolbachia infection on pathogen infection replication determined.
Planned Impact
In the past two decades, honey bees have suffered from declines across Europe and North America. This is due to a number of incursions of non-native pests and pathogens which damage honey bees, but also due to a change in the prevalence and virulence in viruses posited to be induced by the introduced parasitic mite, Varroa destructor. There are no controls for these viruses and management of Varroa is challenging due to widespread resistance to chemical pesticides and few new products available for this market. A potential new route to pathogen tolerance in bees will therefore be of great interest to a wide range of stakeholders beyond academic researchers (for whom see 'academic beneficiaries' above), including apicultural communities, the many farmers large and small who depend on bee-delivered pollination services, and those interested in sustainable agriculture and food security more generally.
The Pirbright Institute and the PI have extensive experience of successful commercialisation of novel technologies. Though this project does not set out to develop stably-transfected lines of bees, it lays the foundation for such future work both in terms of resources such as Wolbachia-infected bee cells, and data, such as the relative effects of different strains of Wolbachia. There are clear commercial uses if strains of bees showing beneficial traits were to be produced. Correspondingly, these data and resources will be of significant value to commercial enterprises such as queen rearing in the honey bee industry. We will therefore consider the commercial sensitivity of these data prior to publication; the IP generated in this project will be identified and appropriately exploited. Since current GM legislation is uncertain and expensive in terms of time-to-market, at least in the UK and Europe, applications leading to non-GMO products are likely to be of more interest, and we have taken this into account in developing the project. Beneficial insect companies have showed interest in insect genetic technology, and the idea of this research; we aim to gain more detailed understanding of relevant commercial needs, and possibly develop formal collaborations, over the course of the project.
This research will be of interest to beekeepers and bee farmers across the UK. We will communicate directly with interested stakeholders, e.g. through attendance and presentation at apiculture conferences, such as the National Honey show and the BBKA convention. We have established contacts allowing us to engage with the beekeeping community on this research. The Pirbright Institute and the scientists involved have a strong track record in communicating to both scientific and lay audiences, and of taking novel applied genetic approaches from initial conception through to successful commercialisation and dissemination.
The Pirbright Institute and the PI have extensive experience of successful commercialisation of novel technologies. Though this project does not set out to develop stably-transfected lines of bees, it lays the foundation for such future work both in terms of resources such as Wolbachia-infected bee cells, and data, such as the relative effects of different strains of Wolbachia. There are clear commercial uses if strains of bees showing beneficial traits were to be produced. Correspondingly, these data and resources will be of significant value to commercial enterprises such as queen rearing in the honey bee industry. We will therefore consider the commercial sensitivity of these data prior to publication; the IP generated in this project will be identified and appropriately exploited. Since current GM legislation is uncertain and expensive in terms of time-to-market, at least in the UK and Europe, applications leading to non-GMO products are likely to be of more interest, and we have taken this into account in developing the project. Beneficial insect companies have showed interest in insect genetic technology, and the idea of this research; we aim to gain more detailed understanding of relevant commercial needs, and possibly develop formal collaborations, over the course of the project.
This research will be of interest to beekeepers and bee farmers across the UK. We will communicate directly with interested stakeholders, e.g. through attendance and presentation at apiculture conferences, such as the National Honey show and the BBKA convention. We have established contacts allowing us to engage with the beekeeping community on this research. The Pirbright Institute and the scientists involved have a strong track record in communicating to both scientific and lay audiences, and of taking novel applied genetic approaches from initial conception through to successful commercialisation and dissemination.
