Amphibians as animal models for laboratory research in physiology

The concept of animal models is well honored, and amphibians have played a prominent part in the success of using key species to discover new information about all animals. As animal models, amphibians offer several advantages that include a well-understood basic physiology, a taxonomic diversity well suited to comparative studies, tolerance to temperature and oxygen variation, and a greater similarity to humans than many other currently popular animal models. Amphibians now account for approximately 1/4 to 1/3 of lower vertebrate and invertebrate research, and this proportion is especially true in physiological research, as evident from the high profile of amphibians as animal models in Nobel Prize research. Currently, amphibians play prominent roles in research in the physiology of musculoskeletal, cardiovascular, renal, respiratory, reproductive, and sensory systems. Amphibians are also used extensively in physiological studies aimed at generating new insights in evolutionary biology, especially in the investigation of the evolution of air breathing and terrestriality. Environmental physiology also utilizes amphibians, ranging from studies of cryoprotectants for tissue preservation to physiological reactions to hypergravity and space exploration. Amphibians are also playing a key role in studies of environmental endocrine disruptors that are having disproportionately large effects on amphibian populations and where specific species can serve as sentinel species for environmental pollution. Finally, amphibian genera such as Xenopus, a genus relatively well understood metabolically and physiologically, will continue to contribute increasingly in this new era of systems biology and "X-omics."     

Establishment risk and potential invasiveness of the selected exotic amphibians from pet trade in the European Union

The pet trade has grown in recent years and become the most important pathway for the introduction of non-indigenous species of amphibians and reptiles worldwide. Amphibians traded on the pet market have been widely overlooked in systematic invasion studies, so their establishment potential and invasion dynamics remain poorly understood, despite the fact that the impact of invaders from this taxonomic group on native biota may be considerable. The determination of the most common species of amphibians traded as pets was based on a survey of the market in the Czech Republic, which is an export hub for ornamental aquatic animals into the European Union (EU). Subsequently, the establishment potential of the most common species for EU was determined using a proven risk assessment model. Amphibians that scored higher than the established species Xenopus laevis were additionally evaluated for their invasion potential using the Amphibian Invasiveness Screening Kit (AmphISK).Sixteen species of amphibians in pet market (14 Anura, 2Caudata) have an establishment potential in the EU. However, none of these species have reached the invasion potential score of already spreading species X. laevis and Lithobates catesbeianus. The species closest to the threshold value were Lithobates pipiens and Pelophylax saharicus. Amphibians represent a rather small group within all traded animals but this fact should not lead us to an underestimation of their potential invasiveness.     

Population differentiation of temperate amphibians in unpredictable environments

Amphibians are a globally distributed and diverse lineage, but much of our current understanding of their population genetic structure comes from studies in mesic temperate habitats. We characterize the population genetic structure of two sympatric explosive breeding amphibians in the southwestern deserts of the United States: the Great Plains toad ( Anaxyrus cognatus ) and Couch's spadefoot toad ( Scaphiopus couchii ). For both species, we find limited genetic differentiation even between populations in adjacent valleys separated by dispersal barriers such as mountainous habitats. To understand how population genetic patterns in these two arid-adapted species compare to taxa in more mesic environments, we computed a standardized measure of population differentiation for A. cognatus , S. couchii , and for pond-breeding amphibians that inhabit mesic temperate environments. Our results indicate that the arid-adapted species have lower population genetic structure at fine and moderate scales than most other amphibian species we surveyed. We hypothesize that stochasticity in the availability of appropriate breeding sites as well as landscape homogeneity may result in increased population connectivity in desert-adapted frogs. Future work examining fine-scale population structure in amphibians from a diversity of habitats will test the generality of our findings. Intraspecific comparisons among localities with varied seasonality and habitats will be particularly useful for investigating the interaction between species-typical population dynamics and environmental characteristics as determinants of population connectivity in pond-breeding amphibians.     

Facility design and associated services for the study of amphibians

The role of facilities and associated services for amphibians has recently undergone diversification. Amphibians traditionally used as research models adjust well to captivity and thrive with established husbandry techniques. However, it is now necessary to maintain hundreds of novel amphibian species in captive breeding, conservation research, and biomedical research programs. These diverse species have a very wide range of husbandry requirements, and in many cases the ultimate survival of threatened species will depend on captive populations. Two critical factors have emerged in the maintenance of amphibians, stringent quarantine and high-quality water. Because exotic diseases such as chytridiomycosis have devastated both natural and captive populations of amphibians, facilities must provide stringent quarantine. The provision of high-quality water is also essential to maintain amphibian health and condition due to the intimate physiological relationship of amphibians to their aquatic environment. Fortunately, novel technologies backed by recent advances in the scientific knowledge of amphibian biology and disease management are available to overcome these challenges. For example, automation can increase the reliability of quarantine and maintain water quality, with a corresponding decrease in handling and the associated disease-transfer risk. It is essential to build facilities with appropriate nontoxic waterproof materials and to provide quarantined amphibian rooms for each population. Other spaces and services include live feed rooms, quarantine stations, isolation rooms, laboratory space, technical support systems, reliable energy and water supplies, high-quality feed, and security. Good husbandry techniques must include reliable and species-specific management by trained staff members who receive support from the administration. It is possible to improve husbandry techniques for many species by sharing knowledge through common information systems. Overall, good facility design corresponds to the efficient use of space, personnel, energy, materials, and other resources.     

Amphibians as metrics of critical biological thresholds in forested headwater streams of the Pacific Northwest, U.S.A.

1. Amphibians are recognized both for their sensitivity to environmental perturbations and for their usefulness as cost-effective biometrics of ecosystem integrity (=system health).
2. Twenty-three years of research in headwater streams in the Klamath-Siskiyou and North Coast Bioregions of the Pacific Northwest, U.S.A., showed distinct patterns in the distribution of amphibians to variations in water temperature, % fine sediments and the amount of large woody debris (LWD).
3. Here, we review seven studies that demonstrate connections between species presence and abundance and these three in-stream variables. These data were then used to calculate realized niches for three species, the southern torrent salamander, the larval coastal tailed frog and the larval coastal giant salamander, relative to two of these environmental stressors (water temperature and % fine sediments). Moreover, multivariate generalized additive models were used to predict the presence of these three amphibians when these three stressors act in concert.
4. Stream-dwelling amphibians are shown to be extremely sensitive to changes in water temperature, amounts of fine sediment and LWD, and specific thresholds and ranges for a spectrum of animal responses can be used to manage for headwater tributary ecosystem integrity.
5. Consequently, amphibians can provide a direct metric of stream ecosystem integrity acting as surrogates for the ability of a stream network to support other stream-associated biota, such as salmonids, and their related ecological services.     

Spinal Cord Regeneration in Amphibians: A Historical Perspective

In some vertebrates, a grave injury to the central nervous system (CNS) results in functional restoration, rather than in permanent incapacitation. Understanding how these animals mount a regenerative response by activating resident CNS stem cell populations is of critical importance in regenerative biology. Amphibians are of a particular interest in the field because the regenerative ability is present throughout life in urodele species, but in anuran species it is lost during development. Studying amphibians, who transition from a regenerative to a nonregenerative state, could give insight into the loss of ability to recover from CNS damage in mammals. Here, we highlight the current knowledge of spinal cord regeneration across vertebrates and identify commonalities and differences in spinal cord regeneration between amphibians.     

Diseases of amphibians

The development and refinement of amphibian medicine comprise an ongoing science that reflects the unique life history of these animals and our growing knowledge of amphibian diseases. Amphibians are notoriously fastidious in terms of captive care requirements, and the majority of diseases of amphibians maintained in captivity will relate directly or indirectly to husbandry and management. Investigators have described many infectious and noninfectious diseases that occur among various species of captive and wild amphibians, and there is considerable overlap in the diseases of captive versus free-ranging populations. In this article, some of the more commonly reported infectious and noninfectious diseases as well as their etiological agents and causative factors are reviewed. Some of the more common amphibian diseases with bacterial etiologies include bacterial dermatosepticemia or "red leg syndrome," flavobacteriosis, mycobacteriosis, and chlamydiosis. The most common viral diseases of amphibians are caused by the ranaviruses, which have an impact on many species of anurans and caudates. Mycotic and mycotic-like organisms cause a number of diseases among amphibians, including chytridiomycosis, zygomycoses, chromomycoses, saprolegniasis, and ichthyophoniasis. Protozoan parasites of amphibians include a variety of amoeba, ciliates, flagellates, and sporozoans Common metazoan parasites include various myxozoans, helminths (particularly trematodes and nematodes), and arthropods. Commonly encountered noninfectious disease etiologies for amphibians include neoplasia, absolute or specific nutritional deficiencies or overloads, chemical toxicities, and inadequate husbandry or environmental management.     

环境DNA在两栖动物监测中的应用研究进展

两栖动物是我国受威胁程度最高的动物类群,加强两栖动物资源调查和多样性监测,是开展两栖动物保护和濒危物种拯救行动的关键性基础工作。传统的两栖动物监测主要以形态学和声学为基础,耗时费力,且难以发现一些隐蔽性较强的稀有物种。基于环境DNA(environmental DNA, eDNA)的调查方法以其快速、灵敏、高效、无创等独特优势,为两栖动物多样性监测及保护提供了新的工具。综述了eDNA在两栖动物多样性监测、外来入侵和珍稀濒危物种调查、物种丰度或生物量估测等研究领域的应用进展,分析了两栖动物eDNA产生、扩散、迁移和降解的动态变化特征及其关键影响因子,探讨了eDNA应用于两栖动物监测研究的局限性并提出了优化建议,同时对未来的研究方向进行了展望,以充分挖掘eDNA在两栖动物监测中的应用潜力,为两栖动物多样性保护和管理提供新的思路。     

Evaluating multiple causes of amphibian declines of Ecuador using geographical quantitative analyses

Amphibians are declining at alarming rates worldwide; however, the causes of these declines remain somewhat elusive. Here we evaluated three major threats implicated in declines of populations and disappearance of Ecuadorian amphibians: chytridiomicosis, climate change, and habitat loss. We assessed spatial patterns of these key threats to Ecuadorian amphibians using a multi‐species database of endemic frogs along with information on the pathogen's distribution and environmental requirements, species sensitivity to climate change (indirectly based on species geographical distribution and ecological properties) and habitat loss. Our results show that amphibians display a non‐random pattern of extinction risk, both geographically and taxonomically. Further, climate change, chytridiomicosis, and their synergetic effects, are likely currently exerting the greatest impact on amphibians in Ecuador, while habitat loss does not seem to be causing precipitous declines. The most threatened species under the IUCN extinction risk categories are exactly those that appear to be the most affected by these threats. By examining multiple potential causes of amphibian threat level in a spatially explicit framework our study provides new insights about what combination of factors are most important in amphibian declines in a tropical diversity hotspot. Further, our approach and conclusions are useful for studying declines in other regions of the world.     

Medicine and surgery of amphibians

Amphibians are most notably characterized by their glandular skin, which they shed regularly and ingest routinely. It is advisable to handle amphibians only with protective gloves to avoid damaging their skin. These animals absorb water readily across the skin as a means of maintaining hydration. They also easily absorb drugs and anesthetics that are applied directly to the skin. Investigators commonly utilize cutaneous respiration in amphibians and evaluate skin abnormalities via wet mount preparations, skin scrapes, and biopsy. The examination of blood samples can be useful in evaluating the status of ill amphibians, although the similarity in function of amphibian blood cell types and those of other species is largely unknown. If surgery is required, it is necessary to fast the animals before surgery, and to monitor their hydration. The wet environment required for amphibian surgery makes sterile technique challenging, and it is advisable to institute prophylactic antibiotic therapy before the procedure. The anesthetic of choice for amphibian surgery is tricaine methanesulfonate (MS-222). Postoperative recommendations include fluids, nutritional support if necessary, and analgesia. If euthanasia is required, MS-222 overdose or pentobarbital injection are the preferred methods.     

Amphibian biology and husbandry

Extant amphibians comprise three lineages-- salamanders (Urodela or Caudata), frogs and toads (Anura), and caecilians (Gymnophiona, Apoda, or Caecilia)--which contain more than 6,000 species. Fewer than a dozen species of amphibians are commonly maintained in laboratory colonies, and the husbandry requirements for the vast majority of amphibians are poorly known. For these species, a review of basic characteristics of amphibian biology supplemented by inferences drawn from the morphological and physiological characteristics of the species in question provides a basis for decisions about housing and feeding. Amphibians are ectotherms, and their skin is permeable to water, ions, and respiratory gases. Most species are secretive and, in many cases, nocturnal. The essential characteristics of their environment include appropriate levels of humidity, temperature, and lighting as well as retreat sites. Terrestrial and arboreal species require moist substrates, water dishes, and high relative humidity. Because temperature requirements for most species are poorly known, it is advisable to use a temperature mosaic that will allow an animal to find an appropriate temperature within its cage. Photoperiod may affect physiology and behavior (especially reproduction and hibernation), and although the importance of ultraviolet light for calcium metabolism by amphibians is not yet known, ecological observations suggest that it might be important for some species of frogs. Some amphibians are territorial, and some use olfactory cues to mark their territory and to recognize other individuals of their species. All amphibians are carnivorous as adults, and the feeding response of many species is elicited by the movement of prey. Diets should include a mixture of prey species, and it may be advisable to load prey with vitamins and minerals.     

Ultraviolet radiation and Vitamin D3 in amphibian health, behaviour, diet and conservation

Amphibians are currently suffering a period of mass extinction with approximately 20% of species under severe threat and more than 120 species already extinct. In light of this crisis there is an urgency to establish viable ex situ populations and also find the causes of in situ declines. The role of ultraviolet radiation and Vitamin D₃ in amphibian health directly influences both ex situ and in situ populations. Vitamin D₃ can be photosynthesised endogenously via UV-B radiation (UV-B), or acquired through the diet, and then metabolised to calcitriol the biologically active hormonal form. Although, there is a lack of literature concerning Vitamin D₃ requirements and calcitriol synthesis in amphibians, amphibians are likely to have similar Vitamin D₃ requirements and metabolic processes as other vertebrates due to the phylogenetically conservative nature of calcitriol biosynthesis. Deficiencies in calcitriol in amphibians result in nutritional metabolic bone disease (NMBD) and could compromise reproduction and immunity. However, excess biologically active UV radiation has also proven detrimental across all three amphibian life stages and therefore could impact both in situ and ex situ populations. Here we review the role and necessity of UV-B and calcitriol in amphibians and the potential for negative impacts due to excessive exposure to UV radiation. We also identify priorities for research that could provide critical information for maintaining healthy in ex situ and in situ populations of amphibians.     

Species richness of helminth parasites in Mexican amphibians and reptiles

Abstract. Amphibians and reptiles represent an important group of vertebrates in Mexico; on a global scale 10% of the biodiversity of these groups is found in the country, attaining extraordinarily high levels of endemism (60.7% and 53.7%, respectively). However, fewer than 20% of the known species of amphibians and reptiles in Mexico have been surveyed for helminths, so the inventory is far from complete. We assembled a data base that includes a total of 1246 records (entries) of which 460 correspond to helminths in amphibians and 786 to helminths in reptiles. In total, only 41 species of amphibians (14% of those occurring in Mexico) and only 118 species of reptiles (17% of those occurring in Mexico) have been studied for helminth parasites. From amphibians, 119 species of helminths belonging to 60 genera have been recorded, while 239 species of helminths representing 113 genera have been described from Mexican reptiles. One feature of the distribution of helminths of Mexican amphibians and reptiles is its asymmetry, as seen in representation of helminth groups, host groups and geographical range. However, such statistical asymmetry might be an artefact of sampling effort. Based on our data, we estimate that if all the herpetofauna of Mexico could be studied in the following years, approximately 827 additional species of helminths from amphibians and approximately 1403 from reptiles would be described.     

Mitigating amphibian chytridiomycosis with bioaugmentation: characteristics of effective probiotics and strategies for their selection and use

Probiotic therapy through bioaugmentation is a feasible disease mitigation strategy based on growing evidence that microbes contribute to host defences of plants and animals. Amphibians are currently threatened by the rapid global spread of the pathogen, Batrachochytrium dendrobatidis (Bd), which causes the disease chytridiomycosis. Bioaugmentation of locally occurring protective bacteria on amphibians has mitigated this disease effectively in laboratory trials and one recent field trial. Areas still naïve to Bd provide an opportunity for conservationists to proactively implement probiotic strategies to prevent further amphibian declines. In areas where Bd is endemic, bioaugmentation can facilitate repatriation of susceptible amphibians currently maintained in assurance colonies. Here, we synthesise the current research in amphibian microbial ecology and bioaugmentation to identify characteristics of effective probiotics in relation to their interactions with Bd, their host, other resident microbes and the environment. To target at‐risk species and amphibian communities, we develop sampling strategies and filtering protocols that result in probiotics that inhibit Bd under ecologically relevant conditions and persist on susceptible amphibians. This filtering tool can be used proactively to guide amphibian disease mitigation and can be extended to other taxa threatened by emerging infectious diseases.     

Polymorphic adaptations in metazoans to establish and maintain photosymbioses

Mutualistic symbioses are common throughout the animal kingdom. Rather unusual is a form of symbiosis, photosymbiosis, where animals are symbiotic with photoautotrophic organisms. Photosymbiosis is found among sponges, cnidarians, flatworms, molluscs, ascidians and even some amphibians. Generally the animal host harbours a phototrophic partner, usually a cyanobacteria or a unicellular alga. An exception to this rule is found in some sea slugs, which only retain the chloroplasts of the algal food source and maintain them photosynthetically active in their own cytosol – a phenomenon called ‘functional kleptoplasty’. Research has focused largely on the biodiversity of photosymbiotic species across a range of taxa. However, many questions with regard to the evolution of the ability to establish and maintain a photosymbiosis are still unanswered. To date, attempts to understand genome adaptations which could potentially lead to the evolution of photosymbioses have only been performed in cnidarians. This knowledge gap for other systems is mainly due to a lack of genetic information, both for non‐symbiotic and symbiotic species. Considering non‐photosymbiotic species is, however, important to understand the factors that make symbiotic species so unique. Herein we provide an overview of the diversity of photosymbioses across the animal kingdom and discuss potential scenarios for the evolution of this association in different lineages. We stress that the evolution of photosymbiosis is probably based on genome adaptations, which (i) lead to recognition of the symbiont to establish the symbiosis, and (ii) are needed to maintain the symbiosis. We hope to stimulate research involving sequencing the genomes of various key taxa to increase the genomic resources needed to understand the most fundamental question: how have animals evolved the ability to establish and maintain a photosymbiosis?     

Amphibian behavioral diversity offers insights into evolutionary neurobiology

Recent studies have served to emphasize the unique placement of amphibians, composed of more than 8000 species, in the evolution of the brain. We provide an overview of the three amphibian orders and their respective ecologies, behaviors, and brain anatomy. Studies have probed the origins of independently evolved parental care strategies in frogs and the biophysical principles driving species-specific differences in courtship vocalization patterns. Amphibians are also important models for studying the central control of movement, especially in the context of the vertebrate origin of limb-based locomotion. By highlighting the versatility of amphibians, we hope to see a further adoption of anurans, urodeles, and gymnophionans as model systems for the evolution and neural basis of behavior across vertebrates.