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Frog Cross Breeding: A Comprehensive Overview



A scientist at The Manchester Museum in England allowed the two species of endangered Central American leaf frogs housed within the same chamber to interbreed to better understand how closely these parents are related. Understanding the genetic relationships between, and even within, species is important when trying to protect them.


This was a match made in lab heaven. The parents, Agalychnis annae and Agalychnis moreletii, wouldn't cross paths on their own, since they occupy different regions in Central America. In the past 30 years, populations of endangered leaf frogs have completely disappeared, particularly at cooler, high elevations. The amphibian-devastating chytrid fungus is implicated.




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On warm, rainy nights over the next few weeks, Cornell University biology students and members of the campus Herpetology Society will gather along a stretch of road in the Ringwood Preserve, about six miles from campus. Their mission: to stop and slow down automobile traffic, giving frogs and salamanders right of way to cross from the forest to a mating pond.


Millions upon millions of adult amphibians are killed on the first warm, rainy nights of spring as they cross roads throughout the Northeastern United States in their migration from breeding grounds to breeding ponds.


The Cornell students want to make a small contribution to end this slaughter between now and May 31, the end of the breeding season. "As a class and as conservation-minded herpetologists, we plan to decrease the number of animals killed this spring by assisting them to cross the road from the forest to where they reproduce," says Jacqualine B. Grant, a graduate student in neurobiology and animal behavior. "We want do this as safely as possible for them and for us."


A few months ago, students from the society and from the university's herpetology class, taught by Harry W. Greene, Cornell professor in ecology and evolutionary biology, contacted Ward A. Hungerford, superintendant of the local Tompkins County Highway Department, to explain their idea for a frog-crossing project. The department has cooperated by lending the group full-coverage reflective vests, slow/stop paddles, reflective barrels and traffic cones, work-zone signs, flagger signs and "be prepared to stop" signs. And county road technician Jerry Stern has marked the road so the students will know where to place their signs. The highway department has even trained the frog-crossing team in safe night-flagging techniques. The students have handed out leaflets that explain to nearby residents why traffic on Ringwood Road is slowing down. With everything in place, the students need one more ingredient to make this project a success: "Now, we are just waiting for it to rain," says Grant.


"Long before humans built a road, there was a pond. And all these pond-breeding amphibians need water to lay their eggs," says Kelly R. Zamudio, Cornell assistant professor of ecology and evolutionary biology. "If they don't lay their eggs in the pond, the eggs dry up. The eggs develop into tadpoles, the tadpoles develop into froglets, and then, as frogs, they leave the pond. This migration is not something they have choice about."


The frogs' and salamanders' reproduction is critical, says Zamudio. Most creatures do not migrate across roads to breeding areas en masse like this. If something goes wrong, or if too many get killed, large numbers of other individuals could be at reproductive risk. Spring peepers live in forests and feast on mosquitoes and flies. Each spring, the peepers climb down from the trees and travel to water, where they meet other peepers, court, breed and lay eggs, says Grant. "This whole procedure takes place in a relatively short period of time," she says. "What this means is that hundreds or even thousands of frogs or salamanders may emerge on a single rainy, warm, spring night and cover the roads on their way to the pond."


Both mules and hinnies are infertile, because donkeys and horses have a different number of chromosomes, the protein structures on which genetic material is present. Yeasts, plants, fish and amphibians (in contrast to mammals) can however produce fertile hybrids. Professor of Molecular Developmental Biology Gert Jan Veenstra: "For instance, in frogs a duplication of chromosomes can appear, in which the whole set of chromosomes from the father and the mother are passed on to the next generation." However, there is another problem: some hybrids are not viable, while the crossbreeding the other way around is. The embryos are genetically identical, but there is a difference in viability, depending on the father species and the mother species. Veenstra: "Even though it is vital for evolution, the mechanisms of viable and non-viable hybrids are to this date unknown."


The scientists showed this phenomenon in research with two related frog species: Xenopus tropicalis and Xenopus laevis (also known as the Western and African clawed frogs, respectively). When a female African clawed frog is crossbred with a male Western clawed frog, the embryos are viable. However, the other way around, crossbreeding a male African clawed frog and a female Western clawed frog leads to embryos that die in the early stages of development. Why that is, remained unclear.


On 10 January, researchers publish what went wrong during this crossbreeding: the maternal molecular machinery of the Western clawed frog cannot fully recognize the paternal chromosomes of the African clawed frog. Two specific pieces of the paternal chromosomes are incompatible with the maternal cell and thus the separation of the chromosomes during cell division is disrupted. These cells now lack a large number of important genes, such as genes for metabolism, and therefore quickly die.


This shows there is a strong asymmetry when it comes to hybrids, depending on the father species and mother species. "These findings are important, because these type of hybrids are present in nature and in some cases lead to new species. When new species are formed, there seems to be a period of transition: closely related species are able to produce viable offspring, but if the chromosomes are no longer compatible it leads to asymmetric results of crossbreeding. When species further separate, crossbreeding no longer leads to viable offspring. We here show the cellular mechanism behind this phenomenon," states Gert Jan Veenstra.


The viable hybrid of a male Western clawed frog and a female African clawed frog also revealed a molecular mechanism: parasitic DNA elements (transposons) are activated in one of the genomes. Veenstra: "The female's immune system is not wired to recognize the paternal transposons and hence does not repress them. As a consequence, parasitic DNA elements are now able to fulfill a new role: they can act as regulatory DNA that influences gene activity. This may have a large influence on formation and characteristics of a new species." These findings were published in Genome Biology (Elurbe et al, 2017).


Spotted salamander, Jefferson-blue spotted salamander complex, and wood frogs emerge from winter hibernation on rainy nights in March and early April, after the ground has thawed and evening air temperatures stay above 40ºF.


But why are these amphibians so frequently seen crossing the road? Migration distances to woodland pools can vary from a few hundred feet to more than a quarter of a mile. Unfortunately, because forest and wetland habitats are often disconnected by development, many migrating amphibians need to cross roads and long driveways, leading to mortality of slow-moving wildlife, even in low traffic areas.


The Amphibian Migrations and Road Crossings (AM&RC) Project enlists volunteers to find locations where migrations cross roads; document weather and traffic conditions; record migrating amphibians; and help them across the road. Since the project started in 2009, more than 850 volunteers have counted at least 46,000 amphibians and observed 20 species. They assisted 27,000 amphibians across roads during migrations and counted nearly 14,000 migrating amphibians killed by passing vehicles.


Volunteer observations are building our understanding of where mole salamanders and wood frogs are especially vulnerable during their annual migrations and where their habitats are located. This information can be used for community planning and for groups of volunteers interested in starting "crossing guard" programs for the amphibian breeding season.


Amphibians have undergone dramatic declines and extinctions worldwide. Prominent among these have been the stream-breeding frogs in the rainforests of eastern Australia. The amphibian chytrid fungus Batrachochytrium dendrobatidis (Bd) has been postulated as the primary cause of these declines. We conducted a capture-mark-recapture study over a 7-year period on the endangered Fleay's barred frog (Mixophyes fleayi) at two independent streams (30 km apart) in order to assess the stability of these populations. This species had undergone a severe decline across its narrow geographic range. Mark-recapture modelling showed that the number of individuals increased 3-10 fold along stream transects over this period. Frog detection probabilities were frequently above 50% but declined as the populations increased. Adult survival was important to overall population persistence in light of low recruitment events, suggesting that longevity may be a key factor in this recovery. One male and female were present in the capture record for >6 years. This study provides an unambiguous example of population recovery in the presence of Bd.


For the first time in a laboratory, Australian scientists have produced tadpoles more tolerant of climate change. A team from the University of Western Australia has cross-bred frogs from wetter regions with other species from drier areas to make them more resilient.


The black-eyed and gray-eyed recessive color mutants seen in several frog species lack the normal iridophores and melanophores, respectively, resulting in pale or albino frogs6,7,8,9,10,11,12,13. To create translucent frogs from these two color mutants of the Japanese brown frog Rana japonica12,14,15,16 and to clarify the mode of inheritance in the resulting see-through frog, we crossed two mutant frogs through artificial insemination. F2 offspring produced frogs with skin that remains translucent throughout life. We also observed the dermal chromatophores in the skin of wild-type, black-eyed, gray-eyed and see-through frogs with an electron microscope to examine the microstructure of the dermal chromatophores in the dorsal skin of see-through frogs. The present paper reports our experiments in detail, although a brief note on this topic has been previously published as a News-in-Brief (Nature)17. 2ff7e9595c


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