Team:Wageningen UR/overview/solution

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Revision as of 14:58, 16 October 2014

Wageningen UR iGEM 2014

 

 


Solution

Engineering a bacterial platform to control the panama disease threatening banana cultivars all over the world is the main purpose of BananaGuard. A combination of a variety of strategies will prevent Fusarium oxysporum infection. Detection of F. oxysporum presence in the soil forms the first part of the BananaGuard project. Expression of four fungal growth inhibitors coupled with the sensor for fusaric acid form the second part of the BananaGuard project. Making the expression of the fungal growth inhibitors dependent on fusaric acid sensing will enable BananaGuard to have a low impact on non-target organisms. Resistance to fusaric acid is very important in this context. Even though a moderate fusaric acid resistance is endogenous for our host organism Pseudomonas putida [1] , fusaric acid resistance is engineered since the system is intended to be transferable to different host organisms.

When releasing a genetically modified organism into the soil, horizontal gene transfer as well as the risk that non-target organisms are outcompeted need to be taken into account [2]. Therefore safety forms the third part of the BananaGuard project. Horizontal gene transfer will be avoided by a double dependent plasmid system. The loss or gain of one of the two modified plasmids will cause the host to die. Additionally, a self-destructive kill switch that will be automatically activated after BananaGuard fulfilled its purpose and will protect the natural balance within the soil. Engineering a bacterial platform for biological control of F. oxysporum based on Pseudomonas putida shows great potential. This soil-borne bacterium possesses already a moderate fusaric acid resistance [3] and naturally inhibits F. oxysporum to a certain degree [4]. Since the system is designed to sense and respond to fusaric acid it is very important that the host possesses a resistance to this toxin.

BananaGuard time line.

Application

The control of soil-borne pathogens by biological control is not a new idea and has been studied since the early twentieth-century [4]. For a long time commercial applications were thought to be not feasible but, in the last decades techniques for application progressed tremendously.

For BananaGuard, we believe that distributing the final product via drip irrigation is the best method. In drip irrigation a network of pipes is used that has emission points (drippers) delivering the water in a precisely measured volume into the rhizosphere. Distribution of growth substances such as fertilizers and chemicals, such as herbicides and fungicides, has been shown to be successful before [5, 6]. In 2008, Boari et al. showed that distribution of suspensions containing small particles via drip irrigation shows great potential. When considering shelf live and transport as well as the fact that, in some growing regions, no mechanical irrigation is used, BananaGuard could be applied as dry products such as granules or powders as seen with comparable products [8]. Furthermore, these dry pelets can be stored for a longer time period. Since BananaGuard needs to be applied in a preventive manner rather than curative, usage of more dilute suspensions culture applied over a longer time period or repeated application of the dry product is conceivable. Seed coating can be considered when excluding the kill switch from the system. Colonization of the banana roots by BananaGuard at the very early seedling stage secures continuous protection.

Application of BananaGuard could possibly be combined with the application of products such as fertilizer. Since farmers generally already supplement their plants with fertilizer or pesticides, this would only add a small extra step to their current practices. The farmers can also grow the bacteria themselves with a minimal amount of materials and training, since P. putida is an easily grown bacteria as long as there is the possibility to work in sterile conditions. The bacteria can be harvested over a series of generations. However, new samples would need to be purchased to assure the quality of the product and to remove contamination that arise during harvesting.

Although the application would be simple, regulations concerning an engineered soil bacterium for bio-control are complex and different in most countries. With the current regulations in Europe, for example, application of our product would not be possible in the near future. Luckily, however, our main target area, South America and Asia have far less stringent rules about the use of GMO products. Combining the acceptance of GMO bacteria in most banana producing countries and maintaining availability of a “GMO free” banana makes our product a strong candidate for introduction into the global market.

With the rise of more highly engineered crops, whether created with traditional breeding or genetic modification, the inter-species diversity in crops grown commercially decreases. Even though it will not be as dramatic as is the case with bananas, crops will become less diverse and therefore weaker against threats they were not specifically selected for. Fusarium has been known to also infect other crops such as wheat, cucumbers and tomatoes. If successful, our system could be rapidly changed to work with other crops or even other fungus. Future applications could even include the production of plant growth promoters or increased nitrogen fixation in the soil. By combining the modularity and rapid modification time possible in bacteria but not plants, our system can be an efficient way to safeguard the crops of the future.


References

  1. Utsumi R, Yagi T, Katayama S, Katsuragi K, Tachibana K, Toyoda H, Ouchi S, Obata K, Shibano Y, Noda M. (1991) Molecular cloning and characterization of the fusaric acid-resistance gene from Pseudomonas cepacia. Agric Biol Chem. Jul;55(7):1913-8.
  2. Wright O., Stan GB., Ellis T. (2013) Building-in biosafety for synthetic biology. Microbiology (2013), 159, 1221–1235
  3. Lemanceau P. Alabouvette C. (1991) Biological control of fusarium diseases by fluorescent Pseudomonas and non-pathogenic Fusarium. Crop Protection. Vol. 10 Issue 4: 279–286
  4. Cook, R. J. (1993). Making greater use of introduced microorganisms for biological control of plant pathogens. Annual review of phytopathology, 31(1), 53-80.
  5. Hebbar SS, Ramachandrappa BK, Nanjappa HV, Prabhakar M (2004) Studies on NPK drip fertigation in Weld grown tomato (Lycopersicon esculentum Mill.). Eur J Agron 21:117–127
  6. Gollehon N (1990) Chemigation: a technology for the future? Economic Research Service, Washington
  7. Boari, A., Zuccari, D., & Vurro, M. (2008). ‘Microbigation’: delivery of biological control agents through drip irrigation systems. Irrigation Science, 26(2), 101-107.
  8. Schisler, D. A., Slininger, P. J., Behle, R. W. & Jackson, M. A. (2004) Formulation of Bacillus spp. for biological control of plant diseases. Phytopathology 94, 1267–1271.