Molecular evidence of Francisella tularensis in nature, 10 years after a large outbreak of tularemia in Spain

Resumen del trabajo presentado a la 9th International Conference on Tularemia, celebrada en Montréal (Canada) del 16 al 19 de octubre de 2018.[Background and aim]: Tularemia is a zoonosis caused by Francisella tularensis, a pathogen that can use a variety of transmission routes in nature as well as in humans. Since 1997 more than 1,000 human cases of tularemia have been declared in Spain in two major outbreaks, but knowledge about the epidemiology of this infection is very limited. The ability of F. tularensis to find mechanisms to persist in the environment and cause new outbreaks makes it necessary to identify, among others, the potential reservoirs during inter-epizootic periods, the main objective of this study. [Material and methods]: The sampling points were located in an area of Castilla y León, well known as “hot spots” for tularemia. They corresponded to surface water and sediments, as well as crayfish, mosquitoes, hares and ticks. Specifically, 8 zones divided into 4 subzones were sampled, including three rivers, three irrigation channels and three lagunes. Samples were collected in July and November of 2016 and 2017. For detection of Francisella lpnA was amplified by conventional PCR and hybridization by reverse line blotting with specific probes. Positive samples were confirmed by a real-time multitarget TaqMan PCR, using tul4 and ISFtu2 assays. For additional characterization sdhA gen and VNTR Ft-M19 were amplified and sequenced. [Results]: A total of 102 samples of water, 111 sediments, 343 crayfish, 156 mosquitos, 64 hares and 126 ticks from 14 hares were collected. Overall, 2 samples of water, 3 of sediments, 7 hares and 17 ticks tested positive for Francisella. Interestingly, F. tularensis subspecie holarctica was detected in water, hares and ticks while F. hispaniensis was the species detected in water and sediments. [Conclusions]: In this study we provide evidence of Francisella presence in nature during periods in which no large outbreaks of tularemia occur, and suggest that hares and ticks play an important role in the maintenance of the bacteria in the terrestrial environment. Moreover, to the best of our knowledge this is the first description of F. hispaniensis in abiotic environment, which enforces the need to assess the risk for the population and establish the necessary measures for its prevention and control.Peer Reviewe