Differential attractiveness of humans to the African malaria vector Anopheles gambiae Giles : effects of host characteristics and parasite infection

Abstract

The results of a series of studies designed to understand the principal factors that determine the differential attractiveness of humans to the malaria vector Anopheles gambiae are described in this thesis. Specific attention was paid to the role of body emanations and infection (of humans) with the malaria parasite Plasmodium falciparum . The main findings of these studies are summarised in the following sections.Differential attractiveness of humans to Anopheles gambiae ( Chapter 1 )Although it has frequently been reported that human beings differ in their degree of attractiveness to mosquitoes, the principal causes that make certain individuals to be preferred more than others are not well known. This gap in knowledge has hindered the understanding of the transmission dynamics of malaria and other mosquito-borne diseases. From an epidemiological point of view, high malaria transmission rates are expected if mosquito vectors preferentially select infective humans for a blood meal, become infected, survive long enough for the parasites to develop to infective stages and proceed to bite uninfected individuals selectively. In terms of fitness, mosquito vectors would be better off if they select hosts that (1) are docile and less defensive so as to minimise feeding-associated risks of mortality, (2) have blood of a high nutritional value, (3) are free of (gametocyte) parasites, healthy and (4) have no anti-mosquito immunity. The answers to these epidemiological and fitness factors are still lacking. Furthermore, since host seeking is odour-mediated, the identification of chemical compounds responsible for attracting mosquitoes to their vertebrate hosts would help in developing traps that are useful for vector surveillance and control. Research in this area is rapidly advancing but has not produced tools for field application. It is felt that more research effort is needed so that new approaches towards understanding and combating disease vectors can be developed.Characterisation of mosquito blood meals using DNA markers (Chapters 2 & 3)The analysis of arthropod blood meals using molecular genetic markers was reviewed and forensic techniques borrowed from the Federal Bureau of Investigation (FBI), USA, used to evaluate the effect of blood meal size and extent of digestion on the ability to identify human DNA extracted from blood meals of Anopheles gambiae . The review recommended that proper and appropriate storage, determination of the concentration of host DNA and collection within few hours after ingestion are important parameters for improving the success of identifying blood meal sources of field-collected mosquitoes. Further, microsatellite markers were highlighted as being more appropriate than minisatellites in analysing blood meals that have been highly degraded e.g. through prolonged digestion. Also, mitochondrial DNA targets were recommended to be better than nuclear DNA targets for analysing blood meals that have been highly degraded. Blood meal size and (microsatellite) locus (analysed) were shown not to affect the success of amplifying human DNA extracted from blood meals of An. gambiae after having been digested for 0, 8, 16, 24 and 32 hours. However, a significant negative relationship between the time since ingestion and the success probability of obtaining positive PCR reactions among blood meals digested for between eight and 32 hours was demonstrated. There was no significant difference in the success probability of amplifying human DNA from blood meals of mosquitoes killed at zero and 8 hours after ingestion. The research demonstrated that not the quality of ingested blood, but the time since ingestion determined the success of blood meal analysis.Host characteristics and differential attractiveness of humans to An. gambiae (Chapter 4, 5 & 6) A tent olfactometer that accommodates complete humans as sources of host-seeking stimuli was designed, developed and tested. The olfactometer was used to study (1) differential attractiveness of humans to host-seeking An. gambiae and (2) how the differences, so elicited, are affected by human breath, body odour, heat and moisture. Nine human subjects were successfully ranked for their attractiveness to the mosquitoes based on (mosquito) responses to their complete body emanations encompassing body odour, heat and moisture. The nine subjects were classified into least (3 persons), medium (4 persons) and most attractive groups (2 persons). Breath was shown to reduce mosquito responses, whereas body odour was highly attractive. Breath was also shown to be an important contributor to between-person differences in relative attractiveness to An. gambiae . Whereas differential attractiveness of two human subjects for the mosquitoes could be demonstrated based on their total body emanations (breath plus body odour), the attractiveness of the two subjects did not differ significantly based on body odour alone. Body odour from either individual was consistently more attractive than total emanations from the other. The same results were obtained with another pair of individuals. It was concluded that breath, although known to contain attractive semiochemicals like carbon dioxide, may also contain compounds that inhibit attraction and may thus serve as an important contributor to between-person differences in relative attractiveness to this important malaria vector. The inhibitory effect of breath was postulated to be allomonal as it benefits the emitter (human being) but does not harm the recipient (mosquito vector). Body heat and moisture were shown to have significant effects on the attraction of An. gambiae to humans. In general, An. gambiae was more attracted to the individual whose body emanations were warmer but less moist than those of an opposing counterpart, in choice experiments. It was concluded that body heat and moisture influence host-selection by An. gambiae at short range and that their effect is probably achieved through interaction with breath components.Parasite infection and differential attractiveness of humans to An. gambiae (Chapter 7 & 8) A rare window of opportunity allowed for the investigation of the effect of clinical symptoms and parasitaemia due to Plasmodium falciparum on variability in human attractiveness to the malaria mosquito An. gambiae using the tent olfactometer previously developed. The relative attractiveness of an individual who was always more attractive than a specific counterpart (when both were uninfected) was suppressed when he (the 'putatively' more attractive individual) displayed clinical symptoms (fever and profuse sweating) of malaria caused by P. falciparum . This finding provided both new and alternative evidence, within the intricate web of Anopheles-Homo-Plasmodium interactions, that the malaria parasite P. falciparum influences the olfactory signals produced by human hosts. Field studies, in which a twin pair of male humans was recruited as a follow-up to the olfactometer study, found that being positive with malaria parasites (trophozoites and possibly gametocytes of P. falciparum ) is associated with higher numbers of attracted mosquitoes ( An. gambiaesensu lato ). This effect was not a consequence of the time of the year and the higher number of mosquitoes present in that period. It was recommended that further investigations be carried out. If these results are confirmed to be true, this work will constitute the first evidence that P. falciparum is capable of enhancing its own transmission by manipulating the physico-chemical characteristics of its vertebrate hosts in such a way that infected individuals are preferentially selected as blood meal sources by host-seeking malaria vectors.What questions remain unanswered (Chapter 9) Much as this thesis has provided some answers to why humans differ in their degrees of attractiveness to mosquitoes ( An. gambiae ), many more questions remain unanswered. The key questions in point include the following: (1) How can we tell whether one is more or less attractive to host-seeking mosquitoes? (2) How do parasites affect the host-seeking behaviour of their insect vectors? and (3) How do evolutionary processes related to host-seeking behaviour affect the fitness of mosquito vectors? The answers to these questions and those to a myriad of others are still waiting to be resolved