Environmental DNA: A promising factor for tuberculosis risk assessment in multi-host settings.

Attaining and maintaining the Official Tuberculosis Free status continues to be a challenge when several domestic and wild hosts contribute to the maintenance of the Mycobacterium tuberculosis complex (MTC). Local tuberculosis hotspots are sometimes identified in cattle in low-prevalence regions. We have, therefore, studied one such hotspot in depth in order to produce an epidemiological diagnosis. Host population size and MTC prevalence were estimated in selected wildlife and in livestock, while on-cattle environmental DNA detection was additionally used as a proxy for risk of exposure at the farm (herd) level. Positive skin test reactors were found on16 of the 24 cattle farms studied in the period 2012-2016. Although all goats tested negative to the skin test during this period, MTC was confirmed in four sheep at slaughter, thus indicating an unknown prevalence of infection in this host species. With regard to wildlife, the prevalence of MTC infection based on culture was 8.8% in the case of wild boar (Sus scrofa), and the only road-killed badger (Meles meles) submitted for culture tested positive. Two criteria were employed to divide the cattle farms into higher or lower risk: tuberculosis testing results and environmental DNA detection. Environmental MTC DNA detection yielded significant differences regarding "use of regional pastures" and "proximity to woodland". This study suggests that on-animal environmental DNA sampling may help when assessing contact risk as regards MTC in livestock at the herd level. This tool opens up new avenues of epidemiological research in complex multi-host settings.This work was supported by the Programa de Tecnologías Avanzadas en Vigilancia Sanitaria (TAVS) from the Comunidad de Madrid (ref. S2013/ABI-2747). JMG holds a FPI pre-doctoral scholarship (BES-2015- 072206), funded by MINECO. This is a contribution to MINECO grant CGL2017-89866 WildDriver and EU-FEDER. This is also a contribution to Valle de Alcudia pilot project SG-2019-02 from PDR-CLM, and to GOSTU project EU-FEADER (AEIAGRI-PNDR-MAPA, ref. 20190020007521). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.S