Project Description

Cardiovascular diseases are medical conditions associated with blockage of the coronary vessels leading to heart organ failure [1]. According to the World Health Organisation, cardiovascular diseases are the leading cause of death globally, with an estimated death of 17.9 million in 2019 total, these numbers represent 32% of all global death, making CVDs statistically the most potent death causing health condition in the human species [1]. However, these conditions are not specific to the human species and can be found across the mammalian class [2]. Under certain environmental and physiological circumstances fish can also develop fatal cardiovascular conditions as referred to by A. C. Smith in his findings [3]. The environmental and physiological conditions contributing to the development of CVDs in fish are discussed by Casillas and Smith in their article [4].

Antithrombotic treatments often feature a use of mixed medicine such as thrombolytics, antiplatelets and anticoagulants, all which function as antithrombotic factors in some way regarding the clotting mechanism [5]. Leeches secrete biologically active antithrombotic substances to allow ease of feeding, one of which is the molecule hirudin, these molecules can serve valuable medical and pharmaceutical purposes in antithrombotic treatments sided with would healing effects [6].

Our team was intrigued by the fact that fish also suffers from CVDs [4][5] and that it might be linked to the unreasoned percentage death within the fish farm industry. There are no clear indications or symptoms that can be observed by fish farmers for identification of fish developing internal coagulation that differentiates this cause of death from regular illnesses that occurs due to abiotic and biotic factor interactions of a fish farm. In both cases fish present a loss of appetite and decreased activity [Learn more in HP segment (HK visit)].

As hirudin molecules extracted from leeches have been studied and applied in treatment for CVD for human patients during the late 19th centuries and early 20th centuries [7], our team were interested to investigate whether the antithrombotic effects of hirudin is also effective in the treatment of fish CVD.

We are biologically synthesising the molecule hirudin and its homogeneous molecules found across different species of leeches. Hirudin genes of 12 different leech species were identified from GeneBank and the literature, these genes will be synthesized by a third-party company under the T7 promotor. Our team will assemble the commercially obtained sequences into the pSB1C3 backbone as provided in the 2024 iGEM Distribution Kit along with the GFP gene (Without LVA/Bsal sites) as a marker, also obtained from the 2024 iGEM distribution kit. The newly assembled plasmids will be transformed into DH5a-competent cells for culturing. Hirudin protein will be extracted from the E. Coli cells and used to perform blood clot assays in fish blood. Fish blood will be obtained from the wet market, collected as side product when fish is slaughter for consumption by humans. Antithrombotic effectiveness, when presented with fish blood, will be compared between the hirudin from the different leech species. If these assays verify that antithrombotic features of hirudin are present in fish blood, the comparison will allow us to identify the most effective hirudin variant for said purpose which will be used in the future stage of our project development.

Once we have identified the leech species that produces the hirudin that is the most effective for use as anticoagulant in fish blood, the hirudin from that species can be developed further to treat fish suffering from cardiovascular disease during the process of fish farming. The visioned future development of the project would be the construction of a delivery system regarding a fish farm setting and a way to identify internal coagulation in fish for early prevention.