Paleozoic Paladin: Horseshoe Crabs and their Contribution to Vaccine Research

Hello, again, loyal MütterEDU readers. Editor Kevin here to introduce another piece from one of our amazing guest writers. Center for Education educator Joanna D'Avella prepared this article on the humble horseshoe crab and its huge contributions to vaccine research.

Does a prehistoric animal hold the key to worldwide vaccination?

With its bright blue blood, (Limulus polyphemus) are the unsung hero of vaccine development in the 21st century as researchers tap into their evolutionary immune system to save the lives of millions. 

How Does a Vaccine Get Approved?

According to the , only after rigorous amounts of testing throughout a variety of stages will a vaccine warrant FDA approval. These stages include the exploratory stage, pre-clinical stage, clinical development, regulatory review and approval, manufacturing, and quality control.

Some stages, similar to the development and manufacturing of the COVID-19 vaccine, can be done in conjunction with each other. This typically happens under an emergency authorization. The completion of stages in conjunction with one another does not reduce the efficacy of the vaccine due to the level of testing that is done prior to distribution.  

Why Do We Need to Test Vaccines on Animals? 

It is imperative that scientists create an environment that mimics that of a human body without placing humans in danger. As a result, scientists use animals as a safer alternative to human testing. Mice, monkeys, dogs, cats, and rabbits were all viable options for scientific research for hundreds of years, especially in early medical history when human dissection was illegal in most countries. Rabbits are the most well-known animal for animal testing, especially during the development of the polio and rabies vaccines. Researchers would use the rabbit pathogen test to confirm if pathogens were present in the vaccine. These animals gave scientists a safer alternative than using human subjects in the early stages of the vaccine development. 

Why Horseshoe Crabs?

Horseshoe crabs are an old species, having been around for over 300 million years. As one of the last connections to the prehistoric era, horseshoe crabs developed disease-prevention measures to maintain their population. This begins with their open circulatory system and their bright blue blood. In “The Role of Horseshoe Crabs in the Biomedical Industry and Recent Trends Impacting Species Sustainability,” the authors summarize the incredible process horseshoe crab blood undergoes when introduced to toxins. 

“When these granular amebocytes come into contact with an endotoxin or 1,3ß-D-glucan (present in the cell walls of Gram-negative bacteria and fungi, respectively), a cascade of defense molecules is released, triggering coagulation and neutralization of the pathogens. The resulting clot effectively immobilizes the threat and prevents an infection from progressing beyond the wound (Isakova and Armstrong, 2003).”

Krisfalusi-Gannon, J., Ali, W., Dellinger, K., Robertson, L., Brady, T. E., Goddard, M. K., Tinker-Kulberg, R., Kepley, C. L., & Dellinger, A. L. (2018). The role of horseshoe crabs in the biomedical industry and recent trends IMPACTING Species Sustainability. Frontiers in Marine Science, 5. 

Clotting helps prevent the toxins from spreading throughout the animal and making them extremely ill. Once researchers understood they could harness this evolutionary process, they have been using it ever since. 

During the 1970s, researchers began using horseshoe crab blood to test for . Endotoxins are a toxin present in the outer membrane of cells that are released once the cell begins to decompose or is destroyed. Once released and introduced outside the cell, these toxins will then present symptoms of disease, making the organism very sick. In hopes of removing this potential threat, medical professionals devised tests to prevent these endotoxins from entering the general public through medical equipment or vaccines that had been contaminated. This was typically done using animals such as rabbits but at the cost of the test subject. Researchers monitored the rabbits for symptoms after injection; if none manifested, then medical equipment or a batch of vaccines were considered safe. However, if the rabbit--similar to a canary in a coal mine--died shortly after being exposed, it was deemed unsafe. Once researchers understood the potential of the horseshoe crab blood, they developed the limulus amebocyte lysate (LAL) which according to a 2010 article in the , “in general were 3 to 300 times more sensitive than the United States Pharmacopeial (USP) rabbit pyrogen test method.” This was a scientific breakthrough for vaccine development. It was a far more effective test that did not necessarily mean the loss of animal life. However this is debatable because  This test effectively replaced the rabbits, and put these Paleozoic paladins on the frontline of medical developments. 

COVID-19 and Horseshoe Crabs

As vaccines are developed all over the world to try and to stop the spread of COVID-19, horseshoe crabs and their blood are being used more than ever due to the sensitivity needed for the LAL test. This species was already under ecological strains due the environmental changes, overfishing, and overuse in synthesis of the LAL test. It is true that researchers do release horseshoe crabs after their blood has been collected, however, the survival rate is not high enough to sustain the current population. There is hope, however, as that will potentially yield similar results.

Sources

• Akbar John, B., K.C.A Jalal, Y.B. Kamaruzzaman, and K. Zaleha. Journal of Applied Sciences 10, Iss. 17 (2010). 
• Cox, Elizabeth. Ted-Ed (September 21, 2017).
• Dhnesha, Neel. Audubon. November 20, 2020. Accessed September 7, 2021.  
• Esteves, P.J., Abrantes, J., Baldauf, HM. et al. The wide utility of rabbits as models of human diseases. Exp Mol Med 50, 1–10 (2018).
• Krisfalusi-Gannon, J., Ali, W., Dellinger, K., Robertson, L., Brady, T. E., Goddard, M. K., Tinker-Kulberg, R., Kepley, C. L., & Dellinger, A. L. (2018). . Frontiers in Marine Science, 5. 
• Merriam-Webster. (n.d) . In Merriam-Webster.com dictionary. Retrieved August 17, 2021.
• Pavid, Katie (3 December 2020, Updated: 15 January 2021) Natural History Muse.
• (2021) Center for Disease Control. https://www.cdc.gov/vaccines/basics/test-approve.html