I recently participated in a partnership to save an incredible species of amphibian: the blue spotted salamander (Ambystoma laterale). This animal is regionally endangered in parts of the Midwest and about 10 years ago, the Iowa Department of Natural Resources started a partnership with Omaha’s Henry Doorly Zoo. They wanted to breed these salamanders for release at locations where they once thrived but have now disappeared. Having had little success in doing so thus far, the organizations reached out and requested my amphibian “sexpertise”. I travelled to Omaha to deploy some of my special reproductive techniques with the blue spotted salamander assurance colony living there.

It comes as no surprise that these salamanders proved very difficult to breed. Blue spotted salamanders need an extraordinary amount of “salamander romance” in order to breed naturally. These animals undergo winter brumation (a hibernation-like state). In the spring, they will emerge from brumation when it begins to rain and when the temperature rises in order to migrate to breeding ponds. Their migrations can be over several miles, and most salamanders return to the same pond every breeding season. In captive breeding situations, we attempt to recreate the cues that the salamanders take from the wild in order to get them “in the mood” for breeding. We cool them down and offer them the opportunity to undergo brumation. We place them in rain chambers so they can experience an “indoor storm”. We also attempt to create a naturalistic pond for them to breed in with proper vegetation and leaf litter.

Sometimes, despite our best efforts, the natural cues we provide artificially are not enough to trigger breeding. Many amphibians are having difficulty breeding in the wild, due to changes in the climate and their habitat. Given that the natural environment is not currently providing ideal cues for breeding, it is especially difficult to perfectly recreate environmental breeding cues for animals in human care. When captive animals do not breed in response to the cues we provide, we often use hormone treatments to give them a little boost. These hormone protocols are meant to trigger breeding behaviors as well as the release of sperm and eggs. In the best-case scenario, the application of hormones will result in the animals breeding on their own. In the worst-case scenario, the animals will not breed on their own after the application of hormones and an in vitro fertilization (an artificial fertilization of the eggs with the sperm in a petri dish) will take place.

We used a combination of natural cues and hormones to stimulate breeding in the blue spotted salamanders. The amount of hormones I gave to the female blue spotted salamanders was determined by how developed their eggs looked on ultrasound. While giving the animals hormones resulted in many interesting behaviors, the salamanders did not breed on their own. After giving them time to breed on their own without success, I attempted in vitro fertilization. In order to perform this technique, I needed to collect eggs from the females and sperm from the males. There was one big problem with attempting this technique – the male blue spotted salamanders were not producing sperm. Fortunately, I had a back-up plan!

The blue spotted salamander is no ordinary salamander. In the wild, many female blue spotted salamanders breed through a special adaptation called kleptogenesis, meaning that they steal sperm. You read that correctly: sperm thieves. Most salamanders undergo internal fertilization. This occurs when the male deposits the sperm in a ball – called a spermatophore – into the environment. Through a courtship dance, the spermatophore is collected by the female. These special female blue spotted salamanders will find and collect spermatophores deposited by males from other, closely related, species (including Jefferson’s salamanders, tiger salamanders, and small-mouth salamanders). This is the part where it becomes complicated. Rather than using the sperm to fertilize their eggs and have hybrid offspring, an enzyme in the sperm activates the egg which begins to grow into an embryo. The rest of the sperm is discarded, and the offspring which develop are essentially clones of the mother. Knowing that blue spotted salamander females are often sperm thieves, I had the Omaha Zoo’s resident male tiger salamanders (Ambystoma tigrinum) on standby. I was able to collect sperm from the tiger salamanders in order to use in the in vitro fertilization of the blue spotted salamander eggs.

Unfortunately, the eggs did not fertilize with the tiger salamander sperm. While we did not achieve offspring, we learned what we need to do differently next time around. We certainly will not give up on these amazing animals. We will try to breed the blue spotted salamanders again later this year, using slightly different techniques.

Thank you to our partners at Omaha’s Henry Doorly Zoo and the Iowa DNR.

– Dr. Ruth Marcec is the director of the National Amphibian Conservation Center at the Detroit Zoo.