Panamanian golden frog | Atelopus zeteki | Brian Gratwicke April 2019 The frogs fight against fungal foe. What are their odds? We are currently in the midst of a global amphibian crisis, caused by perhaps the worst animal disease ever recorded. A fungal pathogen Batrachochytrium dendrobatidis, commonly known as Bd is contributing to the current sixth mass extinction event. This novel pathogen inflicts a deadly and merciless disease on at least 42% of all known amphibians. This leaves the question on the lips of everyone cares about frogs: can this fungus be stopped? Since the 1970’s, Bd has been devastating whole communities of once flourishing frogs. To date 695 species of amphibians have been affected by Bd and with as little as 17% of them sampled, this number is likely to be far greater. Funding, awareness and interest in amphibian conservation is not evenly spread across the globe, meaning areas that appear to be unaffected may be worse off than we believe. In the face of such an overwhelming pandemic it has been hard to find much hope. However, good news stories do exist, even if they’re few and far between. Individual species can recover, with numbers of Australian Common mist frogs currently on the rise. Stabilisations like these have been facilitated by environmental refugia: Bd free areas where populations can persist. Bd requires a unique set of conditions to survive, so amphibians living in regions that are unfavourable to Bd can endure the global pandemic. If we can define and protect these habitats that act as safe zones, we may be able to improve the effectiveness of conservation. However, climatic variation means that these areas too are constantly changing, suggesting they may not provide a permanent solution. Patterns of recovery versus decline in species have been nigh on impossible to predict, fluctuations in numbers happen all the time and it is hard to say with any certainty whether a species is safe. Spotted tree frogs, Littoria spenceri are one species that were thought to be bouncing back but have subsequently gone into remission. These spotted frogs join 43 other Australian species that are 14 threatened by Bd, 7 of which are already believed to be extinct. The future of global amphibians is now in the hands of us humans; we must reduce our own impact on these frog’s precious habitats and seek solutions to aid this animal crisis. How could we help? Bd is not the sole cause of amphibian decline; it is a result of a combination of factors: including habitat destruction and fragmentation. Frogs are very sensitive to changes in their environment and are heavily affected by deforestation, as they live in small territories where they aggregate to breed. If amphibians were to be able to persist even in the presence of this deadly disease, then humans must move to protect the habitats in which they live. Pesticide use also poses a major threat, as amphibian skin is highly permeable to these toxic chemicals. Therefore, biochemical pollution needs to be regulated in areas in which critically endangered species exist, to the benefit of all wildlife, not just frogs. Humans have inadvertently assisted the spread of this killer fungus. The industrial revolution and the sharp rise in international trade over the past 150 years proliferated the movement of frogs for food and the biomedical industry; many of these frogs were released into the wild and Bd alongside. Modern transportation now allows diseases to survive global journeys that they previously wouldn’t. With the ability for pathogens to cross-continents rapidly, Homo sapiens have inadvertently opened Pandora’s box. We have exposed wildlife and humans to novel pathogens like Bd and allowed them to desiccate naïve populations, re-writing millions of years of evolution. It is important to prevent further pathogenic spread of this fungus by enforcing strict biosecurity on both a national and international level. Stopping the trade of infected frogs between countries and ensuring equipment sterilisation between habitats would help mitigate dispersal. With 62% of the world’s countries already affected by Bd, regulating amphibian trade is critical. This would prevent currently unaffected countries like Papua New Guinea where humans haven’t introduced Bd, to remain unscathed. Although this doesn’t provide a solution for infected populations and doesn’t prevent natural spread, it is a key step on the road to preventing further damage. Can we treat Bd? The most effective treatment of Bd has occurred on the Spanish island of Mallorca. Here, scientists have actually been able to eradicate the disease. The fungus was accidentally introduced to the island by the reintroduction of a captively bred population of midwife toads, Alytes muletensis. The midwife toads live in small montane ponds, where the adults come to spawn. Teams of scientists have managed to collect and treat the spawn with fungicides back in the lab. During the treatment the ponds were drained and cured of Bd by sun exposure and use of biocide Virkon S. Upon returning the tadpoles to the pools and subsequently sampling the inhabitants, these measures seem to have worked. Amphibians on Mallorca now appear to be chytrid-free, with no unintended side effects recorded so far. 15 Mallorcan midwife toad | Alytes muletensis | Tuurio and Wallie This treatment has worked in a hot dry climate, where very amphibians few species live. Sadly, the majority of Bd associated declines are observed in cool montane regions and tropical forests. Therefore, it is likely most species couldn’t be treated in this way. However, the midwife toad method could work for other infected Mediterranean populations and provide a framework by which others could be treated. Jamie Bosch who was pivotal to the midwife toad project believes that, “we can’t stand by idly thinking that chytridiomycosis can’t be combatted;” a mindset we should all adopt. A potential treatment for Bd will very likely be found in the frogs themselves. Amphibian skin carries a large array of bacteria that are involved with immune defence. In some species their bacteria are capable of protecting them from Bd. One of these bacteria, Janthinobacterium lividum, could prove a natural fungicidal cure. Focusing research into resistant frogs could hopefully provide an effective treatment in time. Skin bacterial research has been undertaken on captive Panamanian golden frogs, Atelopus zeteki. Isolates of the bacterium J. lividum were taken from mountain yellow-legged frogs, Rana muscosa and transferred to A. zeteki, with the hope that it would work like an antibiotic. The results have not been overwhelming- only five A. zeteki managed to beat Bd infection without treatment. These resistant frogs appear to have survived due to having a different community of skin bacteria to those that didn’t survive infection. Subsequent applications of these bacteria to other frogs have so far been unsuccessful. Further research with Peruvian marsupial frogs, Gastrotheca excubitor also suggests skin bacteria may be behind Bd resistance. However, potential skin bacteria treatments would likely only be able to treat infected individuals, rather than working on whole populations or habitats. Therefore, there is a long way to go before this kind of treatment becomes a sure-fire solution for Bd, but it seems like a good place to place our hope. Alternatively, we could combat Bd another way- maybe we should target the infective thread of the fungus: the spores. Introducing micro-crustaceans that would feed upon the spores and limit infection rates might be an alternative. Similarly, exploring the use of Virkon S treatments, like with the midwife 16 toads could be an option. However, introduction of species and chemical treatments to the water system would likely have fatal consequences to other micro and macrobiota. Therefore, sufficient and vigorous risk analysis would need to be undertaken beforehand. The track record for previous introductions of novel species is not good, but finding an alternative method of limiting Bd infection could be an effective way of fighting the crisis. What can we learn from global declines? There are a disproportionately large number of declines in the Americas, where conservation is critical for the survival of amphibians. Australia is in a similar situation, where the montane temperate forests are ideal habitat for Bd leading to numerous declines. Furthermore, an alternative strain of Bd has recently been discovered that attacks European populations of newts and salamanders, posing a new threat in the Northern Hemisphere. The apparent lack of declines in the rest of the globe may not provide the full picture; most likely sufficient research will not have been undertaken in many parts of the globe. Fig 1. World map, including statistics for Bd driven amphibian decline around the globe. (Scheele et al., 2019) Leading amphibian expert Simon O’Hanlon of Imperial College London has described a particularly virulent strain of Bd, which is responsible for the terrible impact inflicted on global populations. This strain is thought to have arisen from a South East Asian origin, perhaps the Korean peninsula. If Bd has come from Asia, then amphibians may have evolved to coexist alongside it. Therefore, Asian frogs are ideal candidates to look for a solution. Very few cases of amphibian decline are reported in Asia; so could this be the continent that holds the answer? It is of course possible that declines are occurring here and they have not been reported. However, if Simon O’Hanlon’s Korean hypothesis is true, then this region requires further investigation. The skin bacterial communities of these frogs might provide an effective cure for infected frogs and a natural non-invasive alternative to biocides like Virkon S, making Asian research a priority. 17 Are Atelopus toads making an unlikely recovery? One of the groups of amphibians that are most severely affected by Bd are the Harlequin toads (Atelopus).