PATHOGENICITY ISLANDS MEDIATE A NOVEL FORM OF LATERAL TRANSDUCTION

Abstract

Staphylococcus aureus pathogenicity islands (SaPIs) are a family of mobile genetic elements that exploit bacteriophages as helpers for their own production and spread. The SaPI host organism, S. aureus, has become progressively more virulent and resistant to antibiotics, and is continuously evolving. These rapid adaptation abilities imply a highly dynamic evolutionary capability likely driven by horizontal gene transfer, in which SaPIs play a key role. To date, three distinct SaPI-encoded phage interference mechanisms have been identified. These mechanisms reduce phage propagation, and are believed to promote the dissemination and persistence of SaPIs in the host population.

One mechanism, mediated by SaPI-encoded cpm, directs the production of small phage capsids that only accommodate the smaller-sized SaPI genomes and not phage genomes. Yet, despite its classification as a phage interference mechanism, previous studies reported that transfer frequencies for SaPIs lacking cpm were higher than those of their respective wild-type elements, which seemed to suggest that the mechanism is not beneficial to SaPI. To account for this contradiction, we reviewed some possible flaws and inaccuracies in the SaPI literature. Here we identified a previously unrecognised issue with small capsid production and SaPI genome unit length, where direct insertion of a resistance marker originally intended for monitoring SaPI transfer might have increased the SaPI genome size beyond the small capsid packaging capacity, resulting in defective SaPI transducing particles. Thus, we propose that SaPI transduction frequencies may have been underreported. To test this, we engineered a “size-conserved” marked SaPI, and compared the transduction efficiencies of the size- conserved SaPI with the “wild-type” marked SaPI in transduction assays. Correspondingly, we show that direct insertion of a marker indeed affected SaPI transduction frequencies adversely. Moreover, we infected S. aureus at various multiplicities of infection to model the effects of SaPI-mediated phage interference across successive rounds of phage infection, which provides evidence that SaPI-mediated phage interference mechanisms may play a key role in allowing SaPIs to persist in nature. Hence, these indicate that previous SaPI constructs were unable to provide accurate representations of the dynamic relationship between SaPIs and phages in SaPI-mediated phage interference studies.

Furthermore, genomic studies have revealed extensive variation in natural populations of S. aureus. The S. aureus core genome is known to contain large spans of highly variable regions (20-30kb) often enriched in virulence genes. Many of these regions are also found immediately adjacent to SaPIs, or their attachment sites, though a connection between the SaPIs and these variable regions has never been established. In this portion, we sought to elucidate the underlying molecular mechanism that drives the evolution of these distinct regions in the S. aureus genome. Accordingly, we identified a novel mechanism of horizontal gene transfer mediated by the SaPIs. We report that this mechanism promotes the high frequency exchange of chromosomal regions adjacent to known SaPI attachment sites between S. aureus strains. Next, we showed that SaPIs can also initiate packaging and transfer from other sites in the chromosome by integrating into sites other than their bona fide attachment sites. Taken together, we propose that these mechanisms promote genetic recombination in S. aureus, and accounts for some degree of the remarkable levels of genomic plasticity of this highly successful pathogen.