Recap of the problematic
Quagga mussels are an invasive species of mollusc that originally comes from the black sea. Since their first appearance in Switzerland in 2014, this invasive species is now found in at least 6 Swiss lakes, including Lake Geneva. By proliferating in an uncontrolled way, quagga mussels clog the pipes and cause many economic and environmental problems. For example, in the United States, quagga mussels damages cost nearly 1 billion dollars per year.In addition, quagga mussels have a great filtration capacity which result in a considerable decline in phytoplankton, algae proliferation and an alteration of the food chain .
Why we decided to target this issue
Due to its impact on biodiversity and on the economy, us, students of the University of Lausanne,decided to build our project on this topic. Our goal is to find a solution to this global problem by working hand in hand with professionals as well as people affected by this scourge. To do this, our project, named “Quagg’out”, is divided into two parts carried out using synthetic biology and genomic modification of Escherichia coli and Pseudomonas fluorescens bacteria. The first part consists in killing the mussels using the overexpressed fitD toxin and the second one consists in preventing the attachment of the mussels to surfaces using zosteric acid.
Project parts
FitD: the fitD toxin is encoded by the FitD gene and it is known for its insecticides properties, but it has also been previously proven to be effective in killing mussels. This gene is expressed naturally by the bacteria Pseudomonas fluorescens. For this part of the project 2 different strategies are pursued. One technique consists in the overexpression of fitG, the activator of fitD, in pseudomonas. In an additional attempt we overexpress directly the FitD gene in pseudomonas. For both strategies two different plasmids are used: one inducible by the activator lac and one constitutive. The mechanism of the inducible plasmids allows us to produce fitD only under certain conditions, meaning that we can control the production of the toxin and therefore kill the mussels in a more efficient way. Once the plasmids are induced, the bacteria will start to produce the toxin. These bacteria will then be lysate and put, dead, in a small tank containing the quaggas to test its effectiveness in killing the mussels.
Fig1: A) FitD constitutive. B) FitD inducible. C) FitG constitutive. D) FitG inducible
Zosteric Acid (ZA): This compound, as the name said, it’s an acid that has been previously proven to have anti- insecticide, antifungal and anti-adhesion properties; it is naturally produced by algaes. Quagga mussels attach to the surfaces in order to survive. Preventing the attachment with the use of this anti-adhesion molecule, could therefore have an impact on the lifespan of quaggas. For this part of the project we are using E. coli BL21 strain which does not contain the plasmids to produce the enzymes needed for the catalytic steps of Zosteric acid production. In fact in this case 2 plasmids are required: transporters plasmid to allow entry of SO4- (containing cysP +cysDNCQ), and the catalytic plasmid (containing the genes TAL-flojo and SULT1A1) to proceed with the catalytic reactions from SO4- to ZA. Both plasmids, being under the promoter LacI, can be induced with the use of IPTG. ZA will be released by the bacteria into the medium which will be purified to extract the component needed and then applied as a solution into a small aquarium containing the quagga mussels to test its effectiveness.
Fig2: Zosteric acid plasmids: A) and B) are two constructs for the catalytic plasmid, C) and D) are two constructs for the transport plasmid.
