Malaria remains the major tropical disease in the world today, killing almost 1 million people annually; approximately 243 million cases led to 863,000 deaths in 2008 . Incidence rates of malaria are 350-500 million per annum. However, over the last three decades malaria has impinged upon areas where it had previously been eliminated or had successfully been controlled, counterbalancing the achievements accomplished in recent years. Malaria is endemic in 108 countries with nearly 3.2 billion lives at risk of transmission. Half of the world's population is at risk of contracting malaria and individuals living in African countries are most at risk; South of the Sahara in Africa about 60% of malaria cases worldwide occur, 75% of global Plasmodium falciparum malaria cases occur and more than 80% of malaria deaths occur. Plasmodium falciparum is the most common cause of malaria in this region and is responsible for 18% of deaths in children under the age of five. Malaria is contracted from a bite by a female mosquito of the Anopheles family infected from an earlier blood meal from an individual infected by the parasite of the Plasmodium genus. The most common of which are Plasmodium vivax, Plasmodium ovale, Plasmodium malariae, Plasmodium knowlesi, Plasmodium falciparum (also considered to be the most pathogenic.) The World Health Organisation in their 2009 World Malaria report suggests that mosquito and parasite resistance to anti-malarial drugs are a "major threat to achieving global malaria control." Observation of drug efficiency has revealed early resistance to artemisinin containing anti malarial drugs leading to the emergence of resistance containment initiatives. Research from Zanzibar, Zambia and Sao Tome and Principe has highlighted that significant decreases in malaria incidences and deaths have been reflected in substantial declines in 'all-cause' deaths in children under five years of age, indicating that concentrated efforts to reduce malaria through control methods may help reduce child mortality by two thirds in African countries by 2015 . Throughout the last ten years there has been a growing need for alternative methods of vector control which keeps insecticide use to a minimum due to increase in development of resistance to them, environmental pollution, prohibitive costs and logistical difficulties . Currently there are limited studies on the effect of Anopheles larval control aiming to suppress malaria in tropical Africa but their results have indicated that the targeting of larvae, especially in man-made regions, can substantially reduce transmission of malaria under suitable settings. Also these methods are particularly apt for urban regions, where larval density is limited, especially perhaps in combination with indoor residual spraying (IRS) and other preventative approaches such as insecticide treated nets (ITN) .
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Larvivorous fish have recently been introduced in Kenya as a method of mosquito control. Oreochromis niloticus, commonly known as 'Nile Tilapia', is farmed in the highlands of western Kenya and in recent times has demonstrated potential as a bio control agent . It is of African origin and is native from Sudan through to West Africa. Oreochromis niloticus is of high economic value and is widely established outside its natural habitat and is considered to be the mostly universally cultured fish of the Cichlidae family, second to O. mossambicus. Nile Tilapia fish are prolific and rapid breeders, spawning every couple of weeks . There are many benefits to using these fish as a biological control method against the Anopheles mosquitoes: 1) Provide a sustainable source of income to the local community, because following six months after introduction to the wetland areas (ponds, rivers and lakes), the larger fish are ready to be harvested. 2) Oreochromis niloticus are low cost to buy so are commercially viable. 3) The fish would be widely accepted by the local community. 4) They are sizeable, quick to grow; once introduced into a habitat they inaugurate themselves very quickly.5) Mosquito larvae are unable to build up a resistance. 6) Oreochromis niloticus populations are commonly self-sufficient and are not dependent on presence of larvae to survive. 7) Have the ability to tolerate a broad spectrum of water conditions (can thrive in fresh, sea and brackish water). 8)Reports have shown that in the presence of Oreochromis niloticus, Anopheles larvae have a considerably longer developmental period; leading to adult females being smaller, impairing their ability to seek hosts and produce sufficient sized egg batches, therefore reducing their efficiency as vectors of malaria. However, there are some problems associated with introduction of the Nile Tilapia fish as they cause damage to their new habitats through their digging, causing destruction to river banks and increasing turbidity in the water, which decreases the light available to aquatic plants. Their rapid establishment into their new habitats also means that they compete with native fish fauna and because they only eat particular types of aquatic plants they cause changes in the local aquatic plant fauna. Other reports have shown that the use of larvivorous fish, in this case the guppy Poecilia reticulata, is more cost effective than a chemical larvicide 'temephos' by nearly 3 times, when travel, supervision, protective measures and application costs were considered . However the main cost of the project were the fortnightly surveys of the larvae but if these were to be carried out by the local community Kusumawathie and his colleagues found that costs could be reduced by 79%. It was also found that Poecilia reticulata was more effective in reducing larval density than temephos. Mosquito nets have long been a preventative method against mosquito bites and therefore malaria transmission.
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Treatment of the nets with insecticides also greatly increases their efficiency, particularly using a novel method known as 2in1, whereby the net is treated with 2 insecticides, to firstly increase efficiency and to secondly reduce resistance of the mosquito to the insecticides used . Recent projects report applying a resin with the insecticide to the net to prevent a decrease in efficiency following washes, known as PermaNet . On balance the resin used in the PermaNets to bind the insecticide to the net is effective in maintaining the viability of the nets following standard washes, and combination nets are more effective against mosquitoes and slow the development of resistance, but are more prone to contamination when washed, reducing their efficacy.
Figure 1: Map of the Union of Comoros, a grouping of islands in the Indian Ocean, situated off the East African coast in the northern region of the Mozambique Channel, between the northern part of Madagascar and north-eastern side of Mozambique. 
The closest neighbouring countries are Tanzania, Madagascar, Mozambique and the Seychelles.
Eight different Anopheles mosquito species have been documented in the country and the following Anopheles species are malaria vectors, A. Funestus, A. Gambiae, A. Merus and A. Coustani, and the main malaria parasite is Plasmodium falciparum . The islands of the Union of Comoros provide a very good prototype and a good area to carry out this research because the mosquitoes under investigation are confined to the islands, so there is no risk of contamination with other mosquitoes from neighbouring colonies. The National Project on the Fight Against Malaria (PNLP), which is run by the Global Fund, is supported by the government through the exemption of taxes on bed nets, insecticides, and malaria medication .
This research experiment aims to monitor and reduce mosquito larval density and incidence of malaria outbreaks following the introduction of larvivorous fish into wetland areas in combination with the use of insecticide treated bet nets in the Union of Comoros as a prototype for application across the African continent.If proven effective as a combinational method of control, further potential exists for a global use as malaria vector control.
This project will be carried out in two phases. Phase 1 will entail undertaking an environmental assessment of the effects of introducing the larvivorous fish to wetland areas of Grande Comore. Phase 2 is contingent on Phase 1and will involve implementing the experimental stages.
The experimental phase will run for a year on the main island of the Union of Comoros, Grande Comore (Ngazidja), which is a permanent malaria transmission zone made up of 396 square miles of lands. It will involve the introduction of the larvivorous fish, Oreochromis niloticus, commonly known as Nile Tilapia, which will also provide a source of income and food to the local community who will be assisting in the monitoring of larval density. By involving the local community in the project, costs will be reduced and they will be educated about better protecting themselves and their families from malaria.
Insecticide treated nets will also be made available to every house as part of Phase 2, (population ~400,000) on the basis that the average family is 4 members this would cost between $400,000 to $500,000 as ITNs cost between $4 and $5 in Mozambique, one of the nearest mainland countries .
Phase 1-Environmental Assessment
This will consist of commissioning an environmental assessment of introduction of the larvivorous fish, Oreochromis niloticus, which will be reviewed by the government of the Union of Comoros before Phase 2 can be instigated. Initial funding will be required for this assessment which will comply with local requirement but the proposal will be based around UK legislation which is in line with European directives . This initial assessment will also provide a base line larval count which can then be used as a control in the latter stages of the experiment. The environmental assessment will focus on the impact that introduction of the Nile Tilapia fish will have on the surrounding area, in particular the assessment will analyse the effects on: Biodiversity, human health, local fauna, population, local flora, soil, water and landscape. Both in the long term and short term, and whether the introduction of the fish will have a positive or negative impact on the issues listed above. Another point to consider is whether the Nile Tilapia fish will be acceptable to the local community as cultural heritage is an important aspect of the environmental assessment.
Phase 2- Experimental Approach
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My team will consist of myself and two post doctoral workers, it would be preferable to employ two post doctoral workers with the relevant experience from the Union of Comores, but if this was not possible two post doctoral workers from the UK would be acceptable. Funding would be required to cover my team's travel and their salary which would be based on UK rates of pay. A group of forty individuals from the local community will also be required to assist in monitoring the mosquito larval density, and will be paid a salary based on local rates of pay. The group of workers from the local community will also have the opportunity to farm the larvivorous fish. Following introduction of the Nile Tilapia fish to wetland areas and the distribution of ITN's to residents' houses, my team and I will monitor mosquito larval density and the incidence of new cases of malaria. Larval density of the ponds will be monitored weekly using dip tests, whereby 10 larval dips of 2.5 litres each will be taken from the edges of the ponds at random, from which the mean larval density per dip can be calculated. As a control, an untreated fish pond will also be monitored. Anopheline mosquito larvae will be indentified with a morphological key to differentiate from Culicine larvae.[5, 15] Observing the incidence of new cases of malaria is relatively simple and can be done through monitoring local health services and hospitals for the year that the field trial is in place.
Other Considerations- Arrangement of seminars, workshops and lectures open to the public discussing the project, its aims and expected results, in order to convey to the local population the need for this research. Educating the local people to cover drains, to channel water underground or into fast flowing streams, to drain wetland areas, or to build tanks to store water below or above ground (overhead tanks) will all prevent breeding of mosquitoes in stagnant water, therefore decreasing the incidence of malaria. This publicity and education will also require funding.
Justification of Resources- Funding is required for the materials and equipment needed to perform the dip tests, salaries of workers, renting of a lab base in Grande Comore (likely to be the capital, Moroni) and for travel of my colleagues from the UK (if post doctoral workers are UK based rather than from the islands of Comoros.)
Beneficiaries - If this project is successful in reducing Anopheles larval density and the transmission of malaria, many malaria endemic countries will be able to apply the same methods used, i.e. introduction of larvivorous fish and better distribution of insecticide treated nets where they most are needed. This will greatly reduce the number of cases of malaria and therefore deaths in these countries, particularly in Sub-Saharan Africa, where Plasmodium falciparum is responsible for 18% of deaths in children under the age of five.