My research focuses on infectious diseases, especially insect-borne. I am particularly interested on the interactions between the innate immune system of disease vectors and pathogens. Among the three leading global infectious diseases is malaria (others are AIDS and TB) that threatens almost half of the global population, infects over 400 million people every year and kills 1-3 million people, mostly young children in sub-Saharan Africa. It is caused by the protozoan parasite, Plasmodium, transmitted between humans through Anopheles mosquitoes. To study the interactions between vectors and pathogens, we use genomics, functional genomics technologies and reverse genetics, combined together in a systems biology approach. Below are some of our recent scientific discoveries and future directions.
We have pioneered the development of a series of functional genomics platforms for Anopheles research, including various microarray platforms and a full genome RNAi library. The first microarray platform consisted of 4,000 ESTs of the major African malaria vector A. gambiae and was used to study the mosquito immune response and refractoriness to Plasmodium. Later, we produced a 20,000 EST microarray that encompassed approximately 8,000 genes and used extensively by us and others in the research community to address various biological problems, including mosquito responses to viruses, insecticide resistance and developmental programmes. While this platform is still in use, we have generated new full-genome amplicon microarray (MMC2) and, more recently, oligonucleotide microarray platforms, which serve as the main tools in our transcriptomics research to study immune signalling and responses to specific pathogens. The design of MMC2 amplicons have allowed us to also create a double-stranded RNA library of all the mosquito genes. We are currently using this library to finely dissect immune modules and, in collaboration with mathematicians, to study the Anopheles immune system from a systems biological perspective. To complete our genomics toolkit, we called Single Nucleotide Polymorphisms (SNPs) in the A. gambiae genome using sequence traces from past and ongoing genome sequencing projects and are currently developing a SNP chip. We aim to use this to understand the genetic diversity in field mosquitoes that regulates susceptibility vs. refractoriness to P. falciparum and thus contributes malaria transmission.
Transcription profiling of A. gambiae has identified a Leucine-rich repeat encoding gene, LRIM1, as a key player in the mosquito immune system. LRIM1 is a potent antagonist of the development of the rodent malaria parasite, P. berghei, mediating lysis or melanization of ookinetes during their invasion of the mosquito midgut. Genetic epistasis experiments have revealed that the inhibitor of parasite melanization, CTL4, is part of the same immune module and acts downstream of LRIM1. In A. gambiae infections with the human parasite P. falciparum, this module appears not to have the same effects on parasite development. We are currently investigating whether this difference between the human and rodent parasites relates to their differential ability to evade the mosquito immune system or to differences in their levels of infection. In either case, this is thought to be the result of evolutionary co-adaptation between the host and the parasite, which we aim to study using population genetics approaches. Recent data show that LRIM1 is a member of a mosquito-specific gene family, which comprises additional parasite antagonists.
Although A. gambiae is a highly competent vector of human malaria, its sibling A. quadriannulatus is a non-vector. We have shown that A. quadriannulatus is resistant to infections by P. falciparum and the rodent model P. berghei. Resistance is controlled by quantitative heritable traits and manifested by lysis or melanization of ookinetes in the midgut as well as by killing of parasites at subsequent stages of their development in the mosquito. Orthologs of the Leucine-rich repeat proteins, LRIM1 and LRIM2, and the complement-like protein TEP1 are required in this reaction and their silencing transforms A. quadriannulatus into a highly permissive vector. Additional genes involved in this phenotype have been identified and are currently being investigated.
We have shown that the A. gambiae equivalent of the Drosophila Imd pathway is activated in response to bacterial infections and is essential for the survival of adult mosquitoes. This pathway is also involved in the killing of P. berghei in the mosquito midgut, perhaps through transcriptional control of the parasite antagonist, LRIM1. The key recognition receptor of this pathway is PGRPLC, as in Drosophila. PGRPLC exists as three main isoforms, all of which can bind peptidoglycan. Structural modelling has provided insights into how PGRPLC functions to control Imd pathway activation. The Imd pathway and its transcription factor REL2 are not involved with the mosquito fungal infections.
A genome-wide analysis of A. gambiae gene expression revealed a series of developmental transcription programs and tissue-specific patterns. Comparative analysis of these data together with Drosophila developmental expression has revealed a conservation of orthologous gene expression between these two insects. This similarity of expression is not correlated with the CDS similarity, indicating that expression profiles and coding sequences evolve independently. This is the first large-scale comparative transcriptomic analysis between two distantly related organisms. It has also identified clusters of co-regulated antiparasitic immunity genes which are currently being investigated.
In addition to being vectors of devastating parasitic diseases, mosquitoes act as vectors of several viral diseases including Dengue and Yellow fever, various encephalitides and Chikungunya (CHIK). The latter has recently become a major threat in countries of the European Union, with a major outbreak in Italy in 2007. We have developed a research programme to study insect responses to viruses, which may help develop future strategies to control spread of such diseases. The alphavirus O’Nyong Nyong (ONN), which is very closely related to CHIK and transmitted by A. gambiae and A. funestus, is used as a model system. Genome-wide transcriptional analysis of the A. gambiae responses to infection with the ONN virus identified a number of regulated genes; however, only few are part of the classical mosquito immune repertoire. This suggested that the mosquito response against viral infections is distinct from the immune response against bacterial, fungal or parasitic infections. The study of candidate genes and pathways is ongoing.
The genome sequence of the mosquito Aedes aegypti, which is vector of the viral diseases yellow fever, Dengue and CHIK, has allowed us to perform a comparative phylogenomic analysis of the insect immune repertoire. This analysis has revealed distinct and seemingly contrasting modes of evolution of genes involved in the different phases of immune signalling and the melanization genetic module. These dynamics reflect in part continuous readjustment between accommodation and rejection of pathogens and suggest how innate immunity may have evolved. The impact of these modes of evolution on the interactions of the vector with different pathogens is currently under investigation.