We document white-nose syndrome (WNS), a lethal disease of bats caused by the fungus Pseudogymnoascus destructans (Pd), and hibernacula microclimate in New Brunswick, Canada. Our study area represents a more northern region than is common for hibernacula microclimate investigations, providing insight as to how WNS may impact bats at higher latitudes. Hibernacula microclimate is important in hibernaculum selection by bats and may also be an important factor influencing WNS-associated bat mortality rates. To determine the impact of the March 2011 arrival of Pd in New Brunswick and the role of hibernacula microclimate on hibernating bat mortality, we surveyed bat numbers at hibernacula twice a year from 2009 – 2015. We also collected data from iButton temperature loggers deployed at all sites and gathered temperature and humidity data from HOBO loggers at three sites. Bat species found in New Brunswick hibernacula include Myotis lucifugus (Little Brown Bat) and M. septentrionalis (Northern Long-eared Bat), with small numbers of Perimyotis subflavus (Tricolored Bat). All known hibernacula in the province were Pd-positive with WNS-positive bats by winter 2013. A 99% decrease in the over-wintering bat population in New Brunswick was observed between 2011 and 2015. We did not observe P. subflavus during surveys 2013 – 2015 and the species appears to be extirpated from these sites. Bats did not appear to choose hibernacula based on winter temperatures, but dark zone (zone where no light penetrates) winter temperatures did not differ among our study sites, likely due to the geographically small sampling area. Dark zone temperatures did not vary among years, unlike hibernacula entrance temperatures, but both locations were affected by the presence of flowing water. Winter dark zone temperatures were warmer and less variable than entrance or above ground temperatures. We observed visible Pd growth on hibernating bats in New Brunswick during early winter surveys (November), even though hibernacula temperatures were colder (~ 4 – 5°C) than optimum for in vitro Pd growth (12.5 – 15.8°C). This suggests that cold hibernacula temperatures encountered near the apparent northern range limit for Pd do not sufficiently slow fungal growth to prevent the onset of WNS and associated bat mortality over the winter. As WNS continues to spread west across continental North America, the severity of WNS-related mortality may therefore be greater at northern latitudes where bat hibernation periods are longer, despite apparent suboptimal temperatures for Pd growth.

We monitored six limestone caves, two gypsum caves, and three abandoned manganese mines in southern New Brunswick, Canada, 2009 ‒ 2015 (site map in Vanderwolf et al. 2012). We placed air temperature logger iButtons (model DS1921G, ± 1˚C, Maxim Integrated Products, Inc., Sunnyvale, California) in caves and mines in December 2011 and retrieved them August 2017. We set iButtons to record air temperature twice daily (0230 and 1430 hrs.) to capture daily temperature extremes and increase longevity of the devices. We deployed three iButtons at each site: we placed one 1 – 2 m from the ground on a wall ledge near the ceiling in the twilight zone (i.e. the entrance area where some light penetrates 7 ± 5 m into the cave), a second under similar circumstances in the dark zone (zone where no light penetrates), and attached a third to a tree at chest height 50 – 200 m from each hibernaculum entrance. We placed iButtons in the dark zone in passage or chamber areas where bats roosted (if bats were present) 45 ± 26 m SD (range 16 – 200 m) from the entrance, with placement depending on hibernaculum length and roosting location of hibernating bats. Dark zone iButtons were placed in areas with the largest concentration of bats, which was generally not the deepest part of each hibernaculum (total lengths ranged from 74 – 515 m). To avoid loss inside caves, we placed iButtons in perforated (to facilitate airflow) translucent plastic boxes. iButtons outside caves were placed in plastic camouflaged boxes (painted brown to deter theft) with a hole drilled in the bottom to allow water drainage. Due to iButton failures, we placed two iButtons in each box starting in 2013 (paired iButtons). When available we used the mean value of paired iButtons for analysis.

We deployed temperature/relative humidity loggers (HOBO model U23-001; ± 0.45˚C, ± 2.5% from 10% to 90% RH and ± 5% at >90% RH, Onset Computer Corporation, Bourne, MA) at three sites in conjunction with iButtons: White Cave, Markhamville Mine, and Berryton Cave. We deployed HOBOs April 2014 and retrieved them August 2017. We placed three HOBOs at each site near existing iButtons (twilight zone/entrance, dark zone, and outside the hibernaculum) and programmed them to record air temperature and relative humidity twice daily (0230 and 1430 hrs.). HOBOs outside hibernacula were tied at chest height to the same tree as camouflaged iButton boxes, with the HOBO placed just above the iButtons and the sensor pointing towards the ground. We wrapped HOBOs in a tube of black foam pipe insulation for protection and camouflage, while ensuring that the sensor was open to the air at the bottom of the tube. Loggers tied to trees were below the forest canopy, which probably modulated local extremes in temperature and humidity. We downloaded data from iButtons and HOBOs twice a year during bat counts and replaced malfunctioning iButtons. The relative humidity sensor of the HOBO logger deployed in the dark zone of White Cave failed September 2016, so no data were available after that date.

During visits March 2012 ‒ May 2015 we also measured air temperature and relative humidity with a Kestrel 3000 Handheld weather meter (MPN# 0830, ± 0.04˚C, ± 1% RH, Boothwyn, Pennsylvania, USA). Kestrel measurements took some time to stabilize, particularly for relative humidity, so the device was temporarily attached to a tree adjacent to the hibernaculum entrance for measurements above ground, and positioned on the hibernacula floor for measurements inside passages while we performed bat counts.

We converted relative humidity to water vapor pressure because relative humidity is less informative and potentially misleading (Kurta, 2014). We calculated equilibrium vapor pressure using temperature data and the quadratic formula of Tabata (1973), and then determined actual vapor pressure (hPa) by multiplying the saturation vapor pressure by the relative humidity as recorded by HOBOs. We incorporated previous iButton temperature data collected from the same sites October 2009 to October 2010 (Vanderwolf et al., 2012) in the analysis.

Kurta, A. (2014) ‘The misuse of relative humidity in ecological studies of hibernating bats’, Acta Chiropterologica, 16(1), pp. 249–254. doi: 10.3161/150811014x683444.

Tabata, S. (1973) ‘A simple but accurate formula for the saturation vapor pressure over liquid water’, Journal of Applied Meterorology, 12, pp. 1410–1411.

Vanderwolf, K. J. et al. (2012) ‘Bat populations and cave microclimate prior to and at the outbreak of white-nose syndrome in New Brunswick’, Canadian Field-Naturalist, 126(2), pp. 125–134.

There are temporal gaps in the data due to logger failure but these do not show up as missing values. The 'iButton data' tab also include temperature data from HOBO loggers at the bottom. The 'Kestral data' tab has multiple missing values indicated as a blank cell. Any reference to 'Mark small' refers to Markhamville Mine 2. Water temperature data was collected during a previous study and collection methods are described there (Vanderwolf et al. 2017).

Vanderwolf, K. J., McAlpine, D. F. and McGuire, L. P. (2017) ‘Hibernacula water chemistry and implications for hibernating bats’, Journal of Mammalogy, 98(6). doi: 10.1093/jmammal/gyx111.