RESUMO
Introduction: Thermosynechococcus elongatus BP1 is a thermophilic strain of cyanobacteria that has an optimum growth at 57°C, and according to previous analysis by Yamaoka et al, T elongatus BP1 cannot survive at a temperature below 30°C. This suggests that the thermophilic property of this strain may be used as a natural biosafety feature to limit the spread of genetically engineered (GE) organisms in the environment if physical containment fails. Objective: To further explore the growth and survivability range of T elongatus BP1, we report a growth and survivability assay of wild-type and GE T elongatus BP1 strains under different conditions. Methods: Wild-type and GE T elongatus BP1 cultures were prepared and incubated in the laboratory (high temperatures and constant light source) and greenhouse conditions (lower/varied temperatures and sunlight) for 4 weeks. The cell density was monitored weekly by measuring the optical density at 730 nm (OD730). To assess the survivability, a sample of each culture was added to fresh media, placed in laboratory conditions (42.2°C and 30 µE m-2 s-1) in multi-well plates and observed for growth for up to three weeks. Lastly, the number of viable cells were determined by plating a diluted sample of the culture on solid media and counting colony-forming units (CFU) after 1 day, 2 weeks and 4 weeks of incubation in laboratory or greenhouse conditions. Results: Our experimental results demonstrated that growth was hindered but that the cells did not entirely die within 2 to 4 weeks at warm temperatures (31.42°C-36.27°C). The study also showed that 2 weeks of exposure to cool temperature conditions (15.44°C-25.30°C) was enough to cause complete death of GE T elongatus BP1. However, it took 2 to 4 weeks for the wild-type T elongatus BP1 cells to die. Conclusion: This study revealed that the thermophilic feature of the T elongatus BP1 may be used as an effective biosafety mechanism at a cool temperature between 15.44°C and 25.30°C but may not be able to serve as a biosafety mechanism at warmer temperatures.
RESUMO
Synthetic biology with genetically engineered (GE) cyanobacteria has the potential to produce valuable products such as biofuels. However, it is also essential to assess the potential risks of synthetic biology technology before it can be widely used. In order to address key concerns posed by the application of synthetic biology to microorganisms, studies were designed to monitor the horizontal transfer of engineered genes from GE cyanobacteria Thermosynechococcus elongatus BP1 to Escherichia coli through co-incubation. The results of these experiments demonstrated that the genetically engineered DNA construct containing alcohol producing genes and kanamycin resistance can be horizontally transferred from GE T. elongatus BP1 to wild-type E. coli following two days of liquid co-culturing. The rapid and facile transfer of foreign genes, which include antibiotic resistance, between bacterial communities signifies the need to continue to deepen our understanding of the process of horizontal gene transfer, chromosomal integration as well as further biosafety-oriented research efforts. In the era of synthetic biology, the natural microbial process for sharing genetic material will also significantly impact risk assessments, containment approaches and further policy development.