Amphibian Mortality Research Articles

Amphibian Chytrid Prevalence in an Amphibian Community in Arid Australia

[Extract] The amphibian disease chytridiomycosis, caused by the pathogen Batrachochytrium dendrobatidis (Bd), has dramatically affected amphibians, causing population declines in over 200 species worldwide (Fisher et al. 2009). The disease is widespread, driving amphibian declines in North America (Muths et al. 2003; Briggs et al. 2005), Australia (Berger et al. 1998), Central America (Lips et al. 2006) and South America (Catenazzi et al. 2011). The variation in susceptibility to disease and mortality seen among host species, populations and locations is at least partially driven by interplay between external environmental and internal host-specific factors (Woodhams et al. 2007; Searle et al. 2011; Blaustein et al. 2012). While at a single location some species may be locally extirpated, others may persist (Lips et al. 2006). Amphibian infection prevalence and mortality rates due to chytridiomycosis are correlated with ambient environmental conditions: being highest during cooler months and at higher elevations (Berger et al. 1998; Woodhams and Alford 2005; Kriger and Hero 2008).

Malformations and mortality in the Asian Common Toad induced by exposure to pleurolophocercous cercariae (Trematoda: Cryptogonimidae)

Malformations and increased mortality due to infection by the digenetic trematode, Riberioa ondatrae have been reported for many species of amphibians. Severe malformations have also been reported in the Common Hourglass Tree Frog, Polypedates cruciger induced by pleurolophocercous cercariae in Sri Lanka in addition to the changes in the behaviour, development and survival of the host. We exposed pre-limb bud stage tadpoles (Gosner stages 25–26) of the Asian Common Toad, Duttaphrynus melanostictus to the same pleurolophocercous type cercariae under laboratory conditions. Molecular and morphological identification showed that these cercariae belonged Acanthostomum burminis infecting freshwater snakes as definitive hosts. These cercariae induced malformations (27.8%) and reduced survival to metamorphosis (53.8%). The magnitude of the effects increased with the dose of cercariae. Types of malformations were mainly axial, such as scoliosis and kyphosis. Severe limb malformations such as extra or missing limbs as reported for amphibians exposed to R. ondatrae were not observed in the D. melanostictus. Same authors reported a higher percentage of malformations previously when P. cruciger was exposed to the cercariae A. burminis compared to D. melanostictus. However, tadpoles of D. melanostictus, which are smaller compared to those of P. cruciger, experienced higher mortality than P. cruciger tadpoles. Trematode induced malformations and mortality in amphibians are highly variable and depend on multiple factors such as host species differences such as resistance to infection and tolerance, life-history characteristics such as size at metamorphosis and length of the metamorphosis period, and other factors such as size of the amphibian at the time of trematode exposure.

Evidence of chytrid‐mediated population declines in common midwife toad in Serra da Estrela, Portugal

AbstractThe emergence of a novel infectious disease, chytridiomycosis, is now widely recognized as a major cause of amphibian declines and biodiversity loss across local and global scales. Amphibian mortalities caused by the pathogenic chytrid fungus, Batrachochytrium dendrobatidis (Bd) were first recorded in Iberia, Europe over a decade ago. In August 2009, hundreds of post‐metamorphic common midwife toads (Alytes obstetricans) were found dead in the water and margins of a pond in the Serra da Estrela Natural Park, north‐central Portugal. Histological and genetic analyses confirmed their infection with Bd. Given the likelihood of a new outbreak of chytridiomycosis, we evaluated the possible impacts of this disease on populations of A. obstetricans within the Park by conducting field surveys during 2010 and 2011. We compared the present distribution and abundance of A. obstetricans with historical records, and quantified the present prevalence and intensity of infection by Bd. Results showed that (1) A. obstetricans disappeared from 67% of the 1 × 1 km squares where it was recorded in the past, (2) breeding is currently limited to 16% of the confirmed breeding sites in the past, and that (3) larvae are now less abundant, as well as are highly infected by Bd in the remaining sites. These effects were most pronounced at altitudes above 1200 m. Our findings suggest that an outbreak of chytridiomycosis is responsible for the rapid decline of A. obstetricans in Serra da Estrela, and we believe that urgent conservation measures are needed to prevent local extinction of the species.

Fungicide‐induced declines of freshwater biodiversity modify ecosystem functions and services

Although studies on biodiversity and ecosystem function are often framed within the context of anthropogenic change, a central question that remains is how important are direct vs. indirect (via changes in biodiversity) effects of anthropogenic stressors on ecosystem functions in multitrophic-level communities. Here, we quantify the effects of the fungicide chlorothalonil on 34 species-, 2 community- and 11 ecosystem-level responses in a multitrophic-level system. At ecologically relevant concentrations, chlorothalonil increased mortality of amphibians, gastropods, zooplankton, algae and a macrophyte (reducing taxonomic richness), reduced decomposition and water clarity and elevated dissolved oxygen and net primary productivity. These ecosystem effects were indirect and predictable based on changes in taxonomic richness. A path analysis suggests that chlorothalonil-induced reductions in biodiversity and top-down and bottom-up effects facilitated algal blooms that shifted ecosystem functions. This work emphasises the need to re-evaluate the safety of chlorothalonil and to further link anthropogenic-induced changes in biodiversity to altered ecosystem functions.

Road mortality of pond-breeding amphibians during spring migrations in the Mazurian Lakeland, NE Poland

The effects of road traffic mortality on amphibian populations are in many cases hard to estimate and speculate, primarily due to the inaccuracy of the methods employed in studies. We have analyzed amphibian road mortality during two breeding seasons on selected road sections adjoining spawning ponds and have critically reviewed methods used for estimating the impact of road traffic on migrating amphibians. The mortality rates of particular amphibian species differed significantly and varied according to year, site, and traffic volume. The highest mortality was recorded for Bufo bufo and the lowest for Lissotriton vulgaris. Species-specific parameter estimates of mortality, evaluated on a daily basis counts of roadkills, were positively correlated with the mean body mass of the amphibian species. The share of a given species among carcasses found on the road, commonly used in such studies, proved to be an unreliable measure of mortality. We found that nearly 60% of amphibian roadkill victims had disappeared within 24 h of exposure, and the number of missing carcasses was inversely related to body mass.

Spatial analysis of amphibian road mortality levels in northern Portugal country roads

Animal mortality caused by vehicle collisions is one of the main ecological impacts of roads. Amphibians are the most affected group and road fatalities have a significant impact on population dynamics and viability. Several studies on Iberian amphibians have shown the importance of country roads on amphibian road mortality, but still, little is known about the situation in northern Portugal. By being more permeable to amphibian passage, country roads represent a greater source of mortality than highways, which act as barriers. Thus, mitigation measures should be applied, but due to the extensive road network, the identification of precise locations (hotspots) and variables related to animal-vehicle collision is needed to plan these measures successfully. The aim of the study was to analyse the spatial occurrence and related factors linked to amphibian mortality on a number of country roads in northern Portugal, using spatial statistics implemented in GIS and applying a binary logistical regression. We surveyed 631 km of road corresponding to seven transects, and observed 404 individual amphibians: 74 (18.3%) alive and 330 (81.7%) road-killed. Bufo bufo represented 80% of the mortality records. Three transects showed clustered distribution of road-kills, and broadleaved forests and road ditches were the most important factors associated with hotspots of road-kill. Logistic regression models showed that habitat quality, Bufo bufo’s habitat preferences, and road ditches are positively associated with amphibians’ road mortality in northern Portugal, whereas average altitude and length of walls were negatively associated. This study is a useful tool to understand spatial occurrence of amphibian road-kills in the face of applying mitigation measures on country roads from northern Portugal. This study also considers the necessity of assessing the condition of amphibian local populations to understand their road-kills spatial patterns and the urgency to apply mitigation measures on country roads.

Varying Responses of Northeastern North American Amphibians to the Chytrid Pathogen Batrachochytrium dendrobatidis

Chytridiomycosis, caused by Batrachochytrium dendrobatidis (Bd), is widespread among amphibians in northeastern North America. It is unknown, however, whether Bd has the potential to cause extensive amphibian mortalities in northeastern North America as have occurred elsewhere. In the laboratory, we exposed seven common northeastern North American amphibian species to Bd to assess the likelihood of population-level effects from the disease. We exposed larval wood frogs (Lithobates sylvaticus) and postmetamorphic frogs of six other species to two different strains of Bd, a northeastern strain (JEL404) and a strain that caused die-offs of amphibians in Panama (JEL423), under ideal in vitro growth conditions for Bd. Exposed American toads (Anaxyrus americanus) all died; thus, this species may be the most likely to die from Bd-caused disease in the wild. Both Bd strains were associated with mortalities of wood frogs, although half the metamorphs survived. The Bd strain from Panama killed metamorphic green frogs (L. clamitans), whereas the northeastern strain did not, which means novel strains of Bd may lead to death even when local strains may not. No mortality was observed in four species (bullfrogs [L. catesbeianus], northern leopard frogs [L. pipiens], spring peepers [Pseudacris crucifer], and blue-spotted salamanders [Ambystoma laterale]) and in some individuals of green frogs and wood frogs that we exposed. This finding suggests these six species may be Bd vectors. Our results show that systematic exposures of amphibian species to Bd in the laboratory may be a good first step in the identification of species susceptible to Bd-caused declines and in directing regional conservation efforts aimed at susceptible species.

Factors contributing to amphibian road mortality in a wetland

Abstract To understand road characteristics and landscape features associated with high road mortality of amphibians in Zoige Wetland National Nature Reserve, we surveyed road mortality along four major roads after rainfall in May and September 2007. Road mortality of three species, Rana kukunoris, Nanorana pleskei and Bufo minshanicus, was surveyed across 225 transects (115 in May and 110 in September). Transects were 100 m long and repeated every two kilometers along the four major roads. We used model averaging to assess factors that might determine amphibian road mortality. We recorded an average of 24.6 amphibian road mortalities per kilometer in May and 19.2 in September. Among road characteristics, road width was positively associated with road morality for R. kukunori and B. minshanicus. Traffic volume also increased the road mortality of B. minshanicus in September. Of the landscape features measured, area proportions of three types of grassland (wet, mesic and dry) within 1 km of the roads, particularly that of wet grassland, significantly increased road mortality for R. kukunori and total mortality across all three species. To most effectively reduce road mortality of amphibians in the Zoige wetlands, we suggest better road design such as avoiding wet grasslands, minimizing road width, underground passes and traffic control measures. The implementation of public transit in the area would reduce traffic volume, and hence mortality.

Primer registro de Saprolegnia sp. en una población de anfibios en Colombia.

<p><strong> </strong></p> <p><strong>Objective: </strong><strong>Most </strong>research related to the decline of amphibians has been focused on the detection of the pathogenic fungus <em>Batrachochytrium dendrobatidis</em>. This fungus is the main pathogen detected around the world. However, research has shown the presence of another fungus, <em>Saprolegnia ferax</em>, as a cause of mortality in amphibians in North America. Our study suggests a possible interspecific transmission caused by the presence of rainbow trout; thus, amphibian declines may not be attributable only to the presence of a single pathogen, but to other organisms and factors. <strong>Materials and methods: </strong>Our study revealed the presence of <em>Saprolegnia</em> sp. in the Andean frog <em>Atelopus mittermeieri</em> using the imprinting technique with lactophenol blue staining, which allowed the typical structures of this fungus to be observed. <strong>Results:</strong> The importance of this discovery is the presence of two pathogenic fungi, <em>B. dendrobatidis </em>and <em>Saprolegnia</em>, which affecting simultaneously a population of amphibians. This finding brings attention to the eventual presence of other microorganisms that might be involved individually or collectively in the decline of amphibian species. <strong>Conclusions:</strong> This record suggests a possible transmission between rainbow trout (<em>Oncorhynchus mykiss</em>), an introduced species in the highlands of Colombia, which shares the same habitats with different species of amphibians in the Sanctuary of Flora and Fauna Guanentá in the upper river Fonce in the mid Cordillera Oriental of Colombia.</p> <p><strong>Key words: </strong>decline, Amphibians,<em> Saprolegnia</em>, Fishes, <em>Atelopus</em>, Colombia.</p> <p> </p> <p><strong> </strong></p><br />

Ranavirus-associated mass mortality in wild amphibians, The Netherlands, 2010: A first report

In 2010, a mass die-off of over 1000 wild water frogs (Pelophylax spp.) and at least 10 common newts (Lissotriton vulgaris) occurred in a pond in The Netherlands. Haemorrhagic disease with hepatomegaly and splenomegaly was evident. Microscopically, multiple organs presented cells with multifocal intracytoplasmic inclusion bodies, in which ranavirus-like particles were demonstrated ultrastructurally. All specimens examined tested positive for ranavirus by PCR. The sequence obtained showed a 100% identity with the one deposited for common midwife toad virus (CMTV). This is the first report of ranavirus-associated mortality in wild amphibian populations in The Netherlands. It is also the first time CMTV or a CMTV-like virus has been reported in these two species in the adult stage and outside of Spain.

Environmental Refuge from Disease-Driven Amphibian Extinction

Species that are tolerant of broad environmental gradients may be less vulnerable to epizootic outbreaks of disease. Chytridriomycosis, caused by the fungus Batrachochytrium dendrobatidis, has been linked to extirpations and extinctions of amphibian species in many regions. The pathogen thrives in cool, moist environments, and high amphibian mortality rates have commonly occurred during chytridiomycosis outbreaks in amphibian populations in high-elevation tropical rainforests. In Australia several high-elevation species, including the armored mist frog (Litoria lorica), which is designated as critically endangered by the International Union for the Conservation of Nature (IUCN), were believed to have gone extinct during chytridiomycosis outbreaks in the 1980s and early 1990s. Species with greater elevational ranges disappeared from higher elevations, but remained common in the lowlands. In June 2008, we surveyed a stream in a high-elevation dry sclerophyll forest and discovered a previously unknown population of L. lorica and a population of the waterfall frog (Litoria nannotis). We conducted 6 additional surveys in June 2008, September 2008, March 2009, and August 2009. Prevalences of B. dendrobatidis infection (number infected per total sampled) were consistently high in frogs (mean 82.5%, minimum 69%) of both species and in tadpoles (100%) during both winter (starting July) and summer (starting February). However, no individuals of either species showed clinical signs of disease, and they remained abundant (3.25 - 8.75 individuals of L. lorica and 6.5-12.5 individuals of L. nannotis found/person/100 m over 13 months). The high-elevation dry sclerophyll site had little canopy cover, low annual precipitation, and a more defined dry season than a nearby rainforest site, where L. nannotis was more negatively affected by chytridiomycosis. We hypothesize this lack of canopy cover allowed the rocks on which frogs perched to warm up, thereby slowing growth and reproduction of the pathogen on the hosts. In addition, we suggest surveys for apparently extinct or rare species should not be limited to core environments.

Development and Disease: How Susceptibility to an Emerging Pathogen Changes through Anuran Development

Ranaviruses have caused die-offs of amphibians across the globe. In North America, these pathogens cause more amphibian mortality events than any other pathogen. Field observations suggest that ranavirus epizootics in amphibian communities are common during metamorphosis, presumably due to changes in immune function. However, few controlled studies have compared the relative susceptibility of amphibians to ranaviruses across life stages. Our objectives were to measure differences in mortality and infection prevalence following exposure to ranavirus at four developmental stages and determine whether the differences were consistent among seven anuran species. Based on previous studies, we hypothesized that susceptibility to ranavirus would be greatest at metamorphosis. Our results did not support this hypothesis, as four of the species were most susceptible to ranavirus during the larval or hatchling stages. The embryo stage had the lowest susceptibility among species probably due to the protective membranous layers of the egg. Our results indicate that generalizations should be made cautiously about patterns of susceptibility to ranaviruses among amphibian developmental stages and species. Further, if early developmental stages of amphibians are susceptible to ranaviruses, the impact of ranavirus epizootic events may be greater than realized due to the greater difficulty of detecting morbid hatchlings and larvae compared to metamorphs.

The fungicide chlorothalonil is nonlinearly associated with corticosterone levels, immunity, and mortality in amphibians.

Background: Contaminants have been implicated in declines of amphibians, a taxon with vital systems similar to those of humans. However, many chemicals have not been thoroughly tested on amphibians or do not directly kill them.Objective: Our goal in this study was to quantify amphibian responses to chlorothalonil, the most commonly used synthetic fungicide in the United States.Methods: We reared Rana sphenocephala (southern leopard frog) and Osteopilus septentrionalis (Cuban treefrog) in outdoor mesocosms with or without 1 time (1×) and 2 times (2×) the expected environmental concentration (EEC) of chlorothalonil (~ 164 μg/L). We also conducted two dose–response experiments on O. septentrionalis, Hyla squirella (squirrel treefrog), Hyla cinerea (green treefrog), and R. sphenocephala and evaluated the effects of chlorothalonil on the stress hormone corticosterone.Results: For both species in the mesocosm experiment, the 1× and 2× EEC treatments were associated with > 87% and 100% mortality, respectively. In the laboratory experiments, the approximate EEC caused 100% mortality of all species within 24 hr; 82 μg/L killed 100% of R. sphenocephala, and 0.0164 μg/L caused significant tadpole mortality of R. sphenocephala and H. cinerea. Three species showed a nonmonotonic dose response, with low and high concentrations causing significantly greater mortality than did intermediate concentrations or control treatments. For O. septentrionalis, corticosterone exhibited a similar nonmonotonic dose response and chlorothalonil concentration was inversely associated with liver tissue and immune cell densities (< 16.4 μg/L).Conclusions: Chlorothalonil killed nearly every amphibian at the approximate EEC; at concentrations to which humans are commonly exposed, it increased mortality and was associated with elevated corticosterone levels and changes in immune cells. Future studies should directly quantify the effects of chlorothalonil on amphibian populations and human health.

Road mortality of amphibians and reptiles in the Anatolian part of Turkey

The amphibian and reptile specimens, crushed due to vehicle traffic, were recorded during herpetological trips to various regions of Anatolia between 2005 and 2009. In total, 183 individuals were detected, representing 28 species from 4 amphibian [Salamandridae (1), Ranidae (3), Bufonidae (2), and Hylidae (2)], 3 turtle [Testudinidae (1), Emydidae (1), and Geoemydidae (2)], 3 lizard [Agamidae (1), Anguidae (2), and Lacertidae (5)] and 2 snake [Colubridae (6) and Viperidae (2)] families. The measures required for decreasing herpetofaunal mortalities due to traffic are emphasized.

Effective Culvert Placement and Design to Facilitate Passage of Amphibians across Roads

Efficient deployment of culverts to mitigate mortality of amphibians on roadways requires identification of locations within road networks where animals cross (hotspots), points within identified hotspots for culvert placement, and attributes of culverts that make them behaviorally palatable to migrating individuals. In this study, we assessed road crossing frequency of Spotted Salamanders, Ambystoma maculatum, and American Toads, Anaxyrus americanus, along a 700-m transect within a known crossing hotspot, and related these distributions to habitat variables within the hotspot including the presence of existing culverts. We also placed experimental arrays of culverts of varying attributes in the path of migrating Spotted Salamanders to examine culvert preference by salamanders under typical movement environments and appropriate animal behavioral states. Our studies of patterns of road occurrence demonstrated that both species avoided crossing where there was a wetland within 15 m of the downslope of the road and that neither species showed a strong preference for crossing near existing culverts. When considering the choice for experimental culverts by Spotted Salamanders, we found no preference for culverts of varying aperture size, length, or substrate. Our results indicate that patterns of occurrences of the two species of amphibian within a crossing hotspot may be linked to the physical attributes at the site. For Spotted Salamanders in particular, predicting where they will cross within a hotspot may not be easy. Spotted Salamanders showed little preference for culverts of different design, indicating that a variety of culvert designs can suffice for mitigation if placed in appropriate locations.

Amphibian Encounter Rates on Roads with Different Amounts of Traffic and Urbanization

Although amphibians have relatively high rates of road mortality in urban areas, the conditions under which traffic threatens the survival of local amphibian populations remain unclear. In the Sandhills region of North Carolina (U.S.A.), we counted living and dead amphibians along two transects (total length 165 km) established on roads in areas with varying degrees of urbanization. We found 2665 individuals of 15 species, and amphibian encounter rates declined sharply as traffic and urban development increased. Regression-tree models indicated that 35 amphibians/100 km occurred on roads with <535 vehicles/day, whereas the encounter rate decreased to only 2 amphibians/100 km on roads with >2048 vehicles/day. Although mortality rate peaked at higher traffic levels (47% dead on roads with >5200 vehicles/day), the number of dead amphibians was highest at low levels of traffic. This suggests that areas where amphibian mortality is concentrated may actually contain the largest populations remaining on a given road transect.

The impact of roadside ditches on juvenile and sub-adult Bufo melanostictus migration

The amphibian species play an important role in much ecosystem functioning. However, it is severely impacted by civil engineering, which causes a high level of amphibian mortality. This is partly because roads isolate the amphibians’ habitats, causing the amphibians to cross them, during which time they can be killed by vehicles. However, secondly, roadside ditches can also have a fatal impact on the amphibians when they fall into them. This study focuses on this second issue and comprises two parts: field experiments and control experiments in the laboratory. In the field experiments, research into the roadside ditch impact on juvenile/sub-adult Bufo melanostictus migration was carried out by looking at different combinations of the ditch's angle, construction material and depth, and amount of plant growth. In the control experiments in the laboratory, the main focus of the study was the relationship between the body length of the juvenile/sub-adult Bufo melanostictus and its maximum jumping distance and jumping height, and the maximum gradient that Bufo melanostictus could climb. This research indicates that amphibian migration should be taken into account when roadside ditches are being planned. Civil engineering should consider the needs of the amphibians’ living space and thereby reduce its impact on the amphibians’ original habitat, so that human beings live in harmony with nature.

Minimising exposure of amphibians to pathogens during field studies

Many of the recent global amphibian mass mortalities, declines and extinctions have been attributed to the emerging infectious disease chytridiomycosis. There have been mass mortalities due to ranaviral disease but no major declines or extinctions. Controlling the transmission and spread of disease is of utmost importance, especially where there is the potential for human involvement. We have reviewed current hygiene guidelines for working with wild frogs, identified potential flaws and recommended those most suitable and effective for the field environment. Our within-site hygiene measures aim to reduce the risk of transmission among individuals. These measures encompass the capture, handling and holding of amphibians, skin disinfection before and after invasive procedures, marking frogs, sealing open wounds and treatment of accessory equipment. Our between-site hygiene measures aim to mitigate the risk of pathogen spread among populations. We have designed a risk calculator to help simplify and standardise the decision-making process for determining the level of risk and appropriate risk mitigation strategies to reduce the risk of increasing pathogen spread above background levels. Calculation of an overall risk score for pathogen spread takes into account the prior activity of field workers, the proposed activity, remoteness of the site, presence of known pathogens and the consequences of increased pathogen spread for amphibians in a given area.

The role of multiple stressors in ranavirus-caused amphibian mortalities in Acadia National Park wetlands

Recent studies suggest that multiple sublethal stressors compromise amphibian immune systems and increase susceptibility to disease. We examined two aspects of multiple stressors and incidence of ranavirus-caused amphibian mortalities in free-living amphibian populations: (1) among-pond differences in physical, chemical, and biological stressors that may exacerbate mortality events, and (2) temporal changes in within-pond stressors that coincide with mortality events. At the among-pond scale, we used principal components analysis and logistic regression followed by Akaike’s information criterion (QAICc) to identify stressors associated with disease incidence. Of the stressors we investigated, aluminum, temperature, and conductivity were most correlated with outbreaks, but it was unclear whether they increased ranavirus-caused mortality events. Sublethal stressors were difficult to isolate in the field and few were significantly associated with ranavirus across all breeding ponds. Our results suggest that each wetland, because of varied physical, biological, and chemical settings, will have its own suite of stressors that sublethally affect amphibians.

Road kills of amphibians in different land use areas from Sharavathi river basin, central Western Ghats, India

A survey of amphibian mortality on roads was carried out in the Sharavathi river basin in the central Western Ghats. Road kills in three different land use areas: agricultural fields, water bodies and forests were recorded for four days along three 100m stretches in each type of area. One-hundred-and-forty-four individuals belonging to two orders, eight families, 11 genera and 13 species were recorded in the survey. Kills/km observed were: in forest 55, agricultural fields 38 and water bodies 27, for an overall average of 40 kills/km. Kill species compositions varied significantly between land use areas, but not overall kill rates.