A stroll along a beach in Maryland or Delaware in the spring of the year will often bring you to the partial remains of a horseshoe crab. On first glance it looks like something from the movie Aliens, especially if the shell is turned making some of its twelve legs visible. Often only part of the shell serves as a witness that there was once an entire organism here.
Although called a crab, the Limulus polyphemus, an arthropod, does not belong with the crustacean branch of the arthropod phylum, but instead, it is more closely related to the arachnids like spiders, mites, and scorpions.
Though not very pretty, the Limulus has the distinction of age; as a species it has survived for 455 million years!
Like its arachnid relatives the Limulus has blood (more accurately, "hemolymph") that is blue instead of red. The Limulus hemolymph uses copper to transport oxygen coloring it blue when it is oxygenated.
Limulus has a heart and arteries but the blood (hemolymph) is in an open system with no veins to carry the blood back to the heart. The pumping of the heart delivers blood (hemolymph) to the tissue spaces called hemocoels that allow the animal's cells to bathe in oxygenated blood and to exchange cell wastes. When the animal moves, the blood sloshes back to the sinus (the pericardium) that surrounds the heart, passing the gills along the way to rid itself of wastes and to be re-oxygenated.
Because the Limulus has an open circulatory system that allows potential pathogens ready access to its hemolymph system, the animal has evolved a unique defense element to its immune system.
Among the potential bacterial invaders that may threaten the Limulus are Gram-negative bacteria such as Escherichia coli, Salmonella, Shigella, Pseudomonas, Neisseria, Haemophilus influenzae, Bordetella pertussis, and Vibrio cholerae which all have cell walls that contain a substance called endotoxin. When an organism is exposed to endotoxin, the immune system is stimulated to release cytokines but the cytokines can cause excessive inflammation that can lead to organ failure and death.
Patrolling the Limulus hemolymph are amoeba-like cells called amebocytes. If one of these detect the presence of endotoxin, it releases a granule that begins a protein cascade (similar to the clotting cascade in mammals) that results in the production of the protein coagulogen. The coagulogen immediately forms an insoluble gel (coagulin) around the endotoxin molecules rendering them harmless.
While the Gram-negative bacteria can be killed by high heat, the endotoxin retains it ability to cause serious illness. In addition, they are difficult to detect and can cause severe problems in even very small amounts. Contamination of medicine or medical devices by endotoxin is therefore a serious matter.
Before 1970, pharmaceutical companies kept herds of rabbits onsite to use as test animals because a sick or dead rabbit was the only reliable clue to endotoxin contamination. Then, in 1956, Frederick Bang who was interested in possible contribution from marine biology to medical science reported that the endotoxin of Gram-negative bacteria would cause the blood of the Limulus to turn into a semi-solid mass. Using this knowledge Bang and colleagues created the Limulus amebocyte lysate (LAL) test to detect endotoxin. The test is extraordinarily sensitive and is able to detect as little as a femtogram (a millionth of a billionth of a gram) of endotoxin in a milliliter of water. It is therefore now a standard test used in the pharmaceutical and healthcare industries to detect endotoxin contamination. The key ingredient in the test, the protein coagulogen, is found only in the Limulus.
To obtain sufficient quantities of the coagulogen, it is necessary to collect large numbers of the animals. The harvest comes in the spring when thousands of the Limulus emerge from the Atlantic along the Maryland and Delaware beaches to breed. The collectors use a hollow needle to penetrate the carapace to access the pericardium and drain out the hemolymph-blood. The animal is released alive back into the water.
The hemolymph is centrifuged to separate out the amebocytes which are placed in distilled water. This causes the amebocytes to burst (lyse), releasing the granules that will produce coagulogen. The granules are freeze-dried and sold to the pharmaceutical and healthcare communities.
The collected blood can bring up to $15,000 per quart to the collectors. But there is more to the story of the Limulus.
For thousands of years, the spring breeding cycle of the Limulus polyphemus has attracted the many species of birds migrating along the East Coast Flyway between South America and the Arctic breeding grounds.
The Red Knot (Calidris canutus rufa), a migratory bird that depends on the Limulus. To rest up for the flight to the Arctic, the Red Knot will spend two weeks dining on Limulus eggs, a kind of Red Knot superfood. "They're easy to find, easy to digest, and they're filled with fat, which is exactly what migratory birds need."
The Limulus is also prized by eel and conch fishermen who use the Limulus for cheap bait. Finally, add the occasional disasters like Hurricane Sandy that come blowing up the Atlantic coast scouring the beaches of the sand that the Limulus uses to cover its eggs to the pressure against the survival of the Limulus.
The story of the humble and mostly invisible Limulus polyphemus with its many connections reminds one of Robert Boyle's observation made 400 years ago that it is not possible to investigate one part of the natural world without raising questions about other parts. This connection is well-illustrated by what happens when the walker along the beach turns over the shell of a Limulus polyphemus.
“Shorebirds’ Fate Hangs on Horeshoe Crabs.” National Wildlife Federation
“What is horseshoe crab blood used for? Ask a naturalist.com
Ding, JL (2010) Endotoxin detection--from limulus amebocyte lysate to recombinant factor C. PubMed.gov
Dr. John Holton
Dr. John Holton joined the S²TEM Centers SC in July of 2013, as a research associate with an emphasis on the STEM literature including state and local STEM plans from around the nation.