Phages expressing fluorescent proteins as tools to unveil the mysterious phage-biofilm interaction

Laura Lisá, Anna Sobotková, Pavel Plevka

Central European Institute of Technology, Masaryk University, Kamenice 753/5, 625 00 Brno, Czech Republic

One of the major worldwide health concerns is the increasing occurrence of antibiotic-resistant bacteria. Furthermore, bacteria have the ability to form biofilm, a multicellular structure embedded in a polymer matrix. Biofilm enhances bacterial resistance to the immune system and antibiotics. Biofilm eradication is a challenge which opens an opportunity to develop new approaches such as phage therapy. [1,2]

We focus on visualisation of infection of Staphylococcus aureus biofilm by bacteriophage phi812. The objective is to genetically modify phage phi812 so that it expresses fluorescent proteins in infected cells for the purpose of observation by fluorescence microscopy.

The strategy of the genetic modification lies in the homologous recombination of the phage genome with a vector containing a coding sequence for a fluorescent protein. Bacterial defence mechanism CRISPR-Cas10 will be used for targeting the unmodified regions of the phage genome.[3]

[1]    F. Reffuveille et al., Staphylococcus aureus Biofilms and their Impact on the Medical Field. IntechOpen, 2017. doi: 10.5772/66380.

[2]    F. L. Gordillo Altamirano and J. J. Barr, ‘Phage Therapy in the Postantibiotic Era’, Clinical Microbiology Reviews, vol. 32, no. 2, pp. e00066-18, Jan. 2019, doi: 10.1128/CMR.00066-18.

[3]    S. M. Nayeemul Bari, F. C. Walker, K. Cater, B. Aslan, and A. Hatoum-Aslan, ‘Strategies for Editing Virulent Staphylococcal Phages Using CRISPR-Cas10’, ACS Synth Biol, vol. 6, no. 12, pp. 2316–2325, Dec. 2017, doi: 10.1021/acssynbio.7b00240.