The recent emergence of amphibian chytridiomycosis has precipitated competing hypotheses regarding the endemic versus novel nature of the causative agent, Batrachochytrium dendrobatidis (Bd ). We conducted a retrospective survey of the California Academy of Sciences' (San Francisco, California, USA) amphibian collection, testing for presence of Bd in 4 amphibian species collected from central California between 1897 and 2005. The earliest detection of Bd was found in 2 Rana catesbeiana in 1961, and the data support the hypothesis that Bd was a novel pathogen introduced into central California prior to 1961 that spread out geographically and taxonomically from at least one central location and is now endemic throughout most of central California. The taxonomic pattern of infection prevalence and the ecological constraints of the 4 species we tested suggest that, although Bd was initially detected in R. catesbeiana, the more efficient and most likely local vector for Bd in central California is actually Pseudacris regilla.KEY WORDS: Chytridiomycosis · Batrachochytrium dendrobatidis · Rana draytonii · Natural history collections · Pseudacris regilla · R. catesbeiana · R. boylii · Co-kriging Resale or republication not permitted without written consent of the publisherDis Aquat Org 83: [1][2][3][4][5][6][7][8][9] 2009 Despite these disadvantages, natural history collections nonetheless present our only opportunity for retrospective surveys when seeking answers to questions within an historical context.Examination of current and archived specimens has documented Bd on all continents that harbor amphibians (Bosch et al. 2001, Weldon 2002, Weldon et al. 2004, Une et al. 2008. Weldon et al. (2004) examined 697 archived amphibians collected in South Africa from 1879 to 1999 and detected the earliest known Bd infection in Xenopus laevis collected in 1938. They found that the pathogen was widespread in Africa and concluded that Bd was a stable endemic infection in southern Africa for the 23 yr period before Bd was first detected on another continent. Their museum survey results formed the basis for the hypothesis that Bd originated in Africa and supported the proposal that X. laevis was most likely the original global vector of the pathogen. X. laevis was globally exported out of Africa beginning in 1935, when it became the test species used for human pregnancy assays (Tinsley & McCoid 1996). Rana catesbeiana has also been postulated as a global vector of Bd (Daszak et al. 2004, Garner et al. 2006, Fisher & Garner 2007, as both R. catesbeiana and X. laevis are relatively asymptomatic when infected (Parker et al. 2002, Daszak et al. 2004) and both species are globally transported for the pet, food and laboratory trades (Tinsley & McCoid 1996, Cunningham et al. 2003, Daszak et al. 2004, Fisher & Garner 2007.The earliest known Bd-positive specimens outside of Africa were 2 Rana clamitans collected in July 1961 in Quebec, Canada, as documented by Ouellet et al. (2005), who surveyed amphibian collections from 2 Can...

Amphibian population declines in Honduras have long been attributed to habitat degradation and pollution, but an increasing number of declines are now being observed from within the boundaries of national parks in pristine montane environments. The amphibian chytrid fungus Batrachochytrium dendrobatidis has been implicated in these declines and was recently docu-

Controlled exposure experiments can be very informative, however, they are based on the assumption that pathogens maintained on artificial media under long-term storage retain the infective and pathogenic properties of the reproducing pathogen as it occurs in a host. We observed that JEL284, an in vitro cultured and maintained isolate of Batrachochytrium dendrobatidis (Bd), was becoming less infectious with successive uses. We hypothesized that passing an isolate propagated on artificial media through an amphibian host would make the isolate more infectious and pathogenic in subsequent exposures. To test our hypothesis, we used two discreet steps, a reisolation step (step 1) and a comparative exposure step (step 2). In step 1, we exposed eastern spadefoot toads, Scaphiopus holbrooki, to JEL284 and JEL197, another isolate that had been maintained in vitro for over six years. We then re-isolated JEL284 only from a successful infection and named this new isolate JEL284FMBa. JEL197 did not infect any amphibians and, thus, did not proceed to step 2. In step 2, we compared infectivity and pathogenicity (mortality and survival time) of JEL284 and JEL284FMBa by exposing 54 naïve S. holbrooki to three treatments (JEL284, JEL284FMBa, and negative control) with 18 individuals per group. We found that JEL284FMBa caused higher mortality and decreased survival time in infected individuals when compared to JEL284 and negative controls. Thus, our data show that pathogenicity of Bd can decrease when cultured successively in media only and can be partially restored by passage through an amphibian host. Therefore, we have demonstrated that pathogenicity shifts can occur rapidly in this pathogen. Given the potential for shifts in pathogenicity demonstrated here, we suspect Bd to have similar potential in natural populations. We suggest that, when possible, the use of freshly isolated or cryopreserved Bd would improve the quality of controlled exposure experiments using this pathogen.

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ABSTRACT:The objective of this study was to evaluate the utility of gross morphologic examination of larval mouthpart defects as a diagnostic screening test to detect Batrachochytrium dendrobatidis infection in four California, USA, anuran species. We examined mouthparts of 2,034 tadpoles of Bufo boreas, Pseudacris regilla, and Rana catesbeiana collected in 2003 and 2004 and Bufo canorus collected in 2004. Data were recorded for three morphologic features: upper toothrows, lower toothrows, and combined jaw sheaths. Mouthpart defects were observed in all four species (n5757), but only two species were infected with B. dendrobatidis (n584). Sensitivity and specificity of the mouthparts test were 76% and 58%, respectively. Forty-two percent of B. dendrobatidis-negative animals would have been designated positive based on mouthpart defects. Observed prevalence was 43%, and true prevalence was 3.0%. Tests of the null hypothesis using logistic regression analysis showed that anuran larval mouthpart defects were not associated with B. dendrobatidis infection whether mouthparts scores were tested by individual morphologic feature or in combination (P50.37). We conclude that B. dendrobatidis infection and anuran larval mouthpart defects are two separate processes that may occur concurrently and that evaluation of tadpole oral morphology is neither an accurate nor a reliable diagnostic test for B. dendrobatidis infection for the four species tested.