Molecular insight into how MRSA is becoming increasingly dangerous

Why does methicillin-resistant Staphylococcus aureus (MRSA) remain a dangerous pathogen, particularly in hospital-associated infections? In part, this is due to the fact that MRSA infectious outbreaks occur in “epidemic waves,” meaning that this pathogen constantly produces new clones that are better adapted than previous ones to infect people and/or persist in the hospital environment. However, we only have a very limited understanding of the molecular processes and determinants underlying the rise of new, better-adapted MRSA clones. In a recent study, Li et al. describe a novel surface protein that promotes MRSA colonization and virulence and is transferred via a mobile genetic element. Notably, they provide evidence for a significant spread of that protein among infectious MRSA isolates, analyzing a large series of MRSA hospital infections in China. These findings shed light on the molecular rearrangements occurring in MRSA that lead to the constant creation of more successful and dangerous MRSA clones.

Staphylococcus aureus is a major human pathogen that causes frequent and often severe infections throughout the world. What makes S. aureus infections difficult to treat is the fact that many S. aureus strains are resistant to antibiotics. Most importantly, S. aureus has become increasingly resistant to methicillin, an antibiotic of first choice against S. aureus infections. Methicillin is a penicillinase-resistant penicillin derivative that was developed as a response to the global spread of the penicillinase gene among S. aureus strains, which started in the 1950s. However, methicillin-resistant S. aureus was detected only about one year after the introduction of methicillin into clinical use; and nowadays many countries report methicillin resistance rates among invasive hospital isolates of S. aureus that reach and exceed 50%. More recently, MRSA has also appeared in the community (community-associated MRSA, CA-MRSA), posing an additional threat to public health systems. Nevertheless, hospital-associated cases still represent the by far greater source of morbidity and death among MRSA infections.

Over time, we have seen the disappearance of the original 1960s and the surge of novel MRSA clones, which was accompanied by the appearance of new types of SCCmec, the mobile genetic element that harbors the methicillin resistance gene, mecA. The “archaic” 1960s clone was first found in the United Kingdom and had SCCmec type I. Infections with that clone were limited mostly to Europe. The hospital-associated (HA-) MRSA clones that caused the worldwide MRSA pandemic that started in the 1980s have SCCmec types II and III. These clones still are among the most frequently isolated HA-MRSA clones today and belong to only about five clonal groups. CA-MRSA isolates typically have SCCmec types IV or V. These types are smaller compared with other types of SCCmec, cause less of a fitness cost, and thus likely contribute to the capacity of CA-MRSA to spread and infect healthy people. The molecular basis of virulence and spread of MRSA has recently received much attention. However, it has remained largely unknown why MRSA infections occur in “epidemic waves,” or more specifically, which molecular factors contribute to the rise and epidemiological success of novel HA-MRSA clones.

In contrast to CA-MRSA, extraordinary virulence is not necessarily considered the major driving force behind the success of HA-MRSA. In fact, despite being less aggressive and showing lower virulence characteristics in animal infections models than CA-MRSA isolates, HA-MRSA clones such as USA100 continue to be a major cause of hospital-associated infections. This suggests that molecular factors other than only such contributing to aggressive virulence are behind the epidemiological success of HA-MRSA.

What makes HA-MRSA clones so well adapted to the hospital environment and hospital-associated infections? It has frequently been proposed that HA-MRSA clones thrive in the hospital environment due to an exceptional capacity to colonize patients and hospital personnel. S. aureus predominantly colonizes the nose and many molecular factors involved in nasal colonization have been identified. However, researchers have not yet been able to link a colonization factor to the spread and success of an MRSA clone.

The importance of a recent study by Li et al., who have identified a novel S. aureus surface protein, SasX, that promotes nasal colonization, lies therefore not only in the molecular and functional analysis of that new protein, but mostly in the evidence they provide for the spread of the corresponding gene among HA-MRSA infective isolates. Evaluating data from three large teaching hospitals in eastern China, they showed that the frequency of sasX-positive HA-MRSA infections increased significantly over the last decade. The sasX gene was linked mostly to HA-MRSA strains belonging to sequence type (ST) 239, the most frequent ST in most parts of Asia. Thus, SasX may contribute to the high prevalence of ST239 among hospital MRSA isolates in this geographical area. Interestingly, no ST239 clones were found among community isolates from healthy individuals in the same region, underscoring that this ST is specifically adapted to the hospital environment. These findings shed light on the epidemiology of MRSA in Asia, which is still not as well understood as that in the US or Western Europe. More importantly, however, they exemplify how a colonization factor may contribute to the epidemiological success of a newly arising MRSA clone.

The sasX gene is part of a large mobile genetic element (MGE), namely a ΦSPβ-like prophage of 127 kb. In accordance with the capacity of prophages to be mobilized and transferred to other strains, Li et al. observed a more recent spread of sasX to STs other than ST239. The transfer of a surface protein gene promoting colonization represents a quite unique MGE-associated feature, as compared with the more common transfer of toxin or antibiotic resistance genes via such elements. Notably, it may significantly enhance the recipient strains’ capacity to colonize patients and spread in the hospital environment. In fact, animal colonization studies performed with the frequent sasX-recipient ST5 suggest this is the case. Naturally, the large prophage contains many genes that are transferred together with sasX. However, the lack of other obvious pathogenesis-associated genes in the prophage indicates that the spread of the phage is due to a major extent to sasX and its properties. Furthermore, it is interesting that the sasX gene is at the very end of the prophage, suggesting that it may have been included in the prophage by incorrect phage excision during evolution. It needs to be added that the prophage contains some antibiotic resistance genes potentially conferring resistance to aminoglycoside antibiotics. These may theoretically also play a role in the spread of the prophage. However, supporting the importance of sasX for the spread of ST239 and the distribution of the prophage among MRSA clones, Li et al. found that some strains had prophage deletions in the region containing antibiotic resistance genes, while the sasX region was maintained.

As for the mechanisms underlying the phenotypes associated with sasX, the authors showed that nasal colonization is due to an enhanced capacity of sasX-positive clones to adhere to human nasal epithelial cells. Adhesion could be blocked with purified SasX protein in a competitive fashion, indicating a specific interaction. Interestingly, sasX-positive clones also revealed enhanced virulence in animal lung and skin infection models. This is surprising, given that virulence-promoting properties are usually associated with toxins rather than colonization factors. As a plausible explanation for the observed enhanced virulence, the authors found increased capacity of sasX-positive clones to evade neutrophil phagocytosis and survive in human serum.

Many bacteria evade phagocytosis by forming aggregates. When they form on surfaces, these bacterial aggregates are called biofilms. SasX promoted aggregation and biofilm formation, phenotypes that therefore presumably form the mechanistic basis of the immune evasion and virulence-enhancing properties associated with sasX. Furthermore, biofilm formation may contribute to prolonged survival on abiotic surfaces, which possibly also contributes to the spread of MRSA in the hospital environment.

Many important questions regarding sasX remain. First, where did it come from? The finding that there is a very similar prophage in an S. epidermidis strain, also harboring sasX, indicates that it was acquired from coagulase-negative staphylococci by horizontal gene transfer, as has been proposed for a series of virulence and resistance determinants. Second, what are the mechanisms underlying adhesion to nasal epithelial cells and biofilm formation? Usually, adhesion to eukaryotic cells that is facilitated by a bacterial surface protein is dependent on a specific receptor on the eukaryotic cell. Furthermore, many MSCRAMMs (microbial surface components recognizing adhesive matrix molecules), which connect bacteria to eukaryotic matrix proteins, contain large parts whose function it is to extend the protein through the cell wall, in order to make a terminal part accessible for receptor interaction. In that regard, it is interesting that the SasX protein is rather small in its secreted, processed form, only ~15 kDa, which is likely too small to protrude through the cell wall and expose a domain for interaction. It appears as if SasX uses another mechanism to achieve interaction with the eukaryotic cell. Thus, the eukaryotic SasX receptor and the mechanism by which SasX connects to it remain to be identified. Third, how does SasX accomplish bacterial aggregation and biofilm formation? Several surface proteins were recently described to promote biofilm formation, in addition to their primary function, which often is to facilitate adherence to tissue. Thus, a double function in promoting adherence to eukaryotic cells and bacterial aggregation, as found for SasX, is not uncommon. However, it is not clear whether bacterial aggregation in those cases is also due to a specific, receptor-type interaction, or dependent on a non-specific aggregation phenomenon, possibly based on the physico-chemical properties of exposed protein domains.

In conclusion, the identification of sasX as a driving force of an MRSA epidemic represents an important example helping us to understand how novel MRSA clones arise. By acquiring novel colonization factors they may replace other MRSA clones that are less optimally adjusted to persist in the hospital environment. While research on S. aureus toxins and their potential use as targets for anti-staphylococcal drug therapy has recently been very intense, especially regarding CA-MRSA, this finding suggests that there should also be a strong focus on the molecular factors promoting colonization and the spread of MRSA in hospitals. Given that most MRSA infections originate from colonizing strains, MRSA decolonization has often been proposed as a promising approach to reduce MRSA infection rates. Whether SasX may be used as a target for efforts aimed to reduce colonization or virulence remains to be shown. As a first step in that direction, the use of anti-SasX antibodies in animal models should be tested. Clearly, the use of any drug targeting SasX will be limited to sasX-positive clones. However, it is not uncommon to explore targets that are only present in very successful MRSA clones, such as in the often-discussed case of MRSA harboring Panton-Valentine-leukocidin genes. Probably, all such virulence or colonization-targeted approaches to find anti-MRSA therapeutics will require a mixture of drugs or antibodies targeting different bacterial virulence or colonization determinants.

Li M, Du X, Villaruz AE, Diep BA, Wang D, Song Y, et al. MRSA epidemic linked to a quickly spreading colonization and virulence determinant. Nat Med. 2012;18:816–9. doi: 10.1038/nm.2692.