A Rapid, in-Situ Minimally-Invasive Technique to Assess Infections with Pseudogymnoascus destructans in Bats

Emerging infectious diseases may become serious threats to wildlife, a prominent example being the white-nose disease (WND). In case of WND, the cold-loving fungus Pseudogymnoascus destructans colonizes bats during hibernation, invades the skin and has already lead to the death of millions of bats in North America. P. destructans most likely originated from Europe, where it also causes lesions but without associated mortalities. However, it is still unclear how European bats cope with the fungus. Hence, it is important to have tools that precisely characterise disease progression. Because hibernation is a physiological state during which bats are vulnerable to disturbance, in-situ assessments of the clinical status should be carried out minimal-invasively to avoid detrimental impacts on bats. However, currently available disease assessment methods require handling/touching bats and are therefore invasive: i) UV-light trans-illumination of wing membranes to detect lesions and ii) a qPCR-based quantification of fungal material from wing membrane swabs. Since P. destructans (‘Pd') becomes visible on all furless skin with distinct distribution patterns, we investigated the use of visible symptoms to assess levels of infections without handling/touching bats. We introduce a technique which we termed ‘Visual Pd-score’ (a visual classification scheme), which can be applied without disturbing the animals. To assess its reliability, we used P. destructans infected greater mouse-eared bats (Myotis myotis) to compare the novel method with the two existing golden-standard techniques. Our results show that infection levels obtained from all three techniques are correlated. Importantly, the information carried by the Visual Pd-score is most similar to a composite index combining the information from the qPCR-based and UV-light quantification methods. We conclude that the Visual Pd-score represents a promising index to better characterise disease severity as it is simultaneously representative for fungal colonization and wing damage. We discuss advantages and disadvantages of the applied techniques and conclude that the Visual Pd-score is particularly useful for routine hibernacula counts or large-scale P. destructans-surveillance. In combination with the lesion detection technique the new method is also applicable to immunological studies where both fungal colonization and associated damage have to be investigated, while qPCRs from swabs of all body parts are especially useful if it is necessary to detect cryptic infections, e.g. during the early hibernation period.