Nematode parasitism of arbovirus vectors: effect of mermithids on Culicoides biting midges and their potential use for disease and vector control

Biting midges are a group of tiny flying insects that include a notorious species that plagues parts of Scotland. They are one of the most abundant blood-feeding insects and occur throughout most of the inhabited world. Certain species commonly found on farms are important carriers of livestock viruses in Europe. Methods to control these insects have so far been unsuccessful.
The emergence of the livestock diseases bluetongue (BT) in 2007 and Schmallenberg (SB) in 2011 in the UK highlight that midge transmitted viruses are a potential threat to the UK. They also spread African horse sickness (AHS), a highly lethal disease of horses, considered a risk to Europe. To date, over 50 viruses have been isolated from midges, causing significant economic impact through clinical disease and animal trade restrictions. Outbreaks of BT are ongoing in mainland Europe, so further outbreaks may occur in the UK. Currently we rely on vaccinating animals against some of these diseases, as control of insects transmitting them has been unsuccessful. Such vaccination is not routinely undertaken in UK livestock, and tends to only occur once an outbreak has been confirmed. Vaccines licensed for use in Europe do not exist for many diseases, including AHS and many types of BT. It is therefore paramount to determine other ways to manage such outbreaks to allow informed mitigation procedures in advance of, or in the event of, an outbreak.
My aim is to investigate the effects of a highly prevalent parasitic worm (mermithid nematodes) on the biology of Culicoides midges and to determine whether they affect a midge's ability to be infected by and transmit livestock viruses. Mermithids are a major group of nematode that live inside the larvae of midges, and other insects, often killing them when they emerge; but may be carried into the adult insect, sterilizing them as a result. These mermithids are present in UK midges and the successful use of mermithids to control mosquito populations suggests that there may be potential in their use for midge control, yet no information exists on mermithid parasitism of these midge vectors in Europe.
At the University of Liverpool, using Culicoides from across England and Wales, I aim to study the distribution of these nematodes and assess whether this changes between years. Despite their importance, very few mermithids have been successfully identified so I will use DNA-based techniques to determine the species parasitising UK midges, before establishing reliable identification techniques to enable other researchers to build on such work in the future. Any parasite which may be useful for Culicoides control must have an economical method of mass rearing, so a laboratory rearing method will be established and the possibility of storage and transportation determined, allowing field testing at a later date. Similar rearing methods have been successful in the USA, and this part of the project will be undertaken in collaboration with the University of California, Riverside. Most crucially, the effect of parasitism by mermithids will be assessed on the lifecycle, reproduction and fitness of the midges before being infected with BT and SB viruses and assessed for their ability to transmit virus, the dosage of virus needed to achieve infection, and their survival. Here both wild-caught and colony reared midges, supplied by the Pirbright Institute, will be infected with mermithids and virus at the Liverpool School of Tropical Medicine.
This project will gather information relevant to a wide range of arthropod vectors, such as UK mosquitoes, as well as other humans and livestock vector-borne diseases. In the event of an outbreak the work may provide information for more effective vector control and will arm policy makers with information that can be useful for i) understanding methods to control midge-borne diseases; ii) reducing the chances of an outbreak occurring; iii) more effectively dealing with an outbreak occurrence.

Culicoides biting midges are the most important biological vectors of livestock arboviruses in Europe. The emergence of bluetongue (BTV) and Schmallenberg viruses (SBV), with their economic impacts through clinical disease and animal trade restrictions, provides evidence of the threat they pose to UK livestock. Despite their importance, methods to control these insect vectors remain unsuccessful and control of such diseases relies on virus-specific vaccines. Mermithids are a major group of parasitic nematode worms that utilise the larvae of midges and other insects as definitive hosts, often killing host larvae on emergence, or being carried to the adult stage, sterilizing host insects as a result. The successful use of mermithids to reduce mosquito populations in the USA suggests their potential use in the biological control of Culicoides in Europe, including the UK. To date, almost no published data exists on mermithid parasitism of midges in Europe. The aim of this project is to determine the effect of mermithid parasitism on the life history characteristics, vector competence and survival of Culicoides in the UK, thereby assessing their potential as agents of biological control.

Culicoides collected from 144 sites in England and Wales will be used to determine the spatial and temporal distribution of the nematode species parasitising UK midges and the levels of parasitism and intersex species. Molecular and/or morphological techniques will be established to facilitate identification of mermithids of Culicoides in the UK. An in-vivo culture method and laboratory model for mermithid infection of midges will be established using C. nubeculosus and C. sonorensis colony larvae. The effect of parasitism on fitness parameters and survival of both adult and larval Culicoides will be determined using the infection model, before mermithid-induced effects on vector competence to both BTV and SBV are determined in colony and field-caught Culicoides.

LIVESTOCK INDUSTRIES/ FARMING BODIES: The main aim of the project is to help preserve UK animal and human health by providing information on the ability of mermithids to serve as Culicoides biological control, through effects on survival and vector competence. The National Farmers' Union, committed to the health and welfare of farmed animals, would be interested in on-farm methods of vector control. Entomopathogenic nematode treatment using standard pesticide equipment could be readily employed by farmers/ smallholders, so UK Government could pass disease control responsibilities to industry.
GENERAL PUBLIC/ TOURISM: Certain midges are a biting nuisance impacting on human public health with bites leading, in severe cases, to pruritic wheal-and-flare responses and scarring. They impact on tourism, outdoor pursuits, forestry and agriculture, particularly in Scotland. Research into novel methods of control may lead to benefits for the public and various outdoor industries. While the arboviruses transmitted by midges are largely of veterinary importance, some cause disease in humans. In the event of a future human disease outbreak, as a preventive measure against disease, or in terms of reducing nuisance insects, my work may provide information on effective vector control, leading to direct public benefits.
HORSE OWNERS/ EQUINE INDUSTRIES: Midges cause the hypersensitivity reaction sweet itch in horses, and transmit the highly fatal equine disease African horse sickness. Outbreaks have previously occurred in Spain, but no EU-licensed vaccines are available for this exotic disease. In the UK, organisations such as the Horserace Betting Levy Board, horse charities, and horse owners will be interested in novel methods of vector control which could be applied at yards.
INTEGRATED PEST MANAGEMENT (IPM)/ VECTOR CONTROL AGENCIES: Information on possible biological control agents of midges will be useful to local councils (often responsible for dealing with insect pests), and commercial companies developing insect control tools. This became clear when I visited a company developing low cost IPM programmes for large and small-scale farmers worldwide. Entomopathogenic nematodes fit well in IPM programs because they are non-toxic to humans, relatively specific to their target pest(s), and can be applied with standard pesticide equipment.
UK ECONOMY: The northern European BTV-8 outbreak caused significant costs for national economies, exceeding £1 billion in France in 2007, mainly through lost trade and control costs. The Scottish tourism industry loses around £286 million PA to the biting pests. In this proposal I focus on survival of mermithid-parasitised midges, and vector competence to two midge-borne viruses which can cause high animal production losses to the livestock industry. This study may assist in mitigating outbreaks (of disease or pests) that impact animal health and welfare, as well as tourism, by determining how parasitism can reduce insect numbers plus their ability to transmit disease.
POLICY MAKERS: I will arm policy makers with information for i) more accurate understanding of Culicoides control in the UK; ii) reducing the chances of disease occurrences; iii) more effectively dealing with outbreaks. In 2008, the Science Advisory Council's sub-group on epidemic diseases recommended that Defra should set up a campaign on innovative ways to control midge populations aimed at farmers and also investigate further Culicoides fundamental biology and how this might inform control strategies (Paper: SAC-ED (09) 05; recommendation 7). This project will provide important information for improving contingency plans produced by the devolved administrations (Defra, Welsh Assembly Government, Scottish Executive) and explores an innovative method of controlling midges. As the species to be studied are the vectors throughout northern Europe, the results will be of importance to the European Commission as well as other Members States.