Nonlethal Screening of Bat-Wing Skin With the Use of Ultraviolet Fluorescence to Detect Lesions Indicative of White-Nose Syndrome

Abstract

Definitive diagnosis of the bat disease white-nose syndrome (WNS) requires histologic analysis to identify the cutaneous erosions caused by the fungal pathogen Pseudogymnoascus [formerly Geomyces] destructans (Pd). Gross visual inspection does not distinguish bats with or without WNS, and no nonlethal, on-site, preliminary screening methods are available for WNS in bats. We demonstrate that long-wave ultraviolet (UV) light (wavelength 366-385 nm) elicits a distinct orange-yellow fluorescence in bat-wing membranes (skin) that corresponds directly with the fungal cupping erosions in histologic sections of skin that are the current gold standard for diagnosis of WNS. Between March 2009 and April 2012, wing membranes from 168 North American bat carcasses submitted to the US Geological Survey National Wildlife Health Center were examined with the use of both UV light and histology. Comparison of these techniques showed that 98.8% of the bats with foci of orange-yellow wing fluorescence (n=80) were WNS-positive based on histologic diagnosis; bat wings that did not fluoresce under UV light (n=88) were all histologically negative for WNS lesions. Punch biopsy samples as small as 3 mm taken from areas of wing with UV fluorescence were effective for identifying lesions diagnostic for WNS by histopathology. In a nonlethal biopsy-based study of 62 bats sampled (4-mm diameter) in hibernacula of the Czech Republic during 2012, 95.5% of fluorescent (n=22) and 100% of nonfluorescent (n=40) wing samples were confirmed by histopathology to be WNS positive and negative, respectively. This evidence supports use of long-wave UV light as a nonlethal and field-applicable method to screen bats for lesions indicative of WNS. Further, UV fluorescence can be used to guide targeted, nonlethal biopsy sampling for follow-up molecular testing, fungal culture analysis, and histologic confirmation of WNS.

Detection ofPseudogymnoascus destructanson Free-flying Male Bats Captured During Summer in the Southeastern USA

“…The pattern of fluorescence occasionally seen on bats during summer was not associated with WNS and did not resemble the widespread, miliary pattern typically seen on wing membranes of bats with WNS during winter (Turner et al 2014). Presence of Pd on bats during summer likely represented superficial colonization, which is not identifiable by examination under UV light.…”

“…Basic demographic and morphometric data were recorded from sampled bats using standard techniques (see Supplementary Material Table S1). Wing membranes were assessed for damage (Reichard and Kunz 2009) and inspected under ultraviolet (UV) light (51 bulb LED UV [385 nm] flashlight with filter, LED Wholesalers, Hayward, California, USA) for fluorescence suggestive of WNS (Turner et al 2014). Nonthreatened or nonendangered species of bats with suspicious fluorescence were euthanized and stored at À20 C for diagnostic evaluation; nonlethal wing biopsy samples (3 mm) of similarly fluorescing areas of skin from threatened or endangered species were collected into 10% neutral buffered formalin and stored at room temperature for histopathologic evaluation.…”

Host persistence or extinction from emerging infectious disease: insights from white-nose syndrome in endemic and invading regions

“…We also surveyed a subset of bats' wing and tail membranes for orange fluorescence using an ultraviolet (UV) flashlight (395 nm; Hayward, CA, USA). The presence of orange fluorescence on wing and tail membranes under UV light has been shown to be correlated with the presence of lesions in infected bats [35].…”

Quantification of pathogen levels is necessary to compare responses to pathogen exposure: Comment on Davy et al. “The other white‐nose syndrome transcriptome”

“…In an unpublished study, I have examined whether gene expression patterns vary between adjacent tissues that are uninfected or infected with P. destructans . UV fluorescence (Turner et al., 2014) was used to identify P. destructans ‐positive and  P. destructans ‐negative sites in M. lucifugus wing tissue from bats infected with P. destructans in captivity. The 12 UV‐negative biopsies had low levels of P. destructans reads in the RNA‐Seq data (0.13% ± 0.15% of mapped reads) while 10 of the 12 the UV‐positive biopsies had higher levels of P. destructans reads (4.17% ± 3.27% of mapped reads).…”