Olfactory mechanisms underlying behavioural manipulation of mosquitoes by malaria parasites

Lead Research Organisation: London Sch of Hygiene and Trop Medicine
Department Name: Infectious and Tropical Diseases

Abstract

Parasites have the ability to manipulate the behaviour of the insects that are involved in their transmission to enhance their own reproductive fitness. For example, malaria-infected mosquitoes take larger blood meals than uninfected ones, and will take multiple blood meals. In a pilot study, conducted recently by the applicants, we demonstrated that females of one of the most important African malaria mosquito species Anopheles gambiae sensu stricto infected with Plasmodium falciparum, one of the most important parasites that causes malaria in human beings, were significantly more attracted to human odours than uninfected mosquitoes. This was the first demonstration of a change in An. gambiae s.s. behaviour in response to human odours caused by infection with the malaria parasite P. falciparum. This behaviour is of fundamental importance since it will enhance the efficiency of disease transmission. We propose to explore the mechanisms for this response. The hypothesis of the proposed study is that P. falciparum parasites manipulate the olfactory system of host-seeking An. gambiae female mosquitoes making them more sensitive to host stimuli and this causes infected mosquitoes to be more attracted to human hosts.

We propose to build on our preliminary study to quantify, using a windtunnel bioassay, the behavioural responses of An. gambiae s.s. female infected with human malaria parasites, P. falciparum, compared to uninfected females and determine whether the response depends on the lifecycle stage of the parasites within the mosquito. We will use olfactometer studies, which are designed to measure the behavioural response of the insects to human odours, within specially designed secure laboratories at the London School of Hygiene and Tropical Medicine. To determine the effect of a Plasmodium infection on the olfactory system (i.e. the mosquito's sense of smell), we will use an electrophysiology technique which allows us to record the response of the mosquito's antenna to individual compounds within the complex mixture that makes up human body odour. This, along with other state-of-the-art analytical chemistry techniques, will allow us to identify the volatile compounds that the mosquito can detect and to determine whether infected mosquitoes respond differently to the odorous volatiles emitted by humans to uninfected mosquitoes.

The results of our study will provide information that could be used to illuminate how malaria is transmitted between human beings by An. gambiae s.s. females. Importantly, new attractive compounds could be identified which could be used to develop improved mosquito traps for surveillance or trapping programmes that may even specifically target P. falciparum-infected An. gambiae s.s. females. This study will be a fundamental example of how parasites are able to manipulate their host to increase their own reproductive fitness.

Technical Summary

There is considerable evidence that pathogens manipulate the behaviour of their vectors to enhance pathogen transmission. For example, malaria-infected mosquitoes take larger blood meals than uninfected ones, and take multiple blood meals. We have recently demonstrated that P. falciparum-infected An. gambiae females were significantly more attracted to human odours than uninfected females. This was the first demonstration of a change in An. gambiae behaviour in response to olfactory stimuli caused by infection with P. falciparum. Despite the clear impact on the epidemiology of vector-borne diseases, parasite-mediated behaviour of haematophagous insects in response to host odours, and the chemistry and olfactory mechanisms underlying the behavioural responses, have not been elucidated. The hypothesis of this study is that P. falciparum manipulates the olfactory system of infected An. gambiae mosquitoes, causing them to be more attracted to human hosts.

We propose to build on our preliminary study to quantify, using olfactometer studies within BSL-3 laboratories at LSHTM, the behaviour of An. gambiae females infected with human malaria parasites, P. falciparum, and compare these responses to uninfected females, and determine whether the responses depend on the lifecycle stage of the parasites. Electrophysiological experiments will be done to examine whether P. falciparum infection has an effect on the olfactory system. New compounds (or blends) could be identified, using sophisticated analytical chemistry techniques, which could be exploited to develop a synthetic lure for application in surveillance or trapping programmes specifically targeting P. falciparum infected An. gambiae s.s. females.

This study represents a novel piece of biology exploring the olfactory mechanisms by which parasites enhance their transmission success. This is, to our knowledge, the first study to examine this type of behavioural manipulation by a parasite.

Planned Impact

Although there is ample evidence that pathogens manipulate the behaviour of their vectors to enhance pathogen transmission, and despite the impact on the epidemiology of vector borne diseases, parasite-mediated behaviour of haematophagous insects in response to host odours have hardly been studied. We propose to examine the vector-parasite-host interaction of Anopheles gambiae s. s. - Plasmodium falciparum, one of the most important interactions in the context of humanity, with malaria causing over 200 million human malaria cases and over 770 thousand deaths each year, and having a negative impact on the economic growth of the countries affected. The research in this proposal is likely to be of wide public interest and should generate high impact publications.

The development of novel and highly effective synthetic attractive blends for P. falciparum infected An. gambiae s.s. females will benefit researchers in the area of mosquito control, ecology and behaviour and control programmes that require improved monitoring systems. It will also generate new intellectual property which will be exploited appropriately by the partner institutions and could lead to the development of a product which would be marketed to mosquito surveillance and control programmes as well as the domestic pest control market world-wide. Current methods for monitoring and control of malaria mosquitoes rely mainly on human subjects or CO2. The former has ethical concerns, whereas the latter is not mosquito specific and its attractiveness to An. gambiae s.s. females increases enormously when combined with human odours. Using volatile blends selectively attractive to infected mosquitoes will, in combination with other measures, disrupt contact between infected mosquitoes and humans and therefore malaria incidence. Mass trapping of infected mosquitoes using the newly developed odour baits could, in the long term, create an evolutionary pressure for mosquitoes not being infected. In the long term, the health of travellers and local people in malaria endemic countries (particularly young children and pregnant women) will be substantially improved which will positively affect economic growth in sub-Saharan malaria endemic countries. Additionally, the development and commercial exploitation of novel mosquito control technologies will enhance the economic competitiveness of the UK.

The scientific knowledge gained from the proposed project will enable epidemiologists to improve mathematical models used to predict disease transmission and to plan mosquito control and elimination strategies. Including the effect of parasites on mosquito host-seeking behaviour in the models will increase the reliability of the models and therefore the impact of mosquito control strategies that are implemented based on the predictions of the models.

Additionally, the results of this project will be relevant to other systems where parasites manipulate their hosts and thus, the findings will also be of benefit to evolutionary biologists. It is likely that new avenues of research will be generated (for example understanding the mechanisms of manipulation by parasites at the molecular level; and understanding finer details of the olfactory system, such as olfactory receptors and odourant binding proteins). The project will build on the current knowledge of the individual investigators involved and new skills and knowledge will be gained during the proposed project that will be useful in future projects. The project will also create foundations for a strong collaborative relationship between the investigators and advisors beyond the proposed project, enabling them to target jointly other parasite-insect-host interactions.

Publications

10 25 50
 
Description Mosquitoes infected with malaria parasites have demonstrated altered behaviour that may increase the probability of parasite transmission. Here, we examined the responses of the olfactory system in Plasmodium falciparum infected Anopheles gambiae, Plasmodium berghei infected Anopheles stephensi, and P. berghei infected An. gambiae. Infected and uninfected mosquitoes showed differential responses to compounds in human odour using electroantennography coupled with gas chromatography (GC-EAG), with 16 peaks triggering responses only in malaria-infected mosquitoes (at oocyst, sporozoite or both stages). A selection of key compounds were examined with EAG, and responses showed differences in the detection thresholds of infected and uninfected mosquitoes to compounds including lactic acid, tetradecanoic acid and benzothiazole, suggesting that the changes in sensitivity may be the reason for differential attraction and biting at the oocyst and sporozoite stages. Importantly, the different cross-species comparisons showed varying sensitivities to compounds, with P. falciparum infected An. gambiae differing from P. berghei infected An. stephensi, and P. berghei infected An. gambiae more similar to the P. berghei infected An. stephensi. These differences in sensitivity may reflect long-standing evolutionary relationships between specific Plasmodium and Anopheles species combinations. This highlights the importance of examining different species interactions in depth to fully understand the impact of malaria infection on mosquito olfactory behaviour.
Exploitation Route The findings will advance the understanding of manipulation of mosquitoes by malaria parasites. It has also led to the discovery of some compounds that may be putative attractants for mosquitoes. These will need to be explored further to determine whether they can be used to develop improved traps for mosquitoes.
Sectors Chemicals,Healthcare,Other

 
Description This study is complete. Here, we examined the responses of the olfactory system in Plasmodium falciparum infected Anopheles gambiae, Plasmodium berghei infected Anopheles stephensi, and P. berghei infected An. gambiae. Infected and uninfected mosquitoes showed differential responses to compounds in human odour using electroantennography coupled with gas chromatography (GC-EAG), with 16 peaks triggering responses only in malaria-infected mosquitoes (at oocyst, sporozoite or both stages). A selection of key compounds were examined with EAG, and responses showed differences in the detection thresholds of infected and uninfected mosquitoes to compounds including lactic acid, tetradecanoic acid and benzothiazole, suggesting that the changes in sensitivity may be the reason for differential attraction and biting at the oocyst and sporozoite stages. Importantly, the different cross-species comparisons showed varying sensitivities to compounds, with P. falciparum infected An. gambiae differing from P. berghei infected An. stephensi, and P. berghei infected An. gambiae more similar to the P. berghei infected An. stephensi. These differences in sensitivity may reflect long-standing evolutionary relationships between specific Plasmodium and Anopheles species combinations. This highlights the importance of examining different species interactions in depth to fully understand the impact of malaria infection on mosquito olfactory behaviour. Some of the compounds we identified are putative attractants and repellents and we plan to investigate this further in future projects. These findings have advanced our understanding of the mosquito parasite host interaction significantly and will lead to further research projects. Our findings have also have also been used to inform the research community, our MSc community as part of the teaching programme, masters, and the public, about the work being done on malaria vectors. We have now published the work in one of the Nature journals - Scientific Reports: N. M. Stanczyk, V. A. Brugman, V. Austin, F. Sanchez-Roman Teran, S. A. Gezan, M. Emery, T. M. Visser, J. T. Dessens, W. Stevens, R. C. Smallegange, W. Takken, H. Hurd, John Caulfield, M. Birkett, J. Pickett & J. G. Logan Species-specific alterations in Anopheles mosquito olfactory responses caused by Plasmodium infectionScientific Reports Volume 9, Article number: 3396 (2019 We have also discussed the possible commercial exploitation of this technology with potential licensees including major international companies.
First Year Of Impact 2016
Sector Education,Healthcare
Impact Types Cultural,Societal

 
Description MRC
Amount £650,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 08/2017 
End 08/2020
 
Description ORN collaboration 
Organisation Vanderbilt University
Country United States 
Sector Academic/University 
PI Contribution A collaboration has been set up to investigate which olfactory receptors are associated with alterations in olfaction due to malaria infection. We will supply the collaborator with material from our laboratory and they will perform the molecular studies required to answer this question. We aim to write a joint publication.
Collaborator Contribution The collaborator will perform molecular investigations using material from our studies at LSHTM. We aim to write a joint publication.
Impact No outcomes yet.
Start Year 2014
 
Description Media interest 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Ultimately this has sparked much interest and we receive numerous requests for information from the public, requests for work experience from school students and student placements for university graduates (both local and international), as well as requests for information from other academics.

Impact unknown
Year(s) Of Engagement Activity 2013,2014,2015,2016