Identifying the genetic mechanisms facilitating host range and virulence of a viral pathogen that threatens European amphibian biodiversity
Lead Research Organisation:
Queen Mary University of London
Department Name: Sch of Biological and Chemical Sciences
Abstract
Amphibians are experiencing catastrophic declines, caused by the emergence of infectious diseases. In Europe, the causative agent is predominantly viral (ranavirus), and our work has shown that emergence of ranavirus in amphibian communities can result in at least three different patterns: the mortality and population decline can be host-specific, it can affect the whole host community or infections can be asymptomatic, co-occurring in areas where lethal infections are common. From analysis of viral isolates we have determined that this variation correlates with viral phylogeney: in Spain, a genetic lineage responsible for amphibian community mortality and decline co-circulates with a genetically distinct asymptomatic lineage; in the UK, a third lineage is associated with death and decline of a single host species (common frogs - infection of other host species is rare and does not lead to significant mortality or decline in spill-over hosts).
Here we propose to take advantage of this novel system to ascertain what genetic factors are responsible for these differences in host range and virulence. Our approach is three-pronged: first, we will sequence 130 ranavirus genomes to catalogue genetic variation amongst the European lineages and identify candidate loci. We will also take a more bottom-up phylogenetic approach, identifying any genetic variants that map with differences in host species in a global panel of ranavirus genomes. Second, we carry out controlled infection experiments to verify the different host range and virulence of different viral isolates in three animal models (two anurans and one caudate chosen to encompass the host range we have observed in Spain). Third, we will generate viral knockouts, targeting candidate regions we have identified in the first two objectives, and test hypotheses regarding gene function using recombinants and wild type virus in our animal models.
Our results will be a crucial contribution to our understanding of why ranaviruses are such lethal pathogens in some circumstances. Ranavirus emergence is on the increase across Europe, and is emerging as a novel threat to amphibian and reptile biodiversity in Latin America and Madagascar, two amphibian and reptile biodiversity hotspots. Our research will more widely inform the epidemiological community as to what genetic factors may be important for viruses to exploit novel hosts. Given that most emerging infections are viral and emergence arises through host jumps, we expect our findings to be of broad interest to this community.
Here we propose to take advantage of this novel system to ascertain what genetic factors are responsible for these differences in host range and virulence. Our approach is three-pronged: first, we will sequence 130 ranavirus genomes to catalogue genetic variation amongst the European lineages and identify candidate loci. We will also take a more bottom-up phylogenetic approach, identifying any genetic variants that map with differences in host species in a global panel of ranavirus genomes. Second, we carry out controlled infection experiments to verify the different host range and virulence of different viral isolates in three animal models (two anurans and one caudate chosen to encompass the host range we have observed in Spain). Third, we will generate viral knockouts, targeting candidate regions we have identified in the first two objectives, and test hypotheses regarding gene function using recombinants and wild type virus in our animal models.
Our results will be a crucial contribution to our understanding of why ranaviruses are such lethal pathogens in some circumstances. Ranavirus emergence is on the increase across Europe, and is emerging as a novel threat to amphibian and reptile biodiversity in Latin America and Madagascar, two amphibian and reptile biodiversity hotspots. Our research will more widely inform the epidemiological community as to what genetic factors may be important for viruses to exploit novel hosts. Given that most emerging infections are viral and emergence arises through host jumps, we expect our findings to be of broad interest to this community.
Planned Impact
Our results will provide the pertinent genomic information, host range and virulence data that will enable us to develop diagnostic marker systems for the identification of highly virulent, less virulent and asymptomatic ranavirus lineages. These resources will be of global utility, as ranavirus is an OIE notifiable pathogen yet current tools for detection fail to differentiate between lineages. Ranaviruses are lethal pathogens of fish, amphibians and reptiles - including many economically important (farmed and traded species) as well as taxa significant to biodiversity. Thus, stakeholders interested in the tools we are developing include national and regional veterinary authorities responsible for monitoring notifiable diseases in the trade, wildlife managers assessing risk that ranaviruses pose to wildlife under their care, and the international research community that make up the Global Ranavirus Consortium. This last includes labs identified as equipped for diagnostic identification of ranaviruses. Our experimental investigation of tissue-level infection dynamics as related to pathogen genotype will allow all those who employ molecular diagnostic tools to do so more effectively while incurring less cost per animal.
At the UK level, Garner sits on the UK Wildlife Health Committee with representatives from NGOs (Natural England) and the Government (DEFRA). The outputs from our research are reported to this committee via meetings on a bi-annual basis and provide a scientific basis for decisions on amphibian conservation in the UK. Our Project Partner Bosch informs a similar committee in Spain, where similar efforts are underway to manage amphibian populations with respect to infectious diseases. Both countries are signatories to the OIE and as such decisions regarding amphibian trade and amphibian wildlife management must take into account the relative risk of different genotypes. We can deliver this information directly to the national bodies responsible for making decisions regarding pathogens in trade and movement of potentially infected animals: for example, in the UK the ZSL is tasked with providing quarterly wildlife disease updates to the GB Wildlife Disease Surveillance scheme, which is the UK's reporting scheme that informs the OIE. The ZSL also sits on the Ad Hoc Group on Amphibian Diseases, providing information to the OIE which underpins the classification of diseases as notifiable and guides recommendations as to the risk posed by pathogens according to genotype Our results will provide both the tools and the virulence data necessary for the OIE to start issuing regarding the relative risk of ALRV lineages and how this should feed into appropriate management strategies for traded animals.
Our spatial and genomic data will guide the development of efforts to map the distribution of ranaviruses based on genotypes. We are currently involved in the modification of the GRRS to accommodate virus genotype, alongside projects to genotype ATV ALRVs (Jancovich) and North American FV3s (Waltzek lab, University of Florida).
We also expect our outputs to find a wide academic audience. Our combination of pathogen genomics, multi-host experimental studies of host/tissue specificity and virulence, and our explicit tests of function using knockouts is novel and will be crucial for any current and future academic effort aiming at understanding ranavirus epidemiology, host/parasite interactions, virology, parasite evolution and host/parasite coevolution.
At the UK level, Garner sits on the UK Wildlife Health Committee with representatives from NGOs (Natural England) and the Government (DEFRA). The outputs from our research are reported to this committee via meetings on a bi-annual basis and provide a scientific basis for decisions on amphibian conservation in the UK. Our Project Partner Bosch informs a similar committee in Spain, where similar efforts are underway to manage amphibian populations with respect to infectious diseases. Both countries are signatories to the OIE and as such decisions regarding amphibian trade and amphibian wildlife management must take into account the relative risk of different genotypes. We can deliver this information directly to the national bodies responsible for making decisions regarding pathogens in trade and movement of potentially infected animals: for example, in the UK the ZSL is tasked with providing quarterly wildlife disease updates to the GB Wildlife Disease Surveillance scheme, which is the UK's reporting scheme that informs the OIE. The ZSL also sits on the Ad Hoc Group on Amphibian Diseases, providing information to the OIE which underpins the classification of diseases as notifiable and guides recommendations as to the risk posed by pathogens according to genotype Our results will provide both the tools and the virulence data necessary for the OIE to start issuing regarding the relative risk of ALRV lineages and how this should feed into appropriate management strategies for traded animals.
Our spatial and genomic data will guide the development of efforts to map the distribution of ranaviruses based on genotypes. We are currently involved in the modification of the GRRS to accommodate virus genotype, alongside projects to genotype ATV ALRVs (Jancovich) and North American FV3s (Waltzek lab, University of Florida).
We also expect our outputs to find a wide academic audience. Our combination of pathogen genomics, multi-host experimental studies of host/tissue specificity and virulence, and our explicit tests of function using knockouts is novel and will be crucial for any current and future academic effort aiming at understanding ranavirus epidemiology, host/parasite interactions, virology, parasite evolution and host/parasite coevolution.
People |
ORCID iD |
Richard Nichols (Principal Investigator) |
Publications
Black Y
(2017)
DETECTION AND REPORTING OF RANAVIRUS IN AMPHIBIANS: EVALUATION OF THE ROLES OF THE WORLD ORGANISATION FOR ANIMAL HEALTH AND THE PUBLISHED LITERATURE.
in Journal of wildlife diseases
Campbell L
(2019)
Outbreaks of an Emerging Viral Disease Covary With Differences in the Composition of the Skin Microbiome of a Wild United Kingdom Amphibian
in Frontiers in Microbiology
Duffus ALJ
(2019)
Modelling Ranavirus Transmission in Populations of Common Frogs (Rana temporaria) in the United Kingdom.
in Viruses
Harrison XA
(2019)
Diversity-Stability Dynamics of the Amphibian Skin Microbiome and Susceptibility to a Lethal Viral Pathogen.
in Frontiers in microbiology
Leung WTM
(2017)
A quantitative-PCR based method to estimate ranavirus viral load following normalisation by reference to an ultraconserved vertebrate target.
in Journal of virological methods
Price S
(2016)
Reconstructing the emergence of a lethal infectious disease of wildlife supports a key role for spread through translocations by humans
in Proceedings of the Royal Society B: Biological Sciences
Price SJ
(2017)
Screening of a long-term sample set reveals two Ranavirus lineages in British herpetofauna.
in PloS one
Price SJ
(2015)
Comparative Genomics of Amphibian-like Ranaviruses, Nucleocytoplasmic Large DNA Viruses of Poikilotherms.
in Evolutionary bioinformatics online
Price SJ
(2019)
Effects of historic and projected climate change on the range and impacts of an emerging wildlife disease.
in Global change biology
Price SJ
(2017)
From fish to frogs and beyond: Impact and host range of emergent ranaviruses.
in Virology
Puschendorf R
(2019)
Cryptic diversity and ranavirus infection of a critically endangered Neotropical frog before and after population collapse
in Animal Conservation
Robert J
(2018)
Water Contaminants Associated With Unconventional Oil and Gas Extraction Cause Immunotoxicity to Amphibian Tadpoles.
in Toxicological sciences : an official journal of the Society of Toxicology
Rosa GM
(2017)
Impact of asynchronous emergence of two lethal pathogens on amphibian assemblages.
in Scientific reports
Saucedo B
(2019)
Common midwife toad ranaviruses replicate first in the oral cavity of smooth newts (Lissotriton vulgaris) and show distinct strain-associated pathogenicity.
in Scientific reports
Soto-Azat C
(2016)
Xenopus laevis and Emerging Amphibian Pathogens in Chile.
in EcoHealth
Von Essen M
(2020)
High pathogen prevalence in an amphibian and reptile assemblage at a site with risk factors for dispersal in Galicia, Spain.
in PloS one
Walker JG
(2017)
Host allometry influences the evolution of parasite host-generalism: theory and meta-analysis.
in Philosophical transactions of the Royal Society of London. Series B, Biological sciences
Wynne FJ
(2020)
Choice of molecular assay determines ranavirus detection probability and inferences about prevalence and occurrence.
in Diseases of aquatic organisms
Description | Climate change is often predicted to result in more severe disease epidemics. However, the interactions between disease and the environment can be highly complex and few studies have produced unambiguous results. We addressed the challenge of demonstrating the role of temperature on epidemics caused by ranaviruses - wildlife viruses that are expanding worldwide, and which have spread through southern regions of the UK in the past two decades causing catastrophic declines in populations of frogs. Our research was distinctive in combining three separate lines of evidence - analysis of the temperature effects on viral propagation in cell culture, laboratory studies of animals in controlled conditions showing increased infection and mortality at higher temperature, and an analysis of disease reports wild UK populations showing greater severity during a period of historic warming in the 1990s. All three approaches suggested that increased temperature can drive increased geographic range and severity of the epidemics. The most worrying outcome is that our projections, based on future climate predictions, indicate that the virus may begin to affect the populations in the more vulnerable tadpole life stage. More encouragingly, we were able to suggest management interventions such as planting around ponds, that could counter these trends (Listed in the Global Change Biology paper 2019). |
Exploitation Route | Discovered or developed: A reliable quantitative PCR diagnostic for ranavirus. The technique was presented at the Global Ranavirus Consortium meeting in summer of 2017, and published in the 2018 paper. Further work on temperature using long-term monitoring data and combining in vitro and in vivo assays has revealed a key role for environmental temperature in determining prevalence of infection and severity of disease outcomes. Last, through a collaboration within the IoZ, we have begun to describe an important role for amphibian skin microbial communities in determining prevalence of infection and severity of disease outcomes. Together, these two findings inform our future research plans that we hope will help us develop a disease mitigation strategy for emerging ranaviruses. Intriguingly we found that the patterns of mortality caused by CMTV-like ranaviruses in nature were not replicated in controlled experiments, and our work with Dr Saucedo during his placement has lead us to establish methods for different routes of transmission, and have been granted permission by the HO to infect and freeze-kill tadpoles so we can 'feed' ranaviruses to foraging tadpoles. We should be getting results for this approach next month. Our work on examining virus growth rates in cell culture across temperatures and comparing this to virulence in common frogs is now being used in an NC3Rs-funded studentship and one of our London DTP students to, respectively, examine if cell culture can replace animals for studies of virulence and if ranaviruses evolve in response to climate change. The NC3Rs student is also building on Dr Saucedo's histology work to determine if intervention before the onset of lethal disease in experiments can still yield relevant information regarding virulence through the use of histopathology and qPCR, using the diagnostic developed in 2017. Findings taken forward or used: Uptake of the qPCR diagnostic in the research community has begun. Our work on temperature and microbiomes will undoubtedly inspire others to investigate the role these factors play in ranavirosis in their study systems. |
Sectors | Education Environment Leisure Activities including Sports Recreation and Tourism Transport |
Description | Through presentations and talks, we have raised awareness of the threat posed to UK amphibians, reptiles and fish by the CMTV-like ranaviruses emerging on the continent. |
First Year Of Impact | 2019 |
Sector | Environment |
Impact Types | Cultural |
Description | Reporting of ranavirus infection |
Geographic Reach | Multiple continents/international |
Policy Influence Type | Influenced training of practitioners or researchers |
Description | Chinese Scholarship Council PhD award for Ms Niu Yuhui |
Amount | £64,800 (GBP) |
Funding ID | 201908320461 |
Organisation | Chinese Scholarship Council |
Sector | Charity/Non Profit |
Country | China |
Start | 08/2019 |
End | 08/2023 |
Description | NERC London DTP studentship |
Amount | £86,000 (GBP) |
Organisation | Natural Environment Research Council |
Sector | Public |
Country | United Kingdom |
Start | 09/2017 |
End | 09/2021 |
Description | State Programme for R&D: Challenges facing Society |
Amount | € 122,000 (EUR) |
Funding ID | CGL2015-70070-R |
Organisation | Spanish Ministry of Economy and Competitiveness |
Sector | Public |
Country | Spain |
Start | 08/2016 |
End | 09/2019 |
Title | Viral load estimation by quantitative PCR |
Description | Design of sensitive and specific qPCR methods targeting virus and host loci to be used in tandem to generate viral load estimates |
Type Of Material | Technology assay or reagent |
Year Produced | 2016 |
Provided To Others? | Yes |
Impact | screening of various sample types for presence and intensity of ranavirus infection in six labs on three continents (as of February 2017) yielding more accurate information on host and geographic distribution of a World Organisation for Animal Health listed pathogen. |
Title | Citizen science reports of amphibian mortality in the UK between 1991-2010 |
Description | Records relating to a citizen science report of amphibian mortality in the UK between 1991-2010. The fields cover georeference data and data on the mortality event (numbers of affected animals and signs of disease observed). |
Type Of Material | Database/Collection of data |
Year Produced | 2016 |
Provided To Others? | Yes |
Impact | Enabled modelling of spread of infectious disease in British frog populations and identification of likely drivers of emergence, which lead to publication of findings: Price SJ, Garner TWJ, Cunningham AA, Langton TES & RA Nichols (2016) Reconstructing the emergence of a lethal infectious disease of wildlife supports a key role for spread through translocations by humans. Proc R Soc B 283: 20160952. http://dx.doi.org/10.1098/rspb.2016.0952 |
URL | http://rspb.royalsocietypublishing.org/content/283/1839/20160952 |
Description | Assessing spatial and host distribution of BNV in Galicia |
Organisation | Spanish Herpetological Association |
Country | Spain |
Sector | Learned Society |
PI Contribution | Developed project proposal and permit application for field project. Recruited, trained and supervised student. |
Collaborator Contribution | Helped write and submit permit application as well as providing knowledge of field sites which facilitated project development. Supervision of student in field. |
Impact | Publication in Current Biology, Price et al., 2014. |
Start Year | 2010 |
Description | Gene knockout methodologies |
Organisation | University of Rochester |
Department | Department of Neuroscience |
Country | United States |
Sector | Academic/University |
PI Contribution | Dr. Stephen Price visited Jacques Robert's lab at the University of Rochester for there months to train and access support in virus gene knockout methodologies. Whilst there he collaborated in infection experiments and helped analyse data generated by the Robert lab. |
Collaborator Contribution | Provided training and support in methods essential for meeting one of the project objectives. Provided animals, training and a virus isolate for infection studies. |
Impact | Creation of four ranavirus (knock-out) recombinants Completed two infection trials Collaboration on a study on effects of fracking on virus infection |
Start Year | 2015 |
Description | Amphibian Week for Edinburgh MVetSci |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Public/other audiences |
Results and Impact | One week online workshop including video lectures, discussion groups, group activity. |
Year(s) Of Engagement Activity | 2015,2016,2017,2018 |
URL | http://www.ed.ac.uk/schools-departments/vet/studying/postgraduate/taught-programmes/conservation-med... |
Description | Departmental seminar at University of Liverpool |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Professional Practitioners |
Results and Impact | Invited departmental seminar at University of Liverpool, topic was 'amphibian-associated ranaviruses: the first (and worst?) infectious disease conservation issue for amphibians' |
Year(s) Of Engagement Activity | 2017 |
URL | https://www.liverpool.ac.uk/integrative-biology/events/ |
Description | Graduate student conservation conference (Edinburgh |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Postgraduate students |
Results and Impact | Presentation on the relative importance of ecology vs evolution for mitigating amphibian infectious diseases |
Year(s) Of Engagement Activity | 2015 |
Description | International Ranavirus Symposium, Budapest - conference talk |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Other audiences |
Results and Impact | Regular talk at international conference for researchers and third sector organisations |
Year(s) Of Engagement Activity | 2017 |
Description | International Ranavirus Symposium, Budapest; Keynote talk |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Other audiences |
Results and Impact | Keynote talk at international conference for researchers and third sector organisations |
Year(s) Of Engagement Activity | 2017 |
Description | Invited departmental seminar (Uppsala) |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Postgraduate students |
Results and Impact | Invited departmental seminar for Ecology and Evolution department at Uppsala University |
Year(s) Of Engagement Activity | 2015 |
Description | Invited public talk (Richmond) |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Public/other audiences |
Results and Impact | I was invited to give a talk about wildlife disease and amphibian conservation to the local Richmond Park nature association |
Year(s) Of Engagement Activity | 2015 |
Description | Invited talk (Silwood Park Seminar Series) |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Postgraduate students |
Results and Impact | Seminar to broad academic audience which sparked new contacts and ongoing discussion afterwards |
Year(s) Of Engagement Activity | 2017 |
Description | Keynote address to the Global Ranavirus Consortium |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Co-presented keynote address on the host and geographic distribution of ranaviruses in Europe and their potential and actual threats to biodiversity |
Year(s) Of Engagement Activity | 2017 |
URL | https://www.ranavirus.org/2017-4th-international-symposium-on-ranavirus/ |
Description | Keynote address to the Herpetological Association of Africa outlining potential threat of ranaviruses to African herpetofauna |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Delivered a keynote address to the annual meeting of the Herpetological Association of Africa outlining the potential threat ranaviruses pose to African herpetofauna |
Year(s) Of Engagement Activity | 2017 |
URL | http://www.africanherpetology.org/ |
Description | Lecture on GRC Ranavirus course |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | Live video lecture to raise awareness about ranaviruses among students and professionals. |
Year(s) Of Engagement Activity | 2016 |
URL | http://www.ranavirus.org/online-course-on-ranavirus-biology/ |
Description | NC3Rs Workshop on Amphibian Welfare |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Third sector organisations |
Results and Impact | Two day workshop to engage people from different sectors with shared interest in driving change |
Year(s) Of Engagement Activity | 2017 |
Description | Presentation at ZSL scientific event |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Public/other audiences |
Results and Impact | Presentation regarding the potential impact of invasive species on emergence of threatening amphibian infectious diseases, delivered as part of the ZSL Scientific Event launching the 2016 Living Planet Report |
Year(s) Of Engagement Activity | 2016 |
URL | https://www.zsl.org/science/whats-on/the-living-planet-report-2016-threats-pressures-and-addressing-... |
Description | Presentation to stakeholders |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Professional Practitioners |
Results and Impact | Presentation to a combined meeting of British Amphibian and Reptile groups (ARGs UK) to inform about the potential risk posed to UK herpetofauna by variants of ranaviruses emerging on the European continent |
Year(s) Of Engagement Activity | 2016 |
Description | School visit |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Schools |
Results and Impact | Presentations and conversations with three and four year olds and pre-school staff about wonderful amphibians and reptiles |
Year(s) Of Engagement Activity | 2017 |
Description | Training on IoZ contribution to training on the London DTP and the Oxford DTP |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Postgraduate students |
Results and Impact | I contributed to our annual event hosting PhD candidates on two NERC-funded DTPs. The core content of my presentation was constructed around current research on my NERC-funded projects, and providing PhD candidates the opportunity to become aware of the opportunities and support that could be provided to them through existing NERC projects. Some of the participants contacted me for further information about methods, contacts and the possibility of involvement in the research programmes |
Year(s) Of Engagement Activity | 2017 |