Project

Envenomation by snakebite is one of the most neglected diseases with an estimated 5 million bites. These result in about 100,000 deaths and 400,000 disabilities annually [1]. The only effective treatment is animal derived antivenoms, which frequently causes adverse reactions [2]. As a result they are often only administered as a last resort.

One of the major problems related to treatment of snake bites concerns the lack of identification of the snake. When bitten, it is difficult for most people to remember details about the snake, which makes it almost impossible to confidently administer the correct specific antivenom. Currently, if the snake cannot be identified, multispecies antivenom is used in a consequently higher dose leading to more severe side effects. By solving the venom identification problem it could potentially increase the survival rate of snake bite victims. This is due to decreased diagnosis time and less amount of antivenom necessary for treatment, thereby causing fewer adverse reactions in the patient [3].

We aim to create a novel diagnostic platform by using synthetic biology derived protease activity assay to identify specific venom enzymes that, by developing suitable substrates, can investigate the relative composition of specific venom components in a blood sample. Our goal is to create a diagnostic tool that would make it possible for a clinician to quickly determine which antivenom is necessary, or if it is necessary at all.

Current detection methods

The identification of the snake that has bitten the patient is often a very hard task. Numerous molecular biology techniques such as immunofluorescence, radioimmunoassay and enzyme immunoassay have been described as suitable methods for detecting the snake responsible [4]. However, the drawbacks following these techniques (expensive, need of trained personnel and/or sophisticated equipment, long assay time) usually outweigh the advantages. For these reasons, medical staff and clinicians rely on the symptoms exhibited by the patient, in order to decide which antivenom to administer. This obviously is not optimal, as the time spent for the diagnosis could prove crucial for the survival of the patient.

Results

Our experiments showed that distinguishing between snakes of different or the same genus, using an enzymatic assay detection method, is possible. We identified numerous peptide substrates, cleavable by proteases present in the venom, that can be used to separate snake species, by incubating them with the venom. These peptides can be used as standalone, if they are uniquely cleaved by one of the venoms, or as part of a decision tree using multiple peptides for more accurate detection. We designed composite parts that would utilize these peptides, and display color when they are cleaved for easier and more robust detection. The time necessary for the detection can be significantly reduced when compared to detection methods currently available. Lastly, we developed a prototype of a device that would employ these parts, thus creating a complete and intuitive snake venom detection system.

Future

As mentioned above, our project investigated snake venom detection only as a proof of concept, using three snakes that are a huge health problem and are active in the same geographical region. Future studies would involve optimizing and continuing the tests of the composite parts that allow the identification of these snake venoms, and improving the device used as the detection system. In addition, reducing the time of detection to even less would greatly help the performance and capabilities of the system. Last but not least, continuing the research for more suitable and efficient oligopeptides and expanding the snake species of interest to more regions of the world would be of great importance.

References

[1] Chippaux JP (1998). Snake-bites: appraisal of the global situation. Bulletin of the World Health Organization, 76(5),515-24.
[2] Gutiérrez JM, León G, Lomonte B, Angulo Y (2011). Antivenoms for Snakebite Envenomings. Inflammation and Allergy. Drug Targets 10(5), 369–80.
[3] Warrell D, Gutierrez JM, Padilla A (2007). Rabies and Envenomings: a Neglected Public Health Issue: Report of a Consultative Meeting. World Health Organization, Geneva.
[4] Theakston RD, Laing GD (2014). Diagnosis of snakebite and the importance of immunological tests in venom research. Toxins (Basel), 6(5),1667-95.