Objective: Pathogenic Staphylococcus strains differ in the presence of virulence factors that are encoded mainly by mobile genetic elements, in particular by prophages integrated in the bacterial chromosomes. The study objective was to develop a method for rapid and simple characterization of S. aureus prophages.

Methods: The prophages were induced from lysogenic strains by UV-irradiation. They were picked from one plaque and propagated on a non-lysogenic strain to obtain a low titre phage lysate (10³ PFU/ml). A new method for phage DNA extraction from small volumes (150 µl) of low titre phage lysate was developed using magnetic nonporous microspheres P(HEMA-co-EDMA) and NucleoMag. The phage DNAs were characterized by multiplex PCR assays targeting capsid genes (portal and tail), genes for phage integrases, antirepressors, amidases and virulence associated genes for Panton-Valentine leukocidin, exfoliative toxin A and those of innate immune evasion cluster.

Results: Under optimized induction conditions, prophages were induced from 50 S. aureus strains. The PCR-ready DNA was isolated using new methods and amplified by PCR using newly designed primer sets. The results enabled us to divide the phages into several groups (numbers in brackets) according to capsid structure (9), integrases dictating the attachment site on the host chromosome (10), antirepressor (9), and lytic module (4). We propose updating the phage nomenclature to correspond better to the genomic loci and extensive mosaic pattern of phage genomes.

Conclusion: The rapid and simple method for DNA extraction followed by PCR based diagnosis of phage genomic modules is helpful in effective study of phage dynamics. The characterization of staphylococcal prophages is essential for understanding the role of phages in virulence and evolution of S. aureus strains.

Acknowledgements: Supported by grants LSHM-CT-2006-019064 from EU and MSM0021622415 from the Ministry of Education, Youth and Sports of the Czech Republic.