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Influence of Insecticide Resistance, Wolbachia and their Interaction on Chikungunya transmission

Periodic Reporting for period 1 - IRWIC (Influence of Insecticide Resistance, Wolbachia and their Interaction on Chikungunya transmission)

Reporting period: 2017-05-01 to 2019-04-30

Vector-borne diseases represent one of the most threatening public health challenges, such as malaria but also the emerging arboviral diseases, such as Dengue, Zika and Chikungunya. Vector competence represents the ability of an arthropod vector to acquire a pathogen and subsequently transmit it to another susceptible host. It is a complex trait that depends on specific interactions between vector and pathogen genetic factors, as well as environmental factors, including temperature, diet, pesticide exposure and microbial flora. An important question is how mosquitoes' adaptations (i.e. to insecticides and to symbiotic bacteria) influence its competence to transmit disease pathogens? The overall objective of this project is to assess whether insecticide resistance genes and Wolbachia symbiosis can affect the transmission of Chikungunya virus. It addresses in particular the question whether current (i.e. insecticide) and prospective (Wolbachia) vector control methods interfere with arbovirus transmission in short and long term.
After signing the contract, I first enroll in the BSL3 clearance training and I obtained the authorisation to work in a level-3 confined laboratory on may 22nd 2017. Note that th BSL3 labs at IRD are closed between december 20th to January 31th for decontamination and maintenance.

Biological materials:
In May 2017, I received the three Aedes albopictus mosquito lines from our two collaborators (Dr Calvitti, Italy and Dr Vontas, Greece): one susceptible, one resistant originated from a Greek population and one lab line that were trans-infected by a Wolbachia strain from Culex pipiens. The susceptible line (Malaysia) was from a different origin than the resistant one (Greece), this is problematic because the genetic factors important for vector competence are unknown ans it is important to compare two lines that share a common polymorphism. As a consequence, the Vontas lab had to prepare a field trip to collect the same greek population in the summer (July 2017). Then I reared these lines at ISEM to get the highest density in the cages and to get the mosquito accustomed to new feeding systems for artificial blood feeding (Hemotek® systems) and to new blood source (rabbit blood). In the mean time, I fixed a new insecticide susceptible mosquito line from the newly collected population. Based on a quantitative genetic approch, I was able to select males and females that had not the cce3a gene amplified (i.e. insecticide resistant gene) to make crosses. After mating, the offsprings were reared until adult emergence and analyzed for amplification at the cce3a locus. The ""apparent"" susceptible males and females were mated again to analyze the offspring. This step had been repeated four times to obtain an insecticide susceptible mosquito line from the same origin as the resistant one (october 2017). These two lines now share a common genetic background except around the insecticide resistant locus (cce3a). Then, I moved forward to prepare the asymbiotic mosquito lines.
The Chikungunya virus (Indian Ocean lineage) was already available at IRD. During the time I fixed the new mosquito line, I practiced to infect experimentally some random mosquito with Chikungunya virus in the BSL3 to master all the techniques used at IRD (secondment).

From February 2018, I started to infect experimentally only the susceptible and resistant mosquito lines (with natural Wolbachia symbionts). To this aim, I fed 40-50 mosquito females with rabbit blood containing 10^7 pfu/ml of virus. On day 1, 2, 3 and 6, I collected a batch of mosquito to collect their saliva and to dissect the head, the thorax and the midgut. These samples were stored in Trizol buffer for RNA extraction et virus detection. I repeated the feeding experiments but the molecular analyses were not performed yet. But I plan to continue acquiring data for this project.

Career prospects:
In July 2017, a permanent position had been opened for hiring a research scientist at INRA (french National Institute for Agronomic Research). I decided to apply so I contacted the teams that may be interested in my project and my profil. With the ASTRE unit at INRA, I redefined my research project on West-Nile virus transmission and I applied for this job position on august 2017. I have been selected for an interview in november 2017 and I succeeded."