Although the hellbender has been identified as a Species of Greatest Conservation Need by the Northeast Association of Fish and Wildlife Agencies, there remain substantial data gaps in its distribution. The common mudpuppy shares a significant portion of its habitat with the hellbender, and has been identified as a Species of High Conservation Concern by the Northeast Partners in Amphibian and Reptile Conservation. Given the habitat overlap of these two species, efforts to detect hellbenders may concurrently generate samples or data that are useful in monitoring mudpuppy populations. Our primary objectives are to 1) better document hellbender distribution in the northeast region, and 2) develop standardized methodologies to monitor hellbender populations while collecting opportunistic information about mudpuppy distribution. These objectives will be achieved through stream surveys (including environmental DNA detection), improved communication among individuals working with hellbenders or mudpuppies, and the establishment of a regional stakeholder working group. Within the first year of the project we will produce standardized protocols that ensure the consistency and efficiency of hellbender/mudpuppy surveys while minimizing disturbance of stream boulder habitat. During this time we will also collect environmental DNA (eDNA) samples from a total of ~130 sites in NY, PA, MD, WV, and VA. Samples will be tested for hellbender DNA and archived for future DNA-based detection of mudpuppies or other stream species. In the second year, we will employ conventional surveys to ‘ground-truth’ a subset of eDNA sites. This approach will generate presence/absence data for a broad geographical area and information about abundance, demographics and animal health for a key subset of sites. Project deliverables include 1) a more comprehensive map of hellbender distribution in the northeast, 2) an eDNA archive (for detection of other stream-dwelling species) and 3) a protocol and communication framework to enable coordinated and efficient conservation of hellbenders and mudpuppies.
The eastern hellbender (Cryptobranchus alleganiensis alleganiensis) is declining in many parts of its range  and has been identified as a Species of Greatest Conservation Need by the Northeast Association of Fish and Wildlife Agencies (NAFWA). The species’ historic range in the northeast includes New York, Pennsylvania, Maryland, West Virginia and Virginia. Despite a significant amount of research effort, substantial gaps remain in our knowledge of the hellbender’s current distribution, particularly in NY, PA and VA. Given the broad distribution and cryptic nature of this species, generating a comprehensive distribution map is challenging using traditional approaches. Conventional hellbender surveys rely on rock-turning, which is time-intensive, physically demanding, and potentially destructive to the species’ microhabitat. In contrast, environmental DNA (eDNA) surveys can provide information about species occurrence (and potentially abundance) without disturbing sensitive habitat . Such information is urgently needed to guide ongoing efforts to protect and restore wild hellbender populations.
Original protocol by J. Greathouse
- Following collection of eggs, the eggs should be transported in a covered container with oxygenated water back to the Hellbender Conservation Center.
- Upon arrival at the Hellbender Conservation Center, each egg should be separated from the clutch by cutting the gelatinous membrane between each egg with scissors while a plastic spoon is under the egg to be separated.
- Create hellbender water/methylene blue mix by adding 0.5 cc of methylene blue to a new gallon of spring water and shake well. (Methylene blue is a color indicator of oxygen level. Water will turn green/yellow if oxygen levels drop.)
- Once the egg has been separated from the clutch, place it into a specimen cup with 60 mL of spring water and methylene blue mix.
- Label the cup with the site name, treatment (shaker, incubator, or room temperature), and its individual ID number.
- While in the cup, inspect the egg for any signs of white cottony fungus and abnormalities in the texture or color of the yolk or yolk sac.
- Using the Harrison chart of larval salamander development, determine the approximate stage of the egg and record this on the appropriate form.
- Place as many cups of eggs that will fit on one half of the shaker platform in the shaker. Set the shaker temperature at 14°C and at 40 revolutions per minute.
- Place the remainder of the cups on the top shelf of the incubator with the temperature also set at 14°C. If space in the shaker and the incubator becomes filled, then specimen cups will be stored in cabinets or in empty aquaria that are covered to prevent light disruption.
- Each day, the egg should be gently removed from the cup with a plastic spoon, and the water and methylene blue should be changed at the same volume as listed above.
- Following water changes, eggs should be staged and the data should be recorded on the appropriate form. Any information such as the presence of fungus or abnormalities in the texture or color of the yolk or yolk sac should be recorded as well.
- Upon hatching of the larvae, the individual should still be removed by spoon until it is too large to handle appropriately with the spoon. At that point, the individual should be placed in the specimen cup lid or a coffee cup lid during water changes.
- Developmental staging of the larvae and inspection for yolk sac abnormalities should continue at this point and should continue to be documented.
- One blackworm per individual will be introduced as prey approximately 3 weeks following hatching, dependent upon yolk sac absorption.