Team:Stockholm/Outlook

Outlook

Our proof of concept has been generated in E. coli, since it is a well-characterized bacteria suitable for engineering. It became clear to us that the next step would be to transfer this genetic construct to a strain more suitable for a lung probiotic.

Our basic selection criteria was based on two premises: the selected bacteria should have a shuttle vector compatible with E. coli, and it should be one of the species comprising the lung microbiome. The use of shuttle vectors (plasmids compatible with two different species), allows us to extrapolate our cellular machinery and transfer it to a new bacterial chassis.

After reviewing the available shuttle vectors on the iGEM Registry of Standard Biological Parts we came down to two strong candidates: Pseudomonas aeruginosa and Staphylococcus aureus.

Although both organisms are well defined within the lung microbiota and have available shuttle vectors, the higher compatibility of P. aeruginosa for genetic manipulation would make it a better choice. Nonetheless there is a major drawback for the use of these organisms, which is their pathogenicity. Our initial strategy to cope with this limitation would be to begin with a lung-adapted isolate of P. aeruginosa that does not produce virulence factors.

Another option within this approach could be the use of the part BBa_J153000. This shuttle vector was originally designed to work on cyanobacteria and holds the potential to work with a broad range of gram negative bacteria (Meyer, 2009). This would allow us to attempt to use non-pathogenic gram negative strains from predominant genus such as Fusobacteria or Veillonella (Beck, 2012).

Considering the fact that there are ongoing advancements in genetic engineering, we could consider another approach with a lesser focus on shuttle availability. A great example of this would be the use of S.carnosus, a non-pathogenic strain for which a shuttle vector is currently under development and has successfully expressed hyaluronate lyase from E. coli (Williams, 2002).

References

  1. Beck, J., Young, V. and Huffnagle, G. (2012). The microbiome of the lung. Translational Research, 160(4), pp.258-266.
  2. Meyer, R. (2009). Replication and conjugative mobilization of broad host-range IncQ plasmids. Plasmid, 62(2), pp.57-70.
  3. Williams, R., Nair, S., Henderson, B., Holland, K. and Ward, J. (2002). Expression of the S. aureus hysA gene in S. carnosus from a modified E. coli–staphylococcal shuttle vector. Plasmid, 47(3), pp.241-245.

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