Staphylococcus aureus

Staphylococcus aureus is an opportunistic pathogen that is frequently part of the human microflora and may cause disease when the immune system is compromised. S. aureus accounts for the largest number of nosocomial infections worldwide but is also associated with serious infections in the community and veterinary settings. The situation has been aggravated since these versatile pathogens have acquired resistance to virtually all classes of antibiotics.

1. Global surveillance

A. Health-care associated methicillin-resistant Staphylococcus aureus (HA-MRSA)

National surveillance

Nuno A. Faria, Diana Espadinha, Maria Miragaia

The prevalence of MRSA in Portuguese hospitals has been close to 50% for over 10 years and remains one of the highest in Europe. Having a catalogue of the MRSA clonal types circulating in Portuguese hospitals is crucial to help designing national intervention and prevention strategies.
During the last 20 years studies from our Laboratory allowed us to describe the successive waves of MRSA clones in Portugal, which contrasts with other countries with a similar prevalence of MRSA, where the major clones have been stable for decades. To follow-up the epidemiology of MRSA we continue to periodically analyze the population structure of MRSA in hospitals in Portugal. Moreover, results obtained are part of a major study focused on the epidemiology of nosocomial MRSA in Europe (SRL, from Staphylococcus Reference Laboratories).

Figure 1: Distribution of spa types found in S. aureus collected from blood stream infections in Europe. Source: Grundmann et al. PLOS one. 2010.

International surveillance

MRSA in Portuguese speaking African countries (PALOP) and East Timor

Teresa Conceição, Celine Coelho, Marta Aires-de-Sousa 

Despite the perceived importance and the large volume of research devoted to the study of methicillin-resistant Staphylococcus aureus (MRSA) across the developed world, almost nothing is known about its prevalence and transmission in resource-poor regions such as the Portuguese-speaking African countries (PALOP) and East Timor. Understanding the burden of MRSA colonization/disease in PALOP nations and East Timor, countries that have close demographic relationships with Portugal, will widely contribute to the local and worldwide MRSA control.

Construction of a MRSA reference collection

Nuno Faria, Teresa Conceição, Maria Miragaia, Helena de Deus, Jonas Almeida

Over the years our laboratory compiled a collection of >20,000 S. aureus isolates, including MRSA and MSSA from nosocomial and community origins isolated worldwide. All major epidemic HA- and CA-MRSA clones are represented as well as a high number of sporadic isolates. These isolates were characterized in detail by a combination of molecular typing techniques.

This extensively characterized international collection has been the base of our collaboration with the Sanger Institute in Cambridge UK and will be used to construct a MRSA reference collection. A simple sloppy semantic database (S3DB) relative to this collection is under construction in collaboration with Professor Jonas Almeida.

B. Community-associated methicillin-resistant S. aureus (CA-MRSA)

Once confined to hospital boundaries, methicillin-resistant S. aureus (MRSA) have emerged in the last two decades as a leading cause of severe skin and soft tissue infections in otherwise healthy individuals.

National surveillance

Ana Tavares, Teresa Conceição, Marta Aires de Sousa, Maria Miragaia

In spite of the clinical importance of CA-MRSA, little information exists on its prevalence in Portugal. 

Currently, we are undertaking studies that aim to identify the main clones circulating in the community in Portugal, in different populations, namely in patients at hospital entry, patients with skin and soft tissue infections (SSTI) attending health-care centers and residents in long-term care facilities, and children attending day-care centers. Moreover, we have screened public buses as possible reservoirs of MRSA.
Our preliminary results showed that rates of colonization of children with CA-MRSA are very low, but public buses are reservoirs of MRSA usually observed in the hospital environment, belonging to the so called EMRSA-15 clone.
From these approaches we expect to obtain data that will help in the design of infection control and prevention strategies to contain the dissemination of MRSA in the community in a country with a high prevalence of MRSA in hospitals.

International surveillance

Joana Rolo, Ons Bouchami, Nuno Faria, Maria Miragaia

In Europe the epidemiology of nosocomial MRSA is well understood, but the epidemiology of CA-MRSA is incompletely documented due to poor geographic coverage and outdated data. Under EU Project CONCORD collection of methicillin-susceptible and MRSA from persons in the community in the 16 most populous European countries was gathered and characterized by molecular typing techniques.

Figure 2: Distribution of the most prevalent MRSA and MSSA community associated epidemic and related clones in the 16 most populous European countries. A - MRSA, B - MSSA. Source: Rolo et al. PLOS one. 2012

2. Evolution of methicillin-resistant S. aureus (MRSA)

In order to understand in more detail the origin and evolution of several important pandemic MRSA clones, we a high-throughput genomics approach was used through collaboration with the Sanger Institute. This strategy provided a high-resolution view of the epidemiology and microevolution of the clones under study.

Figure 3: Evolution of MRSA during hospital transmission and intercontinental spead. Square colors indicate different clusters of the same MRSA clone.

3. Novel methods for typing of Staphylococcus aureus

A. SCCmec typing strategies

The mobile SCCmec element carries the mecA gene, the central element of broad-spectrum β-lactam resistance in staphylococci. So far, 11 SCCmec structural types, ranging in sizes of 20 – 60 kb, have been identified which are defined according to specific combinations of the mecA vicinity genetic organization with the ccr allotype, a locus encoding for the recombinases involved in the mobility of this chromosomal island. Assignment of SCCmec types to methicillin-resistance staphylococcal (MRS) strains has been revealed an important addendum to the molecular characterization of clonal types, providing important insights into the assembly mechanisms and dissemination routes of this element within a particular staphylococcal species and across the entire staphylococcal population.

Following our pioneer effort in 2002 in establishing an assay for the rapid assignment of SCCmec types to MRSA strains (Oliveira, D.C. et al., 2002) – SCCmec multiplex PCR and ccrB typing – we are at the moment developing an additional typing system targeting the ccrC, a cassette chromosome recombinase carried within SCCmec type V and widely disseminated among CA-MRSA and FA-MRSA.

ccrC typing

Catarina Milheiriço, Celine Coelho, Maria Miragaia

This consists of the sequencing an internal fragment of the ccrC gene, which enables the characterization of the ccrC allotype. This sequencing-based typing method will allow tracing strain dissemination, to identify new reservoirs of SCCmec V and possibly provide data that will allow to track the evolution of this SCCmec type. We are also developing in collaboration with colleagues of the INESC-ID, Lisbon, Portugal, an online resource for data deposition and analysis – “ccrC typing tool” that will be integrated in the already available ccrB typing tool. This strategy has the advantage to be easily integrated into other widely used DNA sequence-based typing strategies, such as spa typing and MLST.

Selected references

1. Rolo, J., M. Miragaia, A. Turlej-Rogacka, J. Empel, O. Bouchami, N. A. Faria, A. Tavares, W. Hryniewicz, A C. Fluit, H. de Lencastre and the CONCORD working group. 2012. High genetic diversity among community-associated Staphylococcus aureus in Europe: results from a multicenter study. PLoS One. 7:e34768.

2. Deus, H. F., M. C. Correa, R. Stanislaus, M. Miragaia, W. Maass, H. de Lencastre, R. Fox, and J. S. Almeida. 2011. S3QL: A distributed domain specific language for controlled semantic integration of life sciences data. BMC Bioinformatics 12:285.
3. De Lencastre, H. and A. Tomasz. 2011. The CEM-NET initiative: Molecular biology and epidemiology in alliance – tracking antibiotic resistant staphylococci and pneumococci in hospitals and in the community. Int. J. Med. Microbiol. 301:623-629.
4. Conceição, T., M. Aires de Sousa, N. Pona, M. J. Brito, C. Barradas, R. Coelho, T. Sardinha, L. Sancho, G. de Sousa, M. C. Machado, and H. de Lencastre. 2011. High prevalence of ST121 in community associated methicillin-susceptible Staphylococcus aureus lineages responsible for skin and soft tissue infections in Portuguese children. Eur. J. Clin. Microbiol. Infect. Dis. 30:293-7.
5. Harris, S., E. J. Feil, M. T.G. Holden, M. A. Quail, E. K. Nickerson, N. Chantratita, S. Gardete, A. Tavares, N. Day, J. Lindsay, J. Edgeworth, H. de Lencastre, J. Parkhill, S. J. Peacock, and S. D. Bentley. 2010. Evolution of MRSA during hospital transmission and intercontinental spread. Science 327:469-74.
6. Conceição, T., A. Tavares, M. Miragaia, K. Hyde, M. Aires-de-Sousa and H. de Lencastre. 2010. Prevalence and clonality of methicillin resistant Staphylococcus aureus (MRSA) in the Atlantic Azores islands: predominance of SCCmec types IV, V and VI. Eur. J. Clin. Microbiol. Infect. Dis. 29:543-50.
7. Tavares, D. A., R. Sá-Leão, M. Miragaia, and H. de Lencastre. 2010. Large screening of CA-MRSA among Staphylococcus aureus colonizing healthy young children in Portugal. BMC Infect Dis. 10:110.
8. Grundmann, H., D. M. Aanensen, C. C. van den Wijngaard, B. G. Spratt, D. Harmsen, A. W. Friedrich, the European Staphylococcal Reference Laboratory Working Group. 2010. Geographic Distribution of Staphylococcus aureus Causing Invasive Infections in Europe: A Molecular-Epidemiological Analysis. PLoS Medicine. 7(1):e1000215.