Anopheles mosquitoes only bite in the evening and at night. It is possible to protect oneself by wearing protective clothing and using an undamaged mosquito net. Effectiveness is increased by treating the net with pyrethroids (insecticides) such as permethrin (Permas®, Peripel®), lambda-cyhalothrin (Iconet™) or deltamethrin (K-Otrine®). This will increase further in importance in the future. The substances are derived from pyrethrum, a product originating from Tanacetum cinerariifolium (= Chrysanthemum cinerariifolium) and C. coccineum, herbaceous composite-flowered plants (fam. Compositae) which are similar to large daisies. In emergencies where mosquito nets are logistically difficult (e.g. camps after heavy floods), impregnated blankets can be used. These will probably be less effective than impregnated bednets.

• The term "pyrethrum" refers to the powder made from the dried flowers of Tanacetum cinerariifolium.
• The term "pyrethrins" refers to the 6 insecticidal components which are found in the powder. All 6 are organic esters formed from one of two acids (chrysanthemic and pyrethric acid) and one of three keto-alcohols (pyrethrolone, cinerolone, jasmolone). Pyrethrins are unstable molecules which degrade within a few days in light and warmth. They can be stored in a refrigerator at –4°C.
• The term "pyrethroids" refers to the semi-synthetic derivatives of pyrethrins. These products have the advantage that they are much more stable than pyrethrins.
• The plants with the highest concentrations of active constituents grow well in sunny, semi-arid regions at 1600-2000 metres above sea level. Kenya, Tanzania and Ecuador are important producers. They are best harvested during a warm, dry period when the flowers are fully developed. The plants are dried after harvesting, and finely ground. Extraction may be with kerosene, paraffin or alcohol.
• Often a synergist is added, such as piperonal butoxide or various oils.
• Pyrethroids have low toxicity for mammals including humans but are highly toxic, e.g. to fish. They are sometimes used for illegal "chemical" fishing.

Fine-mesh gauze can be applied to windows and ventilation shafts. One good argument for using a mosquito net is the fact that it also protects from nuisance insects such as Culex mosquitoes and bedbugs. In regions where there are few Culex, people are not so ready to use a net: after all they cannot see or hear any mosquitoes. (Anopheline mosquitoes fly noiselessly.)

Insecticides based on pyrethrum can be dispersed by means of spraying (spray gun), evoparation (heated electric plate) or burning (mosquito coil, e.g. with esbiotrin). Insecticides can also be applied to the walls or to the curtains by the windows. There are also various insect repellents. DEET (N,N-diethyl-m-toluamide, now called N,N-diethyl-3-methylbenzamide) is moderately active and can be applied as an alcoholic solution to the skin. This produces a sticky effect when the alcohol evaporates, however. The effectiveness is only moderate. DEET is absorbed through the skin and is eliminated quickly via the urine. There is no accumulation in the body. Other repellants include KBR 3023 (Autan active®) which has the same effectiveness as DEET, but is non-irritant and does not affect plastic (spectacles), dimethyl phtalate (low activity) and ethohexanediol. Essential oils such as citronella, cedar, eucalyptus, neem (obtained from the tree Azadirachta indica) and geranium are short-acting and not very effective. People who need to stay in regions which are heavily infested with mosquitoes, can apply permethrin to their clothing. Taking extra vitamin B and electronic buzzers are of no use.

The product containing IR3535 (ethyl butylacetylaminopropionate) has been subjected to well- controlled field studies. The results demonstrate a protection time for the product of up to three hours or more against mosquitoes. Case studies are not a valid substitute for repellent field studies. Only field studies are to be used to establish efficacy of insect repellants.

Chemoprophylaxis is in the first instance intended as prevention of P. falciparum malaria. No single drug which is taken preventively, is 100% active against sporozoites and no single drug prevents the formation of liver forms (except primaquine). While taking the drugs no vivax or ovale malaria will occur, but after they have been discontinued an attack with these plasmodia is possible in the following months or years. In view of the extensive resistance of P. falciparum, at present no 100% satisfactory protection against this latter parasite is possible. Advice as to whether or not to take medication, and which kind of drug to take, will depend on the region and differ from person to person (short journeys, resident, local population, pregnancy, young children, allergy, chronic diseases, use of other drugs and so on). Recommendations vary from country to country.

In regions with only P. vivax and/or sensitive P. falciparum (WHO zone A) chloroquine 300 mg/week will suffice. In zone B a combination of chloroquine (Nivaquine®) 300 mg/week and proguanil (Paludrine®) 200 mg/d is recommended. If necessary chloroquine 100 mg/day can be taken with proguanil 200 mg/day (= 1 tablet of Savarine® daily). For shorter stays in zone C (<3 months) mefloquine is recommended (Lariam®, 1 tablet per week). An alternative is doxycycline 1 tablet per day (beginning 1 day before departure until 4 weeks after return). Malarone, 1 tablet per day beginning 1 day before departure until 7 days after return, is an easy and effective prophylaxis, but the drug is expensive. For longer stays, the combination Nivaquine®/Paludrine® is advised during the first years, together with the availability of a stand-by treatment in case of a breakthrough. For people who live for many years in such regions it is optional to discontinue the Nivaquine®/Paludrine®, but to still have a treatment on stand-by (e.g. Malarone®, quinine vibramycin, artemether). An annual eye examination to check for chloroquine toxicity is recommended for people who take Nivaquine® for many years. In practice this is sometimes difficult to achieve. The local population should not take chronic chemoprophylaxis (most people are semi-immune). There are some high-risk groups: e.g. pregnancy (during pregnancy, in particular in the second and third trimesters and also immediately post-partum, the immunological resistance to malaria falls). The advice that is given here should take into account the general state of health and local guidelines.

• Personal physical protection (impregnated mosquito net, repellents, clothing that covers, insecticides in rooms)
• Stand-by treatment (Malarone, artemether, quinine + vibramycin)

It has been possible to protect humans against malaria by vaccinating them with irradiated sporozoites. This method is not practical, however, on a large scale. To date there is still no effective vaccine available. Research into a vaccine is based on a number of possibilities. An immune response can be triggered against sporozoites, against liver forms, against erythrocytic forms and/or against gametocytes but this does not necessarily have a protective effect. An effective malaria vaccine is not likely to be developed in the foreseeable future.

Most initial work in developing a vaccine was carried out by studying the proteins found in sporozoites (circumsporozoite surface protein). The idea was that stimulated antibodies, aimed at invariant repetitive amino acid sequences which occur in natural proteins, would eliminate the sporozoites, in the hope of preventing infection (stopping the infection before the liver phase). The repetitive sequences have little or no immunogenicity, however, because they are similar to proteins which are present naturally in the human body. Even if antibodies can be produced, sporozoites only remain in the circulation for 30 minutes. There is therefore little time to neutralise all parasites by means of antibodies. Still, experiments were irradiated sporozoites were used as antigen showed important protection. If sporozoites could be cultured en masse, this would be a watershed. Another route was followed in the development of a cocktail of synthetic antigens. In 1987 Manuel Patarroyo (Colombia) demonstrated that a synthetic peptide vaccine (SPf66) is immunogenic and protects Aotus monkeys against challenge infection with P. falciparum. It also proved safe and partially protective in test subjects in Latin America. It was subsequently tested (double-blind, randomised and placebo-controlled), first in La Tola, Colombia (39% effectiveness) and then (independently) in Idete and Kilombero, Tanzania, regions with high transmission, and in Gambia. In 1994 the results became known. The effectiveness of 3 injections against clinical malaria was 31% (Tanzania) and 0% (Gambia) and the incidence of parasitaemia was the same in test subjects as in the control group. There was no adequate pre-erythrocytic immunity. Other later trials were equally disappointing.

In 1997 the first favourable results became known of a vaccine formed from a fusion protein of

• part of the circumsporozoite antigen of P. falciparum (called RTS, a polypeptide consisting of amino acids 207-395) and
• hepatitis B Surface Antigen (HBSAg) (a protein chain with 226 amino acids). The vaccine was administered with an adjuvant (AS02). The new product is at present still in the experimental phase. The initial results suggest a safe, immunogenic pre-erythrocytic vaccine which offers significant protection against natural P. falciparum infection.

A randomised and controlled study in the Gambia on 306 volunteers showed RTS,S/AS02 to be safe, immunogenic, and to provide significant protection against natural P. falciparum infection. Larger trials are being planned.

Mosquito control is a speciality in itself and includes the use of insecticides, treatment control of breeding grounds and biological control. Each ecological region requires its own individual approach: savannah, primeval forest, agricultural areas with or without irrigation systems, the margins of uplands, desert margins and oases, city environments, coastal and marsh regions. There are many difficulties: interventions need to be maintained over large areas for very long periods of time, mosquitoes quickly become resistant to insecticides, many people will not allow their houses to be sprayed, high costs, shortage of staff, ecological collateral damage due to insecticides, political instability which interferes with long-term planning. Border regions with rapid expansion and unstable social situations, e.g. mining (precious stones, gold), as in Thailand or Brazil, have their own specific problems. An exclusively technical approach will not be possible without simultaneous improvement in the social and economic conditions of the population at risk.

There are three major enzyme families implicated in insecticide resistance: (1) carboxylesterases, (2) glutathione transferases and (3) cytochrome P450s. Genomic analysis of Anopheles gambiae reveals a considerable expansion of these supergene families in the mosquito.

To date the effectiveness of biological control methods (e.g. with fish which eat mosquito larvae, such as gambusias (Gambusia affinis) and guppies (Aplochelius)) has still not been clearly proven. Some fish within the Nothobranchius and Cynolebias genera, have drought-resistant eggs, and can be placed where water has collected temporarily ("instant fish"). Toxorhynchites mosquitoes are large metallic-coloured insects with a curved proboscis [Gr. "toxo" = bowed; "rhinos" = nose) which they use to drink plant juice and nectar. They do not suck blood. The larvae are predatory upon other mosquito larvae, but to date the use of this natural enemy has not produced convincing results.

A female mosquito copulates only once and stores the sperm in a spermatheca. If insemination takes place with a sterile male there can be no reproduction. The massive release of sterile male mosquitoes is a technique which has shown good results against other insect species (screw worms, tsetse fly), but would be of limited use agains malaria transmission.

A certain spider (Evarcha sp.) are only 8 mm long, but stalk female mosquitoes far bigger than themselves. The young spiders particularly target mosquitoes which are full of blood, puncturing their abdomen, killing the mosquito and siphoning out the blood. Because they target Anopheles gambiae mosquitoes -the most effective malaria vector on the planet- the spiders offer the possibility of biological control of malaria mosquitoes.

Among the 400 species of Anopheles mosquitoes, only a limited number can be infected with plasmodia. The other mosquitoes are genetically resistant to this infection. If the natural mosquito population could be replaced by a genetically resistant population it would stop the transmission of malaria. There are a number of problems, however. First, the mosquito genes which make the parasite’s cycle in the mosquito impossible would have to be isolated and understood. It would then be necessary to find a way to bring those genes into a natural vector and to make them function properly. Thirdly and most difficult, the transgenic mosquito must have a competitive advantage over the natural vector in the Darwinian battle for survival. It is highly unlikely that a mosquito strain selected in a laboratory would be able to compete with wild-type mosquitoes. New, malaria-resistant mosquitoes would probably not be able to force out a natural population (so they would need to have a selective survival advantage). They should of course also not transmit other diseases. Fourthly, the Anopheles gambiae populations have various subpopulations which do not mix much. This partially sexual isolation of the natural mosquito populations would make the transgenic vector method extremely difficult.

One possible approach would be to spread a gene which codes for resistance in the mosquito in a wild population, e.g. if it can be coupled to transposons (these small DNA sequences are not inherited according to Mendel’s law, i.e. all the progeny contain the elements, cf. the transposon P in Drosophila). A reasonable amount of work has already been carried out on the transposon Minos which can penetrate the genome of Anopheles stephensi. Research into the use of Wolbachia, intracellular bacteria related to rickettsia, is a hot topic. They take their generic name from the microbiologist Wolbach of Harvard Medical School (1920). These non-pathogenic symbionts can quickly spread in a natural population. They cause a reproductive imbalance in mosquitoes (an infected population replaces a non-infected one). Thus, for example, Wolbachia pipientis infects gonadal tissue from both male and female mosquitoes. [Symbiont-free insects can be cultured in the laboratory by feeding them tetracyclines]. An infected female can pair successfully with both infected and non-infected males and her progeny are themselves fertile whereas a non-infected female can only pair productively with a non-infected male. Insemination of a non-infected female by an infected male leads to non-viable ova (cytoplasmic incompatibility). An infected female lays infected eggs. The general consequence is that infected females have a reproductive advantage. A mosquito population infected with these bacteria quickly replaces a virginal population. Transgenic Wolbachia which interfere with the maturation of malaria parasites in a natural vector, are a theoretical possibility which is being studied. In the genus Anopheles there are, however, no Wolbachia known which infect natural populations. The last word about such transgenic mosquitoes has not yet, however, been said. If the public outcry against genetically manipulated food is a guide, any method which uses transgenic malaria vectors will have to be able to convince public opinion that there is no better alternative.