The challenge addressed by this project is to unravel the causes of stunting disease in UK broiler chickens. Stunting syndrome has been a production issue in commercial broiler flocks since they became industrialised after the second world war, but is also a problem for small producers in low and middle income countries, such as Nigeria, where women often raise small numbers of chickens to supplement the family income. The stunting problem has many names, including infectious stunting, runting stunting syndrome and malabsorption disease. It pertains to a failure to grow properly despite sufficient feed. Birds typically present with stunting problems around the second to third week post hatching, but clinical signs may be observable earlier. As well as the reduction in weight gain, which can be substantial with severely affected chickens only a fraction of their potential weight for age, there may be other signs such as altered appetite, watery droppings, abnormal feathering and enteritis. We and other groups have evidence that the stunting is caused by infectious agents: chiefly, viral aetiology is suspected and experimental infections at AFBI using specific strains of the endemic, enteric viruses, astroviruses and reoviruses separately have resulted in some of the signs observed during stunting, e.g. weight loss or lesions in the gut mucosa. Other groups have also demonstrated some of the signs of stunting from inoculating birds with single viral strains of these and other viral candidates, e.g. birnavirus; however, none have fully recreated the stunting syndrome, and co-infections with more than one viral agent are considered the more probable aetiology. Moreover, diverse strains of these viruses are in circulation which are known to vary widely in pathogenicity with the majority potentially of low or no pathogenicity and accordingly high levels of these virus may be detected in young, healthy, unaffected birds. Other confounding factors in attributing disease aetiology includes the presence of maternally-derived antibodies, route of transmission (from the hen versus their environment) and the age at which birds become infected, since increasing age appears to confer resistance. All these factors have also hampered the identification of the virus(es) responsible for stunting diseases. The specific aims of the proposal are to: 1) identify a virus or viruses that are actively replicating within the lesions associated with stunting disease and that are present in many similar lesions from clinical cases. Histopathology and laser microdissection will identify and excise lesions from which nucleic acids will be sequenced to determine which virus strain(s) is present at levels indicative of an active infection. 2) confirm that the virus or viruses identified are aetiological agents of stunting; firstly, by cellular and tissue-based methods in the laboratory and secondly, by challenge studies of healthy chickens using purified isolates of the identified virus(es), which will demonstrate the development of hallmark lesions and associated clinical signs typical of stunting disease. These objectives will be achieved through the use of state of the art imaging and genomic methodologies, and through collaboration with the Industry (Moy Park, which is one of UK's top broiler meat producers and St David's Poultry Team, the leading poultry veterinary practice in the country). Currently there are no commercial vaccines to prevent stunting or specific treatments for affected flocks so that affected birds are culled. The results of this project will provide evidence of the causal agents of stunting that will enable improved diagnostics and potential interventions for veterinarians, and for vaccine companies to develop appropriate vaccines.

With more than 50 billion chickens reared every year for both eggs (layers) and meat (broilers), poultry is one of the largest and fastest growing food systems worldwide. Poultry already outcompetes other meat markets and by 2025 more than half of all the meat produced globally is forecasted to be chicken. In the UK, growth in the poultry sector has soared in recent years, with an estimated contribution of £3.3bn to UK GDP in 2014. According to Defra, the UK poultry meat production increased to 1.8 million tonnes in 2017, with broilers accounting for around 85%. Currently, the UK is about 75% self-sufficient in poultry meat, and poultry meat represents the only UK livestock sector capable of quickly scaling-up production to support increased self-sufficiency of the UK. Ensuring consistently high fertility rates is key for meeting the demands of this expanding market. Fertility in these flocks is notoriously variable and tends to decline as birds get older, through reproductive ageing. Poor male fertilising efficiency requires increased female exposure to males, which has additional repercussions because males are often aggressive to females, which reduces female condition, health and overall fecundity. Even small improvements in the fertility of breeding stocks have vast financial consequences; e.g. a 1% variation in fertility in broiler flocks was estimated to be worth hundreds of millions of US$ in the US market. It is becoming increasing clear that in several organisms, the proteins contained in the male seminal fluid can have a drastic influence on fertility by modulating sperm swimming velocity, sperm storage within the female reproductive tract, probability of fertilisation and female behaviour after mating including female receptivity to further matings. The advent of proteomics and the publication of a draft genome of the chicken present a unique opportunity to investigate the role of seminal fluid proteins in poultry fertility. Our recent work has characterised the seminal fluid proteome of natural ejaculates of a population of red junglefowl (the species that has given rise to the domestic chicken) and has shown that the seminal fluid proteome of these birds is complex with more than 1500 seminal fluid proteins (SFPs) identified so far, including proteins involved in various known biological functions e.g. immune responses and antibacterial defences, as well as sperm maturation and sperm motility. Our work has shown that some of these SFPs are associated with in vitro measures of sperm quality. Importantly, our work has further shown that the seminal fluid proteome undergoes rapid and marked compositional changes in response to socio-sexual factors such as the sexual familiarity of a female partner and the social dominance of a male, and longer term changes, as males age, with males that are able to retain high sperm quality in advanced age having a distinct seminal fluid proteome. In this project, we capitalise on this wealth of preliminary knowledge to develop a research programme, to identify seminal fluid proteins (SFPs) that are important in maintaining lifelong fertility in male poultry. We use the red junglefowl as benchmark experimental system that has not been influenced by domestication and artificial selection, to characterise proteomic repertoires associated with rapid responses to socio-sexual conditions (objective 1) and reproductive ageing (obj. 2), and identify SFP signatures causally linked to fertility. We then confirm the role of seminal fluid in driving variation in fertility (obj. 3), and validate the commercial relevance and applicability of these findings by investigating patterns of intra- and inter-male variation in fertility-linked SFPs in commercial meat-production domestic chicken lines (broiler breeders, obj. 4). Collectively, these results will help us identify proteins linked to poultry fertility, which will inform new strategies to improve fertility in commercial stocks.

Marek's disease (MD) causes paralysis and tumours in chickens. It is caused by serotype 1 strains of the Marek's disease virus (MDV-1) which is shed from skin of infected chickens and persists for many months in dust in contaminated poultry houses. It is highly contagious and spreads to other chickens by inhalation. MD is a major disease affecting poultry health, welfare, and productivity, with annual estimated loss to the global poultry industry of $2 billion. MD is endemic in UK poultry but is effectively controlled by live vaccine viruses, which are harmless relatives of MDV-1, and include CVI988, HVT, and MDV serotype 2 (MDV-2). MDV-2 vaccines are widely used in the Americas and Asia but not in the UK. However, by testing samples collected from poultry farms, we found MDV-2 is widespread in the UK. MDV-2 strains circulate freely and naturally at high levels and persist long-term in the flock, but little is known about them; are they derived from vaccine strains which 'escaped' from imported poultry, or are they naturally occurring strains? Chickens can be infected with any combination of MDV-1, MDV-2, and vaccine viruses at the same time (co-infection). We have found MDV-2 in healthy chicken flocks, as well as flocks that have MD. We would like to know whether co-infection with MDV-2 affects flock health and disease, and production parameters such as egg production and mortality, and whether certain MDV-2 strains could be used as effective recombinant vaccines against MD and other poultry diseases. Our objectives are to: (1) Investigate prevalence of naturally occurring MDV-2 infection in the field, and it's influence on flock productivity, immune responses and disease, (2) Characterise MDV-2 field isolates and (3) Exploit novel MDV-2 as potential viral vectors for novel recombinant vaccines. The project is a partnership with poultry industry vets. We will select two MDV-2-positive and two MDV-2-negative flocks for two bird types (broiler-breeder, layer) for regular sampling to collect blood samples from chickens and dust from the housing sheds. We will also collect data on flock health and productivity. At Pirbright, we will test the samples by 'polymerase chain reaction' to detect the genetic material of MDV-1, MDV-2 and vaccine viruses to show the kinetics of MDV-2 infection and shedding, and the frequency of co-infection with MDV-1 field strains and vaccine viruses. Using mathematical modelling, we will also investigate dynamics of transmission of MDV-2 within flocks. We will determine variability of MDV-2 strains by sequencing the virus genetic material and comparing with known MDV-2 strains. We will study the characteristics of selected MDV-2 strains by growing these viruses in cell culture then using them to infect chickens under controlled laboratory conditions to examine replication, persistence, clinical signs and transmission of MDV-2. Most MDV-2 strains have characteristics which make them suitable as vaccines against MD: they do not cause disease, they grow well in the chicken and persist for many months, and they are easily transmitted between chickens to maintain a high level of exposure of the flock to vaccine virus. Furthermore, MDV-2 can be genetically engineered to carry genes from other important poultry viral pathogens, e.g., infectious bursal disease virus (IBDV) and Newcastle disease virus (NDV); a recombinant 'vectored vaccine' like this could potentially protect chickens against IBD and ND as well as MD in a single vaccination. We will engineer an appropriate MDV-2 strain to create a 'rMDV2-IBD-ND' virus, then test its ability to protect chickens against these three diseases under controlled laboratory conditions. This study is important to understand the effect of widespread MDV-2 infection on health and productivity of commercial poultry flocks. A new recombinant MDV-2 vaccine would be a useful addition to the set of live virus vaccines used to control MD and other poultry diseases.

Higher agricultural productivity and sustainability is critical to meeting the global challenges of food security in the presence of climate change. Legume crops are a critical source of plant-based proteins for people and animals. As the world demand for animal products increases, the demand for vegetable proteins as animal feedstocks also rises and the UK in common with other countries faces a shortfall in domestic vegetable protein production capability. In the EU 70% of the protein fed to animals is imported, mostly soyabean or soya meal with soya meal accounting for 33% of the protein in UK livestock feeds. In 2011-12 UK imports of soya products reached 1.83 million tonnes, the majority of this being transgenic soya imported from South America. Increasing the amount of UK grown protein to replace imported soya products is recognised as a major challenge for the UK animal feed sector. In this LINK proposal we will develop and apply new genetic approaches to enhance the nutritional value (protein and water soluble carbohydrate) of the pea (Pisum sativum L.) seed, to increase the use of pea as a high quality feed in animal diets, reducing the UK protein deficit from the import of soya products and also delivering environmental benefits to livestock production systems. The proposal builds on knowledge gained in BBSRC, EU, Defra, Innovate UK and levy board-funded research on the genetics and agronomy of pulses that have led to the development of novel lines of pea with higher protein content. We will use our expertise in plant genomics, pea genetics and breeding, agronomy, plant chemistry and animal nutrition to integrate the germplasm with improved grain composition into improved pea varieties. With industry partners from the poultry and pig sector as well as crop developers, we will analyse the impact of replacing soya with these new pea varieties in feed rations on the growth of monogastrics and poultry and the economic and environmental impact of their inclusion. Although the focus is on poultry and monogastrics, the project will provide information on the value of including these new pea lines for other sectors (ruminants and aquaculture).

It is estimated that environmental mitigation costs are in the region of $1 trillion per year of which ~$120 billion is provided largely by government and charitable foundations. Private investment is relatively small, providing a large untapped funding source that needs to be mobilised to help cover the massive shortfall. There are many impediments to private sector investment, including lack of appropriate and marketable financial instruments that can provide a satisfactory level of assurance regarding the outcome of environmental projects. The problem is exacerbated, in terms of biodiversity, due to the lack of financial value attributed to biodiversity and biodiversity assets. Current financial instruments that support the environment comprise mainly of green bonds. While the green bond market is expanding, these assets are not environmental performance related instead representing a commitment to simply fund more "straight forward" sustainable projects such as tree planting and renewable energy. As a result they have been linked to "green washing". FRIBO will identify with stakeholder partners (from the finance, land-use, wildlife, ecosystem service civil society and policy sectors) the opportunities and impediments to developing more fit-for-purpose performance related financial instruments that would be attractive to private investors while incentivising the delivery of real nature-based solutions (NBSs), with a focus on improved and measurable biodiversity outcomes. Examples of candidate instruments are Environmental Impact Bonds (EIBs); they link financial return to the success of the intervention by repaying the initial loan and interest using the financially valuable and marketable benefits of the project, such as carbon offsets, or cost reducing outcomes such as reduced flood damage, to pay off the full project costs. They are termed as "pay for performance" instruments. Sustainability Linked Bonds (SLBs) work in a similar way, but depend on meeting broader pre-defined sustainability goals such as reduction in GHG emissions to avoid penalty payments. FRIBO will analyse the linkages and drivers within the Finance-Biodiversity Nexus to assess opportunities and impediments to progress. Extensive stakeholder interaction, face to face, virtual and via surveys and questionnaires, will be conducted to identify key areas of research needed and critique optimum strategies for developing and incentivising the instruments. Challenges that will need to be taken into account are: how to measure and financially value biodiversity and biodiversity outcomes and NBSs in a market context (FRIBO will exploit links with existing NERC projects in this area), defining an improved biodiversity outcome; developing suitable biodiversity metrics; the role and function of Environmental Impact Assessments, the role and function of biodiversity in NBSs; the evaluation and quantification of wider societal benefits; identifying how to obtain additional payments for the wider biodiversity benefits that are realised in NBSs; the immaturity of EIBs and NBSs requires that their marketability within the financial sector requires study. Finally, there may be unintended consequences such as divesting/offshoring of environmental damage, non-equivalence of some "offsets" such as the carbon and biodiversity value of new tree plantations to offset deforestation of old growth forests;. Comprising a core group of researchers from Queen's University Belfast and Newcastle University, with a range of national and international partners, FRIBO will produce a Strategic Research Agenda that will identify the research required to address these challenges and accelerate investment in biodiversity related NBSs. In addition, FRIBO will produce an Implementation Roadmap that will outline key activities and timelines that need to be undertaken in order for stakeholders to implement Biodiversity focused "Rewards for performance" investment schemes.