diversity Review Effects of Emerging Infectious Diseases on Amphibians: A Review of Experimental Studies Andrew R. Blaustein 1,*, Jenny Urbina 2 ID , Paul W. Snyder 1, Emily Reynolds 2 ID , Trang Dang 1 ID , Jason T. Hoverman 3 ID , Barbara Han 4 ID , Deanna H. Olson 5 ID , Catherine Searle 6 ID and Natalie M. Hambalek 1 1 Department of Integrative Biology, Oregon State University, Corvallis, OR 97331, USA; [email protected] (P.W.S.); [email protected] (T.D.); [email protected] (N.M.H.) 2 Environmental Sciences Graduate Program, Oregon State University, Corvallis, OR 97331, USA; [email protected] (J.U.); [email protected] (E.R.) 3 Department of Forestry and Natural Resources, Purdue University, West Lafayette, IN 47907, USA; [email protected] 4 Cary Institute of Ecosystem Studies, Millbrook, New York, NY 12545, USA; [email protected] 5 US Forest Service, Pacific Northwest Research Station, Corvallis, OR 97331, USA; [email protected] 6 Department of Biological Sciences, Purdue University, West Lafayette, IN 47907, USA; [email protected] * Correspondence [email protected]; Tel.: +1-541-737-5356 Received: 25 May 2018; Accepted: 27 July 2018; Published: 4 August 2018 Abstract: Numerous factors are contributing to the loss of biodiversity. These include complex effects of multiple abiotic and biotic stressors that may drive population losses. These losses are especially illustrated by amphibians, whose populations are declining worldwide. The causes of amphibian population declines are multifaceted and context-dependent. One major factor affecting amphibian populations is emerging infectious disease. Several pathogens and their associated diseases are especially significant contributors to amphibian population declines. These include the fungi Batrachochytrium dendrobatidis and B. salamandrivorans, and ranaviruses. In this review, we assess the effects of these three pathogens on amphibian hosts as found through experimental studies. Such studies offer valuable insights to the causal factors underpinning broad patterns reported through observational studies. We summarize key findings from experimental studies in the laboratory, in mesocosms, and from the field. We also summarize experiments that explore the interactive effects of these pathogens with other contributors of amphibian population declines. Though well-designed experimental studies are critical for understanding the impacts of disease, inconsistencies in experimental methodologies limit our ability to form comparisons and conclusions. Studies of the three pathogens we focus on show that host susceptibility varies with such factors as species, host age, life history stage, population and biotic (e.g., presence of competitors, predators) and abiotic conditions (e.g., temperature, presence of contaminants), as well as the strain and dose of the pathogen, to which hosts are exposed. Our findings suggest the importance of implementing standard protocols and reporting for experimental studies of amphibian disease. Keywords: amphibian population declines; experiments; pathogens; Batrachochytrium; ranavirus 1. Introduction Rapid rates of biodiversity loss have supported the notion that the Earth is heading toward a sixth major extinction event [1–3]. Current species extinction rates are higher than pre-human background rates, suggesting this biodiversity crisis is largely attributed to anthropogenic changes [1–6]. Although numerous species from all taxonomic groups are affected, amphibians are at the forefront of this Diversity 2018, 10, 81; doi:10.3390/d10030081 www.mdpi.com/journal/diversity Diversity 2018, 10, x FOR PEER REVIEW 18 of 33 1. Introduction Diversity 2018, 10, 81 2 of 49 Rapid rates of biodiversity loss have supported the notion that the Earth is heading toward a sixth major extinction event [1–3]. Current species extinction rates are higher than pre-humancrisis [ 3ba,7,ckground8]. Their populations rates, suggesting are declining this biodiversity more rapidly crisis than is thoselargely of birdsattributed or mammals to anthropogenic [8]. Like changes [1–6]. Although numerous species from all taxonomicother groups, groups amphibians are affected, are amphibia affected byns are multiple at the factorsforefront contributing of this crisis to [3,7,8]. population Their declinespopulations [9]. are declining more rapidly than those of birds or mammalsThese [8]. include Like ot habitather groups, destruction, amphibians contaminants, are affected climate by mult change,iple factors over-harvesting, contributing invasive to population species, declines [9]. These include habitat destruction, contaminants,predation, climate and change, infectious over-harvesting, diseases, all of invasive which mayspecies, work predation, independently and infectious or synergistically diseases, all to o affectf which may work independently or synergistically to affect amphibianamphibian populations populations [9–12] [9–12 ](Figure (Figure 1).1). Some Some of of the the research research we we summarize summarize below focused on how a particular pathogen alone affects a host, whereas somea studies particular addresse pathogend how alone a pathogen affects a may host, be whereas affected some by other studies variables addressed that may how interact a pathogen with may pathoge be ns. affected by other variables that may interact with pathogens. Figure 1. Potential abiotic and bioticFigure factors 1. Potential that may abiotic influence and biotic host–pathogen factors that dynamics may influence in amphibian host–pathogen disease dynamics systems. in amphibian disease systems. Among the major threats to amphibians are emerging infectious diseases (EIDs). Several prominent pathogens and associated EIDs affect amphibian populations worldwide. BatrachochytriumAmong thedendrobatidis major threats (hereafter to amphibians referred to areas Bd) emerging is a pathogenic infectious fungus diseases that (EIDs).causes amphibian Several chytridiomycosis [13–15]. Batrachochytrium prominent pathogens and associated EIDs affect amphibian populations worldwide. dendrobatidis (hereafter referred to as Bd) is a pathogenic fungus that causes amphibian chytridiomycosis [13–15]. This disease can cause population declines, local extinctions and contribute to species extinctions [8,16,17]. A related yet highly divergent fungal pathogen that also causes amphibian chytridiomycosis, Batrachochytrium salamandrivorans (hereafter referred to as Bsal), is a newly discovered pathogen primarily infecting salamanders [18]. Iridoviruses of the genus Ranavirus (hereafter referred to as Rv) have been implicated in declines and mass mortalities of amphibians [19–23]. Teacher et al. [22] stated that populations can respond differently to the virus and emergence can be transient, catastrophic, or persistent with recurrent mortality events. Although amphibians are hosts to an assortment of pathogens/parasites, including bacteria, viruses, fungi, water molds and helminths [13,24–27], we focus on Bd, Bsal and Rv, given accumulating evidence of their potentially devastating effects on amphibian populations worldwide. In particular, we focus on reviewing the literature that report the results of experiments (manipulation of key variables [28]) conducted with Bd, Bsal, and Rv concentrating on papers that used live amphibian hosts. Given the complexity of these host–pathogen systems, experimental approaches are crucial for disentangling potential mechanisms driving patterns of transmission and examining variation in lethal and sublethal effects due to host species, host life-history traits, pathogen strain, host populations, and environmental conditions. Prior to 2009, relatively few studies of amphibian diseases employed standard experimental designs [28] (Figure2). Since 2009, there has been a surge in the use of experiments to determine how diseases affect amphibians. Experimental design, methods, and interpretation vary; thus, it is useful to summarize these aspects to assess generality. One problem with experimental work on amphibian Diversity 2018, 10, x FOR PEER REVIEW 3 of 48 Diversity 2018, 10, 81 3 of 49 diseases has been the lack of standardization in experimental methods. Here, we present a synthesis diseasesof experimental has been thestudies lack ofand standardization attempt to inaddress experimental some methods.of the issues Here, weregarding present athe synthesis lack of experimentalstandardization studies and difficulties and attempt in togeneralizing address some about of thethe issuesdynamics regarding of the host–pathogen the lack of standardization systems we andfocus difficulties on. in generalizing about the dynamics of the host–pathogen systems we focus on. Figure 2.2.The The number number of experimental of experimental studies ofstudiesBatrachochytrium of Batrachochytrium dendrobatidis (Bd),dendrobatidisB. salamandrivorans (Bd), B. (Bsal)salamandrivorans and Ranavirus (Bsal)(Rv) and by Ranavirus year. (Rv) by year. SummarySummary of of Pathogen Pathogen Life Life Histories Histories BatrachochytriumBatrachochytrium dendrobatidis dendrobatidis First described by Longcore et al. [[29],29], Bd isis aa fungalfungal speciesspecies inin thethe phylumphylum Chytridiomycota thatthat has multiple hosts on everyevery continent where amphibians exist [[15,16]15,16] and hashas beenbeen associatedassociated withwith numerous population declines and some extinctions [[30–32].30–32]. Recent evidence suggests that that the source of BdBd waswas