Lighting up fleas to understand the complex transmission mechanisms of Yersinia pestis – LUCY

To obtain the results of the project, it was necessary to dissect several hundred Y. pestis-infected fleas to analyze the infectious process. It was also required to produce libraries of Y. pestis mutants (consisting of hundreds of independent clones) and then test the ability of each of these mutants individually in the flea infection model, which involved the use of countless insects. This tour de force was only possible thanks to the development of a new methodological approach. This new method, fast and less expensive than the usual methods, is based on the use of bioluminescent plague bacilli, i.e. capable of producing light autonomously. Finally, after having identified mutants of Y. pestis defective to infect fleas, and thus the genes of the bacillus that are responsible for the transmission of plague by fleas to humans, we deployed bacteriological, biochemical and molecular approaches to better understand the role of the identified genes in the infectious process

The project has produced an innovative methodologies that have allowed the screening of a number of Y. pestis mutants in the flea at an unprecedented level and that will allow new discoveries in the near future. Beyond that, the project has greatly refined the model of how Y. pestis produces a transmissible infection in the flea both from a pathophysiological point of view and in terms of the molecular mechanisms deployed by Y. pestis to produce a transmissible infection. Finally, it has contributed to train several students, recruit a researcher and lead to new national and international interactions.

Our study added several genes to the list of genes required for infection by the fleas, the role of which remains to be elucidated. Moreover, we have only studied a small part of the plague bacillus genome, which comprises just over 4,500 genes. It is hoped that the discovery of other genes and their exact role in the chip will lead to new strategies to control the plague and hopefully eradicate it one day.

To date, the results have led to 2 patent request, 4 publications in international peer-reviewed journals and have been mentioned in The Conversation and National Geographic. Together, they have refined our knowledge of the pathophysiological mechanisms of infection, the environment of the insect digestive tract faced by Y. pestis and the molecular mechanisms used by the bacterium to produce a flea-borne infection.

Flea-borne plague (caused by Yersinia pestis) is an international, re-emerging public health concern and a serious bioterrorism threat. This disease has the ability to rapidly spread, to cause a high mortality rate and huge socioeconomic disruption. The plague threat is accentuated by (i) the emergence of Y. pestis strains resistant to all the antibiotics recommended by WHO for prophylactic and curative treatments, and (ii) the ease with which strains can be engineered to resist all antibiotics. This concern is further aggravated by the absence of a plague vaccine. At present, there are no clear estimations of the consequences of a potential return of plague in Europe (whether natural, accidental or intentional). It is not well established whether this type of re-introduction can be sustained by Europe’s endemic fleas under current and future climate conditions. There is also lack of knowledge on the mechanism of plague transmission, which will hamper future efforts to control the disease. Hence, determining the environmental variables and characterizing the molecular mechanisms leading to successful transmission of Y. pestis by the flea are key steps towards better prevention and protection of our citizens against plague. However, today's investigational methods are not appropriate for rapidly achieving these goals. Here, we propose a fundamental research project that aims at (i) better assessing the risk and potential impacts of plague in Europe, and (ii) reducing and then eliminating the threat. To this end, we intend to develop and apply innovative high-content screening techniques combined with relevant standardized protocols for (i) providing quick, accurate measurements of vector efficiency and capacity of endemic European flea species under various climate conditions and (ii) allowing the rapid identification of genes that are important for flea-borne transmission of plague. The latter will serve as the starting point for more in-depth molecular studies of the flea-borne transmission in plague. From a technical point of view (short-term goals): we anticipate that the novel methodologies and approaches developed within the project will raise our discovery rate to an unprecedented level and could be transposed to other arthropod-borne diseases. From a scientific point of view (short-term goals), we expect to impact the plague community and identify novel transmission mechanisms used by other flea-borne pathogens (including other class A agents). From an economic/political point of view (medium-/long-term goals), better defining the climate variables required for the transmission of plague (using local and foreign flea species) will help to better assess the risk and putative impact of the spread of flea-borne plague (after an attack or natural introduction). By evaluating the sanitary risk and the warning threshold associated with various flea species and climatic variables, the project may be able to provide the authorities with a means of predicting and preventing plague dissemination and epidemics in European populations. Characterizing the mechanisms of flea-borne plague may also provide novel strategies for blocking the spread of the disease. The project may help to limit the number of endemic regions, reduce the ability of criminals to acquire and spread the bacterium (protection at the source), reduce the risk of transmission in (naturally or criminally) infected areas, and thus protect our citizens. Hence, the project’s benefits for society are expected to go well beyond better knowledge of plague.