Understanding a novel mechanism involving pathogenicity islands in the transfer of unlinked chromosomal virulence genes.

In recent decades, most pathogens have become progressively more contagious, more virulent and more resistant to antibiotics. This implies a rather dynamic evolutionary capability, representing a remarkable level of genomic plasticity, most probably maintained by horizontal gene transfer (HGT). HGT can, in a single step, transform a benign bacterium into a virulent pathogen. This is especially true for several notorious pathogens, including Staphylococcus aureus and Escherichia coli, used in this project as models, for which phage-mediated HGT enables relatively benign strains to cause lethal infections. However, in spite of their relevance, the mechanisms underlying transfer of MGEs and emergence of new bacterial virulent clones are far to be completely known.
Here we report the existence of a hitherto unrecognized attribute of pathogenicity islands, namely their ability to contribute to the enhancement of their hosts' pathogenicity independently of their own direct role. We have found that throughout the host chromosome are scattered homologs of the Staphylococcal pathogenicty island (SaPI) pac site, which are recognised by the SaPI coded TerS, leading to the encapsidation and high frequency transfer of chromosomal segments downstream of these pseudo-pac sites. Since SaPIs are very common in staphylococci, and since they encode terminases with different sequence specificities, they are thus capable as a genre, of promoting the exchange of alleles and the acquisition of genes that are important for pathogen adaptation. Although pathogenicity islands can convert an innocuous organism into a deadly pathogen, this is based entirely upon the virulence genes that they carry. The ability of pathogenicity islands to contribute to the pathogenicity of their hosts by directing the transfer of unlinked virulence genes is unprecedented. We note finally, that SaPI-like elements are very widespread, especially among Gram-positive pathogens, and suspect that these may well be capable of the same behavior. Here we will try to decipher the molecular basis underlying this novel mechanism of SaPI-mediated horizontal gene transfer, as well as demonstrate its broad distribution and relevance in the bacterial world.

In recent decades, the notorious pathogen Staphylococcus aureus has become progressively more contagious, more virulent and more resistant to antibiotics. This implies a rather dynamic evolutionary capability, representing a remarkable level of genomic plasticity, most probably maintained by horizontal gene transfer. In this project we will analyse the hypothesis that the pathogenicity islands (PIs) have an unprecedented dual role in gene transfer: they not only mediate their own transfer, but they independently direct the transfer of unlinked chromosomal segments containing virulence genes. While transfer of the islands itself requires specific helper phages, our preliminary results indicate that transfer of unlinked chromosomal segments does not, so that potentially any pac phage will serve. These results reveal that PIs can increase the horizontal exchange of accessory genes associated with disease, shaping pathogen genomes beyond the confines of their attachment sites. In this project we will try to decipher the molecular basis underlying this novel mechanism of SaPI-mediated horizontal gene transfer, as well as demonstrate its broad distribution and relevance in the bacterial world.

The development of novel hypervirulent strains of formerly avirulent or only weakly virulent strains is dramatically fueled by the acquisition of mobile elements carrying virulence factors - a case in point is the emergence of the notorious O157:H7 strains of E. coli and their relatives, whose genomes are nearly twice as large as those of the common garden varieties of E. coli, and the increase consists entirely of acquired mobile genetic elements (MGEs). Closer to home, staphylococcal and streptococcal superantigens are clearly related and have recently had a major role in several fulminant diseases (necrotizing fasciitis, necrotizing pneumonia, etc.). Many of the genes encoding these are carried by highly mobile elements, and we would not be surprised to discover that intergeneric transfer of these genes has had an important role in the development of superantigenic strains. Even more to the point is the emergence of the hypervirulent community-acquired MRSA, typified by USA300 and its relatives, whose hypervirulence is largely attributable to virulence-enhancing genes acquired via the horizontal transfer of MGEs. In spite of its relevance in bacterial pathogenesis, the mechanisms underlying gene transfer among bacteria remain, in most cases, unidentified.
In this project we will try to establish novel pathways by which bacteria exchange genetic information. Although this project is a basic science project, and as such may not immediately result in translational output, however, understanding how bacterial pathogens exchange genetic information and adapt to new hosts is essential if we are to both predict and model the spread and emergence of new virulent clones. Consequently, the expected impact of this project is broad and involves the following areas:
- This project will contribute to the progress of maintaining health and treating diseases by generating a highly needed knowledge base concerning the principles and consequences of MGE transfer in three important pathogens, Staphylococcus aureus, Enterococcus faecalis and Escherichia coli, and by facilitating the transfer of this knowledge to the human and veterinary clinicians.
- Combating infections: Since some antibiotic treatments increase the transfer and spread of MGE-encoded virulence, the identification of molecules that could block the packaging and transfer of virulence genes will prevent the apparition of new virulent clones.
- Phage therapy: This project highlights that there are unexplored mechanism of phage- and pathogencity island-mediated gene transfer that have to be characterised in order to use safely phages to combat infections. Otherwise, some phage treatments could even facilitate the emergence of novel virulent clones.
- Currently, >75% of bacteriophage and pathogenicity island genes are annotated as hypothetical. This application thus responds to the generally recognised need to translate genome data, and the latest developments in Systems Biology into sustainable practical applications for medical and veterinary research and treatment.
- This project will unravel a fundamental understanding of the link between MGEs and disease. Besides S. aureus, E. faecalis and E. coli, this is of general value, because MGEs play a central role for many pathogenic microorganisms.
In summary, a better understanding of the biology of the different MGEs involved in bacterial virulence is urgently required (i.e. the 2011 Escherichia coli O104:H4 outbreaks that originated in Germany and spread to other European countries). The data generated by the experiments proposed here will provide more information on the mechanisms underlying the emergence, spread and emergence of novel bacterial pathogens.