Web-portal for transboundry pathogens

Lead Research Organisation: THE PIRBRIGHT INSTITUTE
Department Name: UNLISTED

Abstract

Abstracts are not currently available in GtR for all funded research. This is normally because the abstract was not required at the time of proposal submission, but may be because it included sensitive information such as personal details.

Technical Summary

We will create of a web portal for all the major animal diseases studied at Pirbright, starting with foot-and-mouth disease and lumpy skin disease. The core purpose will be to allow researchers and policy makers from ODA countries easy and immediate access to the wealth of information gathered by all the reference labs at Pirbright. By exploring an interactive geo-localized map of his/her country any researcher would be able to locate samples for all relevant animal diseases and query their sequence, explore past and present outbreaks, inquire about available vaccines and gather information that can inform decision making. We will also take the opportunity to sequence additional viruses in the reference collection.

Planned Impact

unavailable
 
Description The project has two basic components.

The first one is the sequencing of a number of viral samples originating from animals infected by foot-and-mouth disease virus (FMDV), African Swine Fever virus (ASFV), or Capripoxviruses (such as lumpy skin disease virus, LSDV). Such pathogens have been selected for this project as they all cause livestock diseases that have disastrous consequences on the economy of low- and middle-income countries (LMICs).

The second component consists of a bioinformatics portal from which any scientist operating in LMICs will be able to download sequences according to a number of filtering criteria (country of origin, date range, etc.). Together with the sequences one will be able to download a functional annotation for the genome. In addition to the sequences links to suitable literature and analysis tools might be presented depending on the virus and the sequence.

As for the first component, sequencing protocols for FMDV were already available, specifically optimised for sequencing in containment at Pirbright. We have then proceeded to sequence more than 300 full FMDV genomes to date. The samples were provided by either the FMDV World Reference Lab or other projects (in particular, we selected a number of samples collected from African buffalo, as buffalo is thought to be the natural reservoir through which FMDV persists in the environment).
As for Capripoxviruses we selected a protocol used in Hughes et al, J Virol Meth 243 (2017) 68-73, and proceeded to sequence ~10 viral strains to prototype the technique. The sequencing has been successful, and we are now in the process of identifying a larger number of samples suitable for the project.
ASFV is the most challenging virus, as short-read Illumina-based protocols do not allow a full reconstruction of the viral sequence. It was then decided to sequence the virus externally at Earlham using long-read technologies (Pacific Bioscience) that are not available at Pirbright. While a purification protocol to prepare high quality high molecular weight viral DNA from different sample types has been successfully established, it turns out that the inactivation step needed to extract the material from the SAPO4 restricted area envelope at Pirbright prior to shipment to Earlham significantly degrades the quality of nucleic acid in the sample. We have recently largely overcome this problem by combining new sequencing technologies and library preparations and using our sequencing capacity at Pirbright. We hope to generate the data during the coming year.

New FMDV Sequence Database
We have begun to develop an open-source database (MySQL) to hold foot-and-mouth disease (FMD) virus sequences and associated metadata. The database will consist of both data generated by, research groups, the World Reference Laboratory for FMD (WRLFMD) at the Pirbright Institute and other FMD Network Laboratories, as well as publicly available sequence data retrieved from GenBank. This data will be curated and annotated by FMD experts to a higher level than currently hosted on GenBank. The database is expected to consist of a number of entities with associated information including, but not limited to, 'Sample Batch', 'Virus Sample', 'Virus Genome', 'Virus Sequence', 'Country', 'Host Species', 'Laboratory' and 'Taxonomic Classification'. The database will also require different security levels for users to access data that is considered 'Public', 'Private' or 'Confidential'. The database is now complete and is being populated and expertly curated. This will form the single curated source of data that will be accessed by a web browser that will be publicly facing.
Exploitation Route The viral sequences we are generating will benefit the larger scientific communities that study FMDV, Capripoxviruses and ASFV.

The long-read sequencing protocol we are developing might be really important for all scientists studying ASFV, whose genome is highly redundant and hence difficult to sequence. One outcome would be the possibility of determining better molecular markers for the virus, as the effectiveness of the ones currently used to characterise it has been recently questioned.

A single point of access for reference laboratory data would be of major benefit globally, with the associated reporting and analysis tools
Sectors Agriculture

Food and Drink

Government

Democracy and Justice

Manufacturing

including Industrial Biotechology

Security and Diplomacy
 
Description Horizon2020
Amount € 1,900,000 (EUR)
Funding ID DEFEND 
Organisation European Union 
Sector Public
Country European Union (EU)
Start 04/2018 
End 04/2022
 
Title Method for sequencing of ASFV (African Swine Fever Virus) based on long reads 
Description The ASFV genome is repetitive, due to the presence of families of genes that are present in multiple copies (>10). As a result, sequencing based on short reads only would not be able to reconstruct the genome in its entirety. In addition, good quality full genome sequencing of ASFV is complicated by persistence contamination with fragments of host DNA. We are developing a novel protocol that will allow the sequencing of the virus using long-read technologies (PacBio or Oxford Nanopore). Probe-capture libraries have been designed to mitigate the problem of host contamination by selectively purifying viral DNA from a wide range of different ASFV genotypes. Protocols are being developed to prepare high quality high molecular weight viral DNA from different sample types suitable for generating sequencing libraries for long-read sequencing. 
Type Of Material Technology assay or reagent 
Year Produced 2019 
Provided To Others? No  
Impact More than 100 samples from 18 different LMIC countries have been identified and collated for sequencing with the new protocol. They will be added to the web-portal for transboundary pathogens which is currently being developed thanks to BBSRC funding. 
 
Title FMDV LIMS and webtools 
Description The Integrative Biology and Bioinformatics (IBB) group has been supplying state-of-the-art bioinformatics tools to the World Reference Labs for the Foot-and-Mouth Disease Virus. All tools revolve around a database of genomic sequences and metadata for FMDV samples. Some tools allow members of the lab to enter, process and annotate new data into the database, generating several kinds of reports. Other web-based tools (the "FMDV Toolbox") allow external users to query the database in several ways. Bioinformatics workflows are hosted on the IBB cluster, and have been implemented as REST web services so as to be decoupled from the web interface. The web interface is currently being redeveloped with the help of a specialised company. 
Type Of Material Database/Collection of data 
Year Produced 2016 
Provided To Others? Yes  
Impact The reports generated by our system are used to inform FMDV policy-makers both within the UK and in the rest of the world. External users can compare their sequences to those contained in the database, and get useful biological insights. 
URL https://mallorn.pirbright.ac.uk/FMDVToolbox
 
Title GCRF portal on transboudary diseases 
Description The Integrative Biology and Bioinformatics group is developing a GCRF-funded Portal on Transboundary Diseases. It will be an easy way for all scientists, and those from LMI countries in particular, to access data about several viral diseases generated at Pirbright. The portal will be country-based, presenting the user with a map and dispatching them to the information relevant to the selected region. From that information the user will be able to reach specialised websites for diseases present in the country, and data generated at Pirbright (such as sequences for FMDV, several Capripox viruses, and ASFV). Sequences relevant to the portal are currently being generated. Bioinformatics workflows are hosted on the IBB cluster, and have been implemented as REST web services so as to be decoupled from the web interface. The web interface is currently being developed with the help of a specialised company. 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? No  
Impact The portal will benefit scientists from LMI countries, and all scientist, allowing them to access/download genetic and other data relevant to animal diseases present in a country of choice and studied and Pirbright. Some of the data (for instance the FMDV data generated by the World Reference Lab) is already use to inform policy at international level. Being able to access information in a systematic way will increase the potential for it to be put to good use. 
 
Title South-African buffalo FMDV sequences 
Description We have sequenced FMDV buffalo samples originated from a EEID project entitled: Persistence of a Highly Contagious Pathogen: Ecological and Evolutionary Mechanisms in Foot-And-Mouth Disease Virus. This project aims to understand why a highly contagious pathogen such as FMDV, which induces a rapid host immunity and depletes the supply of susceptible host, is able to persist in isolated buffalo populations and thus avoid auto-extinction. The centre piece of the project is a cohort study which involves an established FMDV-positive breeding herd of ca. 70 buffalo in a 900-hectare enclosure surrounded by double game fencing housing buffalo in isolation from other herds in the Kruger National Park (South Africa). The entire herd is being monitored for three years (animals are sampled every 2 months (serum, tonsil swabs, probang) to trace FMDV transmission events, allowing us to define FMD infection dynamics across the susceptible calf cohorts and amongst adults. So far, FMDV genomes from 101 samples have been deep sequenced by illumina. Samples from the last year captures are currently being analysed and virus is currently being isolated. The second experiment is an experimental study, which involves a group of naïve buffalo experimentally infected with either a SAT-1, SAT-2 or SAT-3 virus. The infected buffalo were then allowed to mingle with susceptible animals and transmission of FMDV to naïve animals was monitored during the acute infection but also from persistently infected animals. So far, FMDV genomes from 48 samples obtained at days 2, 30 and 160 of the experiment have been deep sequenced. 
Type Of Material Database/Collection of data 
Year Produced 2019 
Provided To Others? No  
Impact The samples sequenced come from South Africa, which is an LMIC, While FMDV infection in cattle is very well studied, the mechanisms of persistence in buffalo, which is thought to be the natural reservoir of the infection, are not well understood. Such understanding would be essential to inform better policies to understand and manage FMDV, which is an economically important scourge of cloven hooved animals in LMICs. All the sequences will be made available on the Transboundary Portal which is being developed at Pirbright. 
 
Title Viral assembly pipeline 
Description The Integrative Biology and Bioinformatics group has developed a data analysis pipeline to obtain sensitive and robust viral assemblies out of high-throughput sequencing data. In spite of the relatively short genomic length of most viruses, assembling of viral sequences can be challenging due to several reasons, such as: low amount of material in the sample, which might require amplification, introducing biases, and/or generates low-quality sequences; uneven coverage due to low-quality material or the genomic material of the virus being RNA; the viral nucleic acid being almost lost in the background of nucleic acid of the host. Our pipeline overcomes most of those problems, and is even able to detect different viral strains being present together in the same sample. It can also be used on metagenomic environmental samples. 
Type Of Material Data analysis technique 
Year Produced 2017 
Provided To Others? No  
Impact The pipeline is being deployed as the tool of choice for the nascent sequencing facility at Pirbright. It has been used by several groups at Pirbright, and the FMDV World Reference Lab, in order to assemble very different kinds of viruses. 
 
Title Viral variant calling pipeline 
Description The Integrative Biology and Bioinformatics group has developed a data analysis pipeline to obtain sensitive and robust variant calling for viral sequences out of high-throughput sequencing data. In spite of the relatively short genomic length of most viruses, calling variant for viral sequences can be challenging due to several reasons, such as: low amount of material in the sample, which might require amplification, introducing biases, and/or generates low-quality sequences; very high genome coverage, which slows down most existing variant callers; the presence of a potentially very high number of different haplotypes for RNA viruses, which is an unusual scenario in variant calling. Our fast and sensitive Bayesian pipeline overcomes most of those problems. 
Type Of Material Data analysis technique 
Year Produced 2017 
Provided To Others? No  
Impact The pipeline is being deployed as the tool of choice for the nascent sequencing facility at Pirbright. It has been used by several groups at Pirbright, and the FMDV World Reference Lab, in order to assemble very different kinds of viruses. 
 
Title WRL FMDV sequences for transboundary portal 
Description We have generated a collection of sequences of FMDV (foot-and-mouth disease virus) from samples collected by the FMDV World Reference Laboratory at Pirbright. Those full-genome sequences sample a number of recent exotic strains across different regions/countries of the world, focusing mainly on LMICs in North Africa, Middle East and South East Asia. The main represented FMDV types are O and A. Part of the sequences have already been made available through GenBank -- all will be soon downloadable from the Transboundary Portal which is being developed at Pirbright. 
Type Of Material Database/Collection of data 
Year Produced 2019 
Provided To Others? Yes  
Impact Knowledge of the sequenced strains is essential to inform policy for FMDV control in the LMIC countries where the samples were collected. That chimes very well with the GCRF focus of the project. At a more global scale, knowledge of the features and sequence evolution of the virus in time and across different countries is essential in order to accurately model its behaviour. In general FMDV is a scourge of cloven-hoofed animals, and controlling it better is essential in order to alleviate the huge economic toll claimed by the virus in LMICs. 
 
Title Zimbabwe FMDV sequences 
Description Foot-and-mouth disease virus (FMDV) causes an acute vesicular disease in domestic cloven-hooved animals. However, in the African buffalo (Syncerus caffer) clinical disease is rarely observed and following infection virus is persistently carried in the oesophageal-pharyngeal area of the upper respiratory tract. During the 1990s oesophageal-pharyngeal scrapings were collected from free-living African buffalo in multiple herds in six different geographic areas of Zimbabwe. We sequenced over 140 FMD viruses each belonging to one of the Southern African Territories (SAT) serotypes (SAT 1, SAT 2 and SAT 3) from primary bovine thyroid cells. 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? No  
Impact This dataset has been generated from samples collected in Zimbabwe, which is a low-income country. Those viral sequences will be essential to help elucidating the nature of persistent FMDV infection in African buffalo, which is supposed to be the main virus reservoir in vivo. As the samples track viral infection and evolution over several years and across a number of different herds in different national parks and conservatories, the dataset will also inform better animal management and conservation. A better understanding of FMDV persistence would also be essential to mitigate the economic burden generated by the disease, which is a scourge of cloven-hoofed animals in LMICs. All the sequences will be made available through the Transboundary Pathogen portal that is being developed at Pirbright. 
 
Description EUFMD 2016 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Policymakers/politicians
Results and Impact During the EuFMDV conference taking place from 26 to 28 October 2016, Paolo Ribeca gave a talk on "VIBAsys and FMDV-Tools: Practical resources for FMDV sequence analysis". The conference, organised by the EU, gathers most of the important actors in FMD control. The talk explained how the new resources being developed by the Integrative Biology and Bioinformatics on behalf of the FMDV World Reference Lab can be useful to the community
Year(s) Of Engagement Activity 2016
 
Description Poster by Nick on the sequencing of Zimbabwe FMDV 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Policymakers/politicians
Results and Impact Nick Knowles presented a poster at the 2018 Europic (the meeting of the European Study Group on the Molecular Biology of Picornaviruses) on "Complete Genome Sequence Analysis of Over 140 Foot-and-mouth Disease Viruses Isolated From Free-living African Buffalo (Syncerus Caffer) in Zimbabwe" on behalf of Jemma Wadsworth, Bruce Bolt, Luca Ferretti, Euan C. Anderson, Ashley Gray, Paolo Ribeca, and himself. The sequencing was funded by the Transboundary Pathogens portal project awarded to Pirbright.
Year(s) Of Engagement Activity 2018
 
Description Talk by Nick on the sequencing of Zimbabwe FMDV 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Policymakers/politicians
Results and Impact Nick Knowles gave a talk at the 2018 Open Session of EuFMD (European commission for the control of foot-and-mouth disease ) on "Complete Genome Sequence Analysis of Over 140 Foot-and-mouth Disease Viruses Isolated From Free-living African Buffalo (Syncerus Caffer) in Zimbabwe" on behalf of Jemma Wadsworth, Bruce Bolt, Luca Ferretti, Euan C. Anderson, Ashley Gray, Paolo Ribeca, and himself. The sequencing was funded by the Transboundary Pathogens portal project awarded to Pirbright.
Year(s) Of Engagement Activity 2018
 
Description UK-India bioinformatics workshop 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact On February 9 2018 Pirbright organised a "UK-India bioinformatics workshop" in order to foster ties between Indian and UK bioinformaticians working in the field of livestock and crop research. Paolo Ribeca gave a talk on "Bioinformatics and sequencing at Pirbright".
Year(s) Of Engagement Activity 2018