Vol. 92: 187–191, 2010 DISEASES OF AQUATIC ORGANISMS Published online September 23, 2010 doi: 10.3354/dao02250 Dis Aquat Org Contribution to DAO Special 4 ‘Chytridiomycosis: an emerging disease’ NOTE Batrachochytrium dendrobatidis in the live frog trade of Telmatobius (Anura: Ceratophryidae) in the tropical Andes Alessandro Catenazzi1,*, Vance T. Vredenburg2, Edgar Lehr3 1Department of Integrative Biology, University of California at Berkeley, 3060 Valley Life Sciences, Berkeley, California 94707, USA 2Department of Biology, San Francisco State University, San Francisco, California 94132, USA 3Department of Biology, Illinois Wesleyan University, 303 East Emerson, Bloomington, Illinois 61701, USA ABSTRACT: Species of frogs in the genus Telmatobius are traded and sold for human consumption in the Andes and in coastal cities of Peru and Bolivia. These frogs are harvested from wild popula- tions. We report high prevalence of infection by the pathogenic fungus Batrachochytrium dendroba- tidis in live frogs purchased at the main market in Cusco, Peru, from January 2008 to January 2010. We suggest that the transport of native anurans through the live frog trade could facilitate the spread of this fungus among Andean frogs. The tropical Andes are the most important biodiversity hotspot for amphibians. Because many neotropical taxa are known to be susceptible to chytridiomycosis, the presence of a large reservoir of infection in the frog trade poses a significant threat to amphibian conservation. KEY WORDS: Batrachochytrium dendrobatidis · Amphibians · Trade · Andes · Peru · High elevation Resale or republication not permitted without written consent of the publisher INTRODUCTION peared; Lips et al. 2006). Little is known about how Bd is spreading, but Lips et al. (2008) proposed that the The emerging infectious disease chytridiomycosis, temporal pattern of these declines reflects the epi- caused by the fungus Batrachochytrium dendrobatidis demic spread of Bd from multiple sites of introduction (Bd), has been linked to amphibian declines and ex- in Central and South America. According to this hypo- tinctions worldwide (Berger et al. 1998, Longcore et al. thesis, Bd began infecting amphibians in the Andes of 1999, Lips et al. 2006). This fungus infects the skin of Peru and Bolivia around the year 2000, causing local amphibians, causing osmotic imbalance leading to car- and regional species extirpations. In support of this diac arrest (Voyles et al. 2009). Amphibian species dif- hypothesis, Bd has recently been discovered in this fer in their vulnerability and resistance to chytridiomy- region in a number of amphibian species (Seimon et al. cosis (Alford & Richards 1999, Daszak et al. 2003, 2007, Barrionuevo et al. 2008, Venegas et al. 2008). Woodhams et al. 2007), but this disease often causes Moreover, Catenazzi et al. (in press) reported Bd in 16 catastrophic and quick declines of species-rich amphi- frog species surveyed between 2008 and 2010 in bian faunas (Lips et al. 2006). southeastern Peru. Neotropical montane amphibians have declined or The mechanism for the introduction and spread of collapsed over the past 3 decades (Lips 1998, Busta- Bd is still unknown, but one theory that has been pro- mante et al. 2005, La Marca et al. 2005, Catenazzi et al. posed is that Bd is spread by live trade in amphibians in press). In at least one case in Central America, Bd for human consumption (Daszak et al. 2003, Weldon et caused the collapse of a highly diverse amphibian al. 2004, Schloegel et al. 2009). Most of the attention fauna on a single mountain (22 of 46 species disap- has so far been focused on farming and trade in Litho- *Email: [email protected] © Inter-Research 2010 · www.int-res.com 188 Dis Aquat Org 92: 187–191, 2010 bates catesbeianus (Ranidae; Daszak et al. 2004, Gar- MATERIALS AND METHODS ner et al. 2006). This species is known to carry Bd, yet it is not known to die from chytridiomycosis, and is We visited the San Pedro market in Cusco, Peru raised in farms around the world, including in South (13° 31’ 16” S, 71° 58’ 57” W, 3410 m above sea level) America (Mazzoni et al. 2003, Hanselmann et al. 2004, on 30 January 2008, 11 October 2008, 16 February FAO 2009: www.fao.org/fishery/topic/2017/en). In this 2009, and 3 January 2010. During each visit, we pur- paper we investigate whether trade in native Andean chased 5 live adult Telmatobius marmoratus (the only species (captured in the wild, but traded alive) could species visibly on sale at the market during our visits) be a source of Bd for wild populations. from a stand at the Cusco central market (there was Andean frogs in the genera Telmatobius and Batra- only one stand operating during the course of our chophrynus are traded in montane and coastal cities study). In addition, in 2009, the vendor donated skins for food and because of presumed magical or medici- of 9 recently skinned adults, which we also utilized in nal properties (Lehr 2000, Angulo 2008a). The frogs this study. are sold live, freshly skinned and degutted, dried, or Live frogs (~20 to 30 ind. per container) were housed prepared in frog ‘shakes,’ cooked in soups, or fried in 10 l plastic buckets filled with water (Fig. 1). The (Angulo 2008a). Presumed medicinal properties of vendor was beheading, skinning, and gutting frogs for frogs include treatment of neurological disorders or customers during visits; recently deceased, skinned symptoms, anemia, prostate problems, menopausal frogs ready for consumption were also available at the disorders, and asthma in Cusco (Angulo 2008a) and stand. We recorded price, snout–vent length (SVL), treatment of respiratory diseases, arthritis, diabetes, body mass, and sex of each frog. We housed live frogs frigidity, and hair loss, as well as aphrodisiac action, in individually in plastic bags and swabbed them shortly Lima (Lehr 2000). after purchase. We used frog skin swabs and a real- Peru and Bolivia harbor the highest diversity of Tel- time PCR assay to quantify Bd prevalence and infec- matobius species (Lehr 2005). These frogs are threat- tion intensity (Boyle et al. 2004). Swabs (MW113, Med- ened by habitat destruction and modification, con- ical Wire and Equipment) were stroked across a frog’s tamination from mining and agricultural practices, skin in a standardized way: 5 strokes on each side of harvesting, introduced fish, and disease (Angulo the abdominal midline, 5 strokes on the inner thigh of 2008b, De la Riva & Lavilla 2008, von May et al. 2008). each hind leg, and 5 strokes on the foot webbing of The 3 Telmatobius spp. known from Ecuador were ex- each hind leg. Swabs were air dried in the field and tirpated in the late 1980s and early 1990s and are now stored individually in labeled microcentrifuge tubes presumed to be extinct (Merino-Viteri et al. 2005). before PCR analysis. We used standard DNA extrac- Symptoms of chytridiomycosis were observed in the tion and real-time PCR methods (Boyle et al. 2004, last Telmatobius specimens collected in this region Hyatt et al. 2007), except that swab extracts were ana- (Merino-Viteri et al. 2005). Chytridiomycosis has since lyzed in singlicate instead of triplicate (Kriger et al. been implicated in population declines of T. marmoratus from Peru (Seimon et al. 2007) and 2 Telmatobius spp. from Argentina (Barrionuevo & Mangione 2006). We propose that the trade of live Tel- matobius frogs could facilitate the spread of Bd among wild populations in the tropical Andes. Wild Telmatobius frogs are harvested from lakes and streams and transported live to markets located up to hundreds of km away from their collecting sites. T. culeus, for example, is transported over several hundred km from Lake Titicaca to the main city mar- ket in Cusco in southern Peru (Angulo 2008a) and all the way to Lima (Lehr 2000). Here we report data on the preva- lence of infection and zoospore load of Bd in T. marmoratus frogs sold live for Fig. 1. Stand offering skinned and degutted and live Telmatobius marmoratus human consumption in Cusco. at the San Pedro market in Cusco, Peru, on 3 January 2010 Catenazzi et al.: Bd infection in live frog trade 189 2006). We defined infection intensity as the number of imens were reproductive males bearing nuptial pads ‘zoospore equivalents’ per swab (Zswab). Zoospore and excrescences, except for a female purchased in equivalents were calculated by multiplying the February 2009 and 3 females purchased in January genomic equivalent values generated during the real- 2010. According to the vendor, specimens in October time PCR by 80; this multiplication accounts for the fact 2008 and February 2009 originated from Ocongate, that DNA extracts from swabs were diluted 80-fold 65 km east of Cusco. during extraction and PCR. For calculations of Bd prevalence, swabs were categorized as positive when zoospore equivalents were ≥1 and Bd-negative when DISCUSSION AND CONCLUSIONS zoospore equivalents were <1. Frogs were euthanized with a ventral cutaneous application of 20% benzo- All live Telmatobius marmoratus sampled during caine gel, fixed in formalin, and placed in 70% this study were infected with Bd. This is the first report ethanol. Specimens have been deposited at the Museo of captive T. marmoratus harboring infections in the de Historia Natural, Universidad Nacional Mayor de trade of this species for human consumption. The high San Marcos, Lima. prevalence in our sampled animals might be explained by the fact that Bd has been known to infect wild pop- ulations of this species in the region since at least 2002 RESULTS (Seimon et al. 2007). Additionally, the high prevalence could also be the result of frogs being housed in close All frogs (100%) swabbed between January 2008 contact with each other (e.g. in plastic buckets filled and January 2010 were infected with Bd (n = 20; with water at the market; Fig.