In recent years Batrachochytrium dendrobatidis (Bd), a chytridiomycete fungus pathogenic to amphibians (Longcore and others 1999), has been implicated as the proximate cause of amphibian declines around the world (Berger and others 1998; Daszak and others 2003; Muths and others 2003; Pounds and others 2006). Despite the insidious nature of Bd (Green and others 2002), few published data exist addressing its occurrence in the Pacific Northwest (PNW). When Pearl and others (2007) opportunistically examined 7 PNW amphibian species, they found Bd most often (57% of 21 individuals from 14 sampled populations) in the highly aquatic Oregon Spotted Frog (Rana pretiosa), an Endangered Species in Washington State (WDFW 2009). However, all R. pretiosa they sampled were from Oregon, and Bd was not detected in the Rana cascadae (Cascades Frog) and Anaxyrus boreas (Western Toad) specimens they sampled from Washington. Here, we report the detection of Bd in R. pretiosa from Washington. Since 1997 and 1998, respectively, we have been monitoring R. pretiosa populations at the Trout Lake Natural Area Preserve (TLNAP: UTM Zone 10, 610857-612950E, 5095880-5097574N, WGS84; elev. 594 to 599 m) and Conboy Lake National Wildlife Refuge (CLNWR: UTM Zone 10, 625223-635180E, 5086652-5095491N, WGS84; elev. 552 to 576 m), in Klickitat County, Washington. These sites represent 2 of only 3 areas where R. pretiosa is known to occur in Washington (McAllister and Leonard 1997). At both sites, monitoring included egg mass surveys during the annual breeding season following snowmelt (late February to mid-March at CLNWR; midMarch to early April at TLNAP). Surveys involved area-specific counts of individual egg masses and egg mass groups, each of which was marked with flags and geo-referenced using a Geographic Positioning System (GPS) to avoid double-counting. Sampling at CLNWR addressed 4 hydrologically distinct units, which were surveyed repeatedly until no new egg masses were found; at TLNAP, 3 units consistently used for breeding were sampled. We inferred a 1:1 correspondence between egg mass numbers and number of breeding females based on a combination of our direct observations of oviposition (n 5 13), the recapture of non-gravid females known to have laid eggs (n 5 84), and the relatively short interval over which the laying of new egg masses occurs (about 3 wk). Collectively, these data indicate that females lay only 1 clutch annually, and that egg mass numbers reflect the effective population numbers of adult females. Decline in egg mass numbers in 3 of the 4 surveyed units at CLNWR from 2004 to 2005 (Fig. 1) and 2 of the 3 units at TLNAP over the same period (Fig. 2), coupled with the increasing recognition of Bd as a cause of amphibian declines, motivated us to collect dead frogs found during 2006 and test them for Bd. Five dead adult R. pretiosa were tested for Bd; 1 from CLNWR and 4 from TLNAP. The CLNWR specimen collected on 13 March 2006 displayed feeble vital signs, minimal response to touch, and righting response was lacking. The frog died within 20 min of discovery, was preserved in 10% formalin, and then stored in 70% ethanol before histological examination using a standard wet-mount preparation of its epidermal tissue (Berger and others 1999), which was sloughing extensively as multiple epidermal layers. A dead adult female R. GENERAL NOTES
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