Methicillin-resistant Staphylococcus aureus (MRSA) is an important nosocomial pathogen associated with significant morbidity and mortality. Previous colonisation with this pathogen is a risk factor for the development of subsequent infection. Tolllike receptors (TLRs) are a family of transmembrane receptors of the innate immune system that recognize pathogen-associated molecular patterns. The role of nasal colonisation of S. aureus has started to receive more attention. In spite of this, there are not enough studies looking at its effects on human primary nasal epithelial cells and their response to TLR ligands. The respiratory tract itself seems to pose a contradiction given by the clinical observation that its upper portion (nasal compartment) allows the growth of bacteria, acting like a reservoir, whereas the lower portion (lung compartment) reacts with an exuberant inflammatory response to the same organisms, as noted during pneumonia. The mechanism related with this phenomenon remains to be elucidated. A negative regulator of the TLR signalling cascade called toll-interacting protein (tollip) has been demonstrated to induce hyporesponsiveness in the gastrointestinal tract in the presence of bacteria. So far, tollip has not been demonstrated in the respiratory tract. Aims: To compare the responses of the upper and lower respiratory tract to TLR ligands, to characterise the role of tollip in the respiratory tract and its effects in the induction of tolerance, and to determine the cellular response to nasal carriage of S. aureus. Materials and Methods: The cell line RPMI 2650 (representative of nasal epithelium) and the cell line A549 (representative of type II alveolar epithelium) were used to establish the cytokine response to stimulation with TLR ligands and to demonstrate the presence of tollip protein by immunocytochemistry and enzymelinked immunosorbent assay (ELISA). Primary human nasal epithelial and type II alveolar epithelial cells were isolated and cultured from consented subjects. The cytokine response to stimulation was measured using cytokine bead array and the presence of tollip was determined by immunofluorescence and quantitative polymerase chain reaction. The presence of TLRs was assessed by immunocytochemistry in primary nasal and type II alveolar epithelial cells and the response to stimulation with the TLR9 agonist CpG-C ODN was assessed in these cells as well as in primary human type II alveolar epithelial cells. Subjects were also assessed for nasal carriage of S. aureus and their associated cytokine responses. Results: The RPMI 2650 cell line, despite retaining phenotypic characteristics of the nasal epithelium, appears unresponsive to stimulation with TLR ligands. In contrast, the A549 cell line responded significantly to stimulation with TLR ligands. Primary human nasal epithelial cells responded by secreting higher amounts of interleukin (IL)-8 and IL-6 in response to stimulation with S. aureus peptidoglycan (PGN) and tumour necrosis factor alpha (TNF-α) with a strong trend toward statistical significance. These cells did not respond to stimulation with Pseudomonas aeruginosa LPS. Primary type II alveolar epithelial cells responded significantly to stimulation with S. aureus PGN by increasing the secretion of IL-8, IL-6, IL-1β, TNF-α and IL-10 into cultured supernatant. Cells from the upper respiratory tract displayed a more tolerant phenotype given by the lower levels in cytokine production in response to stimulation with S. aureus PGN, in contrast to alveolar epithelial cells. TLRs were identified in primary nasal epithelial cells. The negative regulator tollip was identified in cell lines as well as primary cells of the respiratory tract in its three segments: nasal, bronchial and type II alveolar. It was not possible to demonstrate an up-regulation of tollip after stimulation with TLR ligands in any of the cell types studied, although, it was possible to observe a significantly higher constitutive level in tollip mRNA transcripts from primary nasal epithelial cells in comparison to type II alveolar epithelial cells. TLR9 was identified in human primary nasal epithelial cells, although it was not possible to observe an increase in cytokine production after stimulation with a TLR9 agonist. TLR9 was expressed strongly in primary type II alveolar epithelial cells which responded by significantly increasing IL-8 production after stimulation with CpG-C ODN. Primary nasal epithelial cells from individuals who carry S. aureus exhibit a proinflammatory profile, as evidenced by higher basal levels of IL-8 and IL-6 in comparison to non-colonised controls. Conclusion: The upper respiratory tract epithelium displays a tolerant phenotype in response to stimulation with TLR ligands in comparison to the lower respiratory epithelium, potentially favouring nasal colonisation by S. aureus. Tollip m-RNA transcripts appear to be up-regulated constitutively in the nasal epithelium which might favour this response. Staphylococcus aureus colonisation is however associated with a local pro-inflammatory state in the nasal epithelium of carrier individuals.

Submitted by Flávia Sousa (flaviabs@ufba.br) on 2013-04-16T16:00:32Z No. of bitstreams: 1 2012 - Mestrado PMPGCF - RNF.pdf: 2912444 bytes, checksum: b76fa79b8643234c9d74ee46adea96a0 (MD5) / Approved for entry into archive by Flávia Sousa(flaviabs@ufba.br) on 2013-05-06T11:43:43Z (GMT) No. of bitstreams: 1 2012 - Mestrado PMPGCF - RNF.pdf: 2912444 bytes, checksum: b76fa79b8643234c9d74ee46adea96a0 (MD5) / Made available in DSpace on 2013-05-06T11:43:43Z (GMT). No. of bitstreams: 1 2012 - Mestrado PMPGCF - RNF.pdf: 2912444 bytes, checksum: b76fa79b8643234c9d74ee46adea96a0 (MD5) / CNPq Universal 014/2008, 13389/2010-0; FAPESB/PPUS 008/2009 / O Staphylococcus aureus é um patógeno envolvido em infecções noscomiais e comunitárias. O objetivo deste estudo foi analisar in vitro a atividade antimicrobiana do extrato etanólico da folha de Phanera flexuosa (Moric) L.P.Queiroz (EFPF), conhecida como escada de macaco, coletada na Floresta Nacional Contendas do Sincorá (BA), sobre o crescimento bacteriano e sobre alguns fatores de virulência de Staphylococcus aureus resistentes a antibióticos (MRSA), isolados de ambientes hospitalares de Vitória da Conquista (BA). O EFPF foi testado nas concentrações entre 4000 a 125 μg/mL na determinação de concentração inibitória mínima (CIM) e concentração bactericida mínima (CBM). Foram estudados Staphylococcus aureus ATCC 43300 e 31 isolados resistentes a oxacilina em antibiograma e que apresentaram o gene mecA em reação de PCR. Na CIM, as placas foram incubadas a 37°C, 24h, sendo interpretadas de acordo com o crescimento bacteriano e confirmado com o corante ressazurina. Como controle dos experimentos, foi utilizado o veículo, etanol (10%, v/v). Verificou-se a inibição de formação do biofilme de MRSA em 3, 5, 7 e 24 h, após a exposição ao EFPF, avaliou-se a formação de biomassa total com cristal de violeta, viabilidade celular com MTT e morfologia do biofilme usando microscopia confocal após 24 h (isolados clínicos 16A e 92). Foi realizada curva de crescimento de MRSA avaliando a absorbância e viabilidade da cultura bacteriana suplementada com 1% glicose, e tratada com o veículo ou EFPF em concentração subinibitória (CIM50). Nos tempos de 5 e 24 h de exposição ao EFPF, da curva foi analisada a expressão gênica de α-hla, PVL, sea, seb, spa, icaA, icaB, icaC, icaD, icaR dos isolados 16A e 112. Foram realizadas 03 triplicatas para cada experimento e análise estatística. Todos os isolados testados e a cepa ATCC apresentaram CIM ente 125 e 4000 μg/mL, sendo resultados relevantes de CIM (125 μg/mL) foram obtidos para a cepa ATCC 43300 e para os isolados 27A, 27B, 28, 29, 33A, 43.1, 47.1, 52, 76, 85.1, 85.2, 92 e 113. Quanto ao CBM, ocorreu para os isolados 47.1 e 92 (4000 μg/mL), isolado 12D (1000 μg/mL), e isolado 29 (500 μg/mL). Na inibição de formação de biofilme, os resultados em redução de células viáveis e formação da biomassa total ocorreram nos tempos de 7 e 24h, sendo que na concentração 5000 μg/mL foi efetiva no biofilme de 24 h. Quanto à curva de crescimento, a atividade antimicrobiana de EFPF ocorreu na fase final exponencial e estacionária inicial avaliada pela redução de absorbância, e também a redução da viabilidade celular do EFPF em relação ao veículo nas cepas testadas. Houve diminuição da expressão de PVL do isolado 16A tratado com EFPF (p<0,05), aumento da expressão de icaC, icaR e spa do isolado 16A em relação ao veículo (p<0,05). O EFPF apresenta atividade antimicrobiana contra S. aureus e influencia a expressão de fatores de virulência das cepas MRSA testadas, sendo um potencial antimicrobiano e promissora fonte para o isolamento de substâncias bioativas contra microrganismos multirresistentes. PIBIC/CNPq, CNPq Universal 14/2008, 14/2010, 13389/2010-0, Decit/SCTIE/MS (CNPq/FAPESB/SESAB), PERMANECER /UFBA, MEC-PET/SESU/SECAD. / Vitória da Conquista - BA

Staphylococcus aureus (S. aureus) is a bacterium that commonly colonises the skin and nares of around one third of otherwise healthy individuals. While colonisation is benign, S. aureus can cross skin and mucosal barriers to cause infections that manifest as clinical disease. Clinical outcomes are diverse and range from mild, non-complicated and often self-limiting skin and soft tissue infections (including boils, abscesses and cellulitis) to more severe and life-threatening conditions including pneumonia, toxic shock syndrome and bacteraemia. Medication isn’t always needed for mild S. aureus infections as often they resolve with time but, for severe or persistent cases, antimicrobial treatment is generally required. Following decades of widespread and intensive usage of topical, enteral and parenteral antimicrobials to treat S. aureus infections; AMR has become an established and ubiquitous problem in the treatment of infections caused by this microorganism, especially when in the methicillin resistant form (i.e. MRSA). The aim of this thesis was to examine aspects of S. aureus epidemiology (including MRSA and methicillin-sensitive S. aureus (MSSA)) in Scotland using statistical methods and data from several large public health databases. More specifically this involved: descriptions of spatial and temporal trends of morbidity and mortality; comparisons of epidemiological and molecular attributes (including antimicrobial resistance) of (1) MSSA and MRSA, and (2) the dominant clones of MRSA (i.e. EMRSA-15 and EMRSA-16); descriptions of spatial and temporal trends of antimicrobial prescribing in primary and secondary care and any associations between prescribing rates and MRSA antimicrobial resistance; and carrying out a hospital-level risk factor analysis of MRSA, testing hypotheses that hospital size, hospital connectivity (through shared transfer patients) and hospital category have an effect on hospital-level incidences of MRSA in mainland Scottish hospitals. Results showed that total S .aureus bacteraemia and MRSA bacteraemia in Scotland statistically declined over time (p < 0.0001), but MSSA bacteraemia did not (p > 0.05). While combined mortality rates (i.e. all MSSA deaths (both primary and secondary cause), or all MRSA deaths (both primary and secondary cause)) mirror these findings; case-fatality ratios (CFR) show no declines over time for either MRSA or MSSA. Results also show that several epidemiological factors point towards a predominant community source for MSSA isolates (i.e. outside healthcare) and hospital source for MRSA. Evidence for this included: (1) the lack of resistance genes in the MSSA population, (2) MRSA was more associated with long-term care and high-risk patients in the specialties care of the elderly, high dependency units/intensive care units (HDU/ICU), and surgery and conversely MSSA with specialties that commonly served outpatients, and (3) the abundance of non-EMRSA-15/non- EMRSA-16 ‘other’ clones in the MSSA population as compared with the hospital-associated CC22 (EMRSA-15) and CC30 (EMRSA-16) clones. EMRSA-15 was by far the most dominant MRSA clone in Scotland with EMRSA-16 declining significantly and non-EMRSA-15/non-EMRSA-16 clones causing an increasing number of infections (over the time period 2003-2013). EMRSA-16 was resistant to a larger number of antimicrobials than EMRSA-15, typically 9 versus 5, and while resistance varied for EMRSA-16 over the study period, resistance remained stable for EMRSA-15. There was little difference between clinical and screening MRSA isolates. Analyses of antimicrobial prescribing showed that prescribing rates of several drugs increased over time (2003-2013). Prescribing was far higher in primary care settings than in secondary care, although this differed between antimicrobials. Significant positive associations between prescribing and resistance rates were found for gentamicin (pr - p<0.0001, se - p<0.0001) and trimethoprim (pr - p<0.01, se - p<0.0001) in both primary (pr) and secondary (se) care, and clindamycin (p<0.0001) in primary care only. Finally, in Scotland there is a threshold of connectivity above which the majority of hospitals, regardless of size, are positive for MRSA. Higher levels of MRSA are associated with the large, highly connected teaching hospitals with high ratios of patients to domestic staff. While there were a number of data limitations, this body of work provides a better understanding of the epidemiology of S. aureus including MRSA in Scotland.

No description available.

Background: Methicillin-resistant Staphylococcus aureus (MRSA) is a multidrug-resistant bacterium that causes serious infections. Specific measures to prevent the spread of MRSA are required within health care, and insufficient knowledge and lack of compliance to these measures have been identified in healthcare professionals. The need to adhere to special guidelines in order to prevent the spread of infection have been shown to create negative feelings for carriers of multidrug-resistant bacterium. Aim: The aim of this study was to investigate different aspects of living and being cared for with known carriage of MRSA. The fist aspect aimed to investigate how people experience their knowledge and the information they receive from the healthcare. The second aspect aimed to investigate their feelings and existence in relation to their carriage. The third aspect aimed investigate their experiences and encounters with health care. Method: Literature study with systematic approach and inductive method. The results were analysed with inspiration from thematic synthesis and based on ten qualitative original articles.  Results: Getting MRSA was shocking and shameful, and many blamed the health care for contracting MRSA. Living with MRSA had a negative impact on daily life. Life felt lonely and isolated, and people were scared to infect others. Being cared for with MRSA was described with experiences of lack knowledge from the health care professionals, inconsistencies in adherence to hygiene measures and unprofessional behaviour. Being cared for in isolation had a negative impact on the persons’ independence, their relations with the health care professionals and the quality of care. Conclusion: Getting MRSA is perceived as stigmatising and frightening, and the health care fails to provide adequate information to counteract these feelings. People feel sadness, abandonment and uncertainty about the future. The behaviour of health care professionals and their compliance with hygiene measures is inconsistent. This results in people feeling extradited to health care, which contributes to enhanced feelings of shame and stigmatisation, and people with MRSA do not receive health care on equal terms. / Bakgrund: Meticillinresistenta Staphylococcus aureus (MRSA) är en multiresistent bakterie som orsakar allvarliga infektioner. För att förhindra spridning av MRSA krävs särskilda hygienrutiner inom vården, och otillräcklig kunskap och bristande följsamhet till dessa har identifierats hos vårdpersonal. För att inte sprida smitta behöver personer med bärarskap av multiresistenta bakterier förhålla sig till särskilda riktlinjer, vilket har visat sig skapa negativa känslor för personerna. Syfte: Syftet var att undersöka olika aspekter av att leva och vårdas med känt bärarskap av MRSA. Den första aspekten syftade till att undersöka hur personerna upplever sin kunskap och informationen de får från vården. Den andra aspekten syftade till att undersöka deras känslor och tillvaro i relation till bärarskapet. Den tredje aspekten syftade till att undersöka deras erfarenheter och upplevelser av mötet med vården. Metod: Litteraturstudie med systematisk ansats och induktiv metod. Resultatet analyserades med inspiration av tematisk syntes och baseras på tio kvalitativa originalartiklar. Resultat: Att få MRSA var chockartat och skamfyllt, och många beskyllde vården för att ha fått MRSA. Att leva med MRSA påverkade vardagen negativt. Livet kändes begränsat, ensamt och isolerat och personerna var rädda för att smitta andra. Att vårdas med MRSA beskrevs med upplevelser av bristande kunskap hos vårdpersonalen, inkonsekvens i följsamhet till hygienrutiner och oprofessionellt bemötande. Att vårdas under isolering för MRSA inverkade negativt på personernas upplevelse av självständighet, relationen till vårdpersonalen och kvaliteten på omvårdnaden. Slutsats: Att få MRSA upplevs stigmatiserande och skrämmande och vården misslyckas med att ge adekvat information för att motverka dessa känslor. Personerna känner sorg, övergivenhet och framtiden känns osäker. Vårdpersonalens beteende och följsamhet till hygienrutiner är inkonsekvent, vilket gör att personerna känner sig utlämnade till vården. Detta bidrar till förstärkta känslor av skam och stigmatisering, och personer med MRSA får inte vård på lika villkor.

Background: Acinetobacter baumannii and methicillin-resistant Staphylococcus aureus (MRSA) represent the most worrying Gram-negative and Gram-positive nosocomial pathogens of the present age. They are of increasing concern in the clinical environment due to their multi-drug resistance and the dwindling therapeutic options available. A. baumannii is the most frequently isolated clinical species of the genus, and is able to rapidly acquire resistance. Hypermutators, most frequently deficient in mismatch repair (MMR) via defects in the mutS gene, have been associated with antimicrobial resistance in several bacterial populations. To date, however, the potential role of MMR-deficient mutators in the development of resistance in clinical Acinetobacter spp. has not been investigated. Biocides, most notably chlorhexidine (CHX), are increasingly used in the hospital environment to prevent bacterial spread. This has led to concerns about the development of reduced biocide susceptibility and associated antibiotic resistance in hospital bacterial populations, where there is frequent exposure to both of these factors. The effect of CHX upon defined clinical MRSA isolates is examined here. Methods: The mutS gene of clinical Acinetobacter spp. isolates with varying sensitivities was sequenced and compared to establish whether any variations were present. Mutation studies were performed on isolates by challenging them with ciprofloxacin to determine whether different mutS types correlated with any variation in their ability to develop significant fluoroquinolone resistance. The response of clinical MRSA isolates to a range of CHX concentrations was examined with susceptibility testing methods, and effects were compared with standard strains. Determination of post-exposure minimum inhibitory concentrations (MICs) of a range of antibiotics enabled evaluation of whether exposure to CHX had an effect on susceptibility to antibiotics. Results: Variation was observed in the mutS gene of clinical Acinetobacter spp. isolates, with greater homology observed as resistance increased. A highly conserved and previously unreported amino acid sequence was discovered in resistant isolates. Nonresistant isolates with this ‘R-type’ mutS sequence appeared to have a greater ability to develop significant ciprofloxacin resistance. Clinical MRSA isolates had varying susceptibility to CHX, and there were differences in the susceptibility of standard strains compared to clinical isolates. CHX residues exerted a prolonged minimal inhibitory effect, and several increases in antibiotic MICs following CHX exposure were observed. Conclusions: The correlation of the mutS sequence with mutation ability suggests that defects in the mutS gene may have a role to play in the ability of certain Acinetobacter spp. to rapidly acquire resistance. This could have implications for the treatment of Acinetobacter spp. infections, and may enable quick determination of which clinical isolates have the potential to develop clinically significant resistance. Incomplete eradication due to the prolonged minimal effect of CHX residues may act as a selective pressure in the hospital environment, allowing survival of reduced susceptibility MRSA isolates. Increases in antibiotic MICs following CHX exposure is of grave concern for the future of biocide usage.

<p>The purpose of this study was to describe the level of knowledge that nurse students in term 6 have about Methicillin-Resistant Staphylococcus aureus (MRSA) and basic hygiene routines. The data collection was carried out by questionnaires. The questionnaire contained questions about knowledge regarding MRSA and basic hygiene routines. The questionnaires were handed out at an obligatory tuition occasion and all of the nurse students who were present (n=57) were asked to take part. The sample consisted of 45 women and 12 men. The youngest participant was 22 years old and the oldest participant was 52 years old. The main results showed a large variation regarding the participants knowledge on the subjects. Regarding questions about basic hygiene routines the majority had given the right answers. The number of right answers in the survey on questions about the participants knowledge of MRSA was lower. Only 12 % had given the right answer to the question about common symptoms of MRSA infection. Regarding the question about whether the nurse students considered themselves knowledgeable enough about MRSA for their future occupation, 32% had answered "to a high degree".</p> / <p>Syftet med studien var att beskriva vilka kunskaper sjuksköterskestuderande i termin 6 har om Methicillinresistenta Staphylococcus aureus (MRSA) och basala hygienrutiner. Datainsamlingen genomfördes med enkäter. Enkäten innehöll frågor om kunskaper gällande MRSA samt basala hygienrutiner. Enkäterna delades ut vid ett obligatoriskt undervisningstillfälle och samtliga, vid undervisningstillfället närvarade sjuksköterskestudenter (n =57) tillfrågades om deltagande. Undersökningsgruppen bestod av 45 kvinnor och 12 män. Yngsta deltagaren var 22 år och den äldsta deltagaren var 52 år. Huvudresultatet visade en stor variation gällande deltagarnas kunskaper i ämnena. På frågor angående basala hygienrutiner hade majoriteten svarat rätt. På de frågor som berörde deltagarnas kunskaper om MRSA var antalet rätta svar i undersökningen lägre. Endast 12 % hade svarat rätt på frågan om symtom som är vanliga vid MRSA infektion. På frågan om sjuksköterskestudenterna ansåg att de hade tillräckliga kunskaper om MRSA för sitt kommande yrke som sjuksköterskor svarade 32 % ”i hög grad”.</p>

The purpose of this study was to describe the level of knowledge that nurse students in term 6 have about Methicillin-Resistant Staphylococcus aureus (MRSA) and basic hygiene routines. The data collection was carried out by questionnaires. The questionnaire contained questions about knowledge regarding MRSA and basic hygiene routines. The questionnaires were handed out at an obligatory tuition occasion and all of the nurse students who were present (n=57) were asked to take part. The sample consisted of 45 women and 12 men. The youngest participant was 22 years old and the oldest participant was 52 years old. The main results showed a large variation regarding the participants knowledge on the subjects. Regarding questions about basic hygiene routines the majority had given the right answers. The number of right answers in the survey on questions about the participants knowledge of MRSA was lower. Only 12 % had given the right answer to the question about common symptoms of MRSA infection. Regarding the question about whether the nurse students considered themselves knowledgeable enough about MRSA for their future occupation, 32% had answered "to a high degree". / Syftet med studien var att beskriva vilka kunskaper sjuksköterskestuderande i termin 6 har om Methicillinresistenta Staphylococcus aureus (MRSA) och basala hygienrutiner. Datainsamlingen genomfördes med enkäter. Enkäten innehöll frågor om kunskaper gällande MRSA samt basala hygienrutiner. Enkäterna delades ut vid ett obligatoriskt undervisningstillfälle och samtliga, vid undervisningstillfället närvarade sjuksköterskestudenter (n =57) tillfrågades om deltagande. Undersökningsgruppen bestod av 45 kvinnor och 12 män. Yngsta deltagaren var 22 år och den äldsta deltagaren var 52 år. Huvudresultatet visade en stor variation gällande deltagarnas kunskaper i ämnena. På frågor angående basala hygienrutiner hade majoriteten svarat rätt. På de frågor som berörde deltagarnas kunskaper om MRSA var antalet rätta svar i undersökningen lägre. Endast 12 % hade svarat rätt på frågan om symtom som är vanliga vid MRSA infektion. På frågan om sjuksköterskestudenterna ansåg att de hade tillräckliga kunskaper om MRSA för sitt kommande yrke som sjuksköterskor svarade 32 % ”i hög grad”.

>Magister Scientiae - MSc / The growing resistance of Methicillin-Resistant Staphylococcus aureus (MRSA) to currently prescribed drugs has resulted in the failure of prevention and treatment of different infections caused by the superbug. Therefore, to keep pace with the resistance, there is a pressing need for novel antimicrobial agents, especially from non-conventional sources. Several natural products (NPs) have displayed varying in vitro activities against the pathogen but few of these natural compounds have been studied for their prospects to be potential antimicrobial drug candidates. This may be due to the high cost, tedious, and time-consuming process of conducting the important preclinical tests on these compounds. Hence, there is a need for cost-effective strategies for mining the available data on these natural compounds. This would help to get the knowledge that may guide rational prioritization of “likely to succeed” natural compounds to be developed into potential antimicrobial drug candidates. Cheminformatic approaches in drug discovery enable chemical data mining, in conjunction with unsupervised and supervised learning from available bioactivity data that may unlock the full potential of NPs in antimicrobial drug discovery. Therefore, taking advantage of the available NPs with their known in vitro activity against MRSA, this study conducted cheminformatic and data mining analysis towards hit profiling, hit-prioritization, hit-optimization, and target prediction of anti-MRSA NPs. Cheminformatic profiling was conducted on the 111 anti-MRSA NPs (AMNPs) retrieved from literature. About 20 current drugs for MRSA (CDs) were used as a reference to identify AMNPs with promising prospects to become drug candidates.

Methicillin-resistenta Staphylococcus aureus (MRSA) har blivit en allt vanligare patogen inom sjukvården och i samhället. MRSA orsakar infektioner som inte kan behandlas med β-laktamantibiotika. För att förhindra spridning genomgår patienter och sjukvårdspersonal screening-tester. I dessa screening-tester ingår PCR-analys av mecA, nuc och/eller Sa442. MecA är lokaliserad på Staphylococcal Cromosomal Cassette mec (SCCmec) och används som en markör för MRSA medan nuc och Sa442 anger S. aureus. PCR-positiva isolat odlas ut på agarplattor efter anrikning i en selektiv buljong. Kolonier av S. aureus resistensbestäms mot cefoxitin som MRSA är resistent mot. Idag används dock PCR-analys av mecA som en referensmetod för diagnostisering av MRSA. Under det senaste årtiondet har även rapporterats fynd av stammar som enligt resistensmönster är MRSA men som har utfallit negativt för mecA i PCR. Under 2011 rapporterades en ny variant av SCCmec och en ny variant av mecA, mecALGA251. I denna studie har en realtids-PCR tagits fram för att identifiera den nya varianten, mecALGA251. Denna PCR användes för att undersöka 43 kliniska isolat, fyra cefoxitin-känsliga S. aureus från rutinen och tre referensstammar. De kliniska isolaten hade samlats in under perioden 2004 – 2011 och hade uppvisat cefoxitin-resistens men gett negativt resultat i mecA-PCR. Totalt 40 av de 43 cefoxitin-resistenta kliniska isolaten visades bära mecALGA251. Resistensbestämningar med diskdiffusion och E-test mot cefoxitin, oxacillin, cefuroxim och cefotaxim visade att denna typ av MRSA inte kan skiljas från klassisk MRSA. Resultaten visar att cefoxitin-resistenta S. aureus isolat som bär mecALGA251 finns bland skånska patienter. De visar också att det finns cefoxitin-resistent S. aureus som saknar både klassiska mecA och mecALGA251. Dessa stammar har inte studerats vidare i denna studie. / Methicillin-resistant Staphylococcus aure s (MRSA) has become a more frequent pathogen within health care facilities and the community. MRSA causes infections that can’t be treated with β-lactamantibiotics. To prevent the spread of MRSA, patients and medical personnel undergo screening-tests. In the screening-tests PCR-analysis of mecA, nuc and/or Sa442 is included. MecA is located at Staphylococcal Chromosomal Cassette mec (SCCmec) and is a marker for MRSA, whereas nuc and Sa442 state regular S. Aureus infections. PCR-positive isolates are grown on agar plates after enrichment in selective broth. Colonies of S. aureus are tested for cefoxitin susceptibility to which MRSA is resistant. PCR-analysis of mecA is the reference method that is being used today when MRSA is being diagnosed. During the last decade cefoxitin-resistant strains that lack mecA in the PCR has been reported. In 2011 a new variant of SCCmec and a new variant of mecA, mecALGA251 was reported. In this study a new real-time-PCR has been developed in order to identify mecALGA251. The new PCR protocol was being used to examine 43 clinical isolates, four cefoxitin-susceptible S. aureus from the routine and three reference strains were examined. The clinical isolates had been collected during the period 2004-2011 and were cefoxitin-resistant but lacked mecA. In total of 40 of the 43 cefoxitin-resistant was PCR positive for mecALGA251. Susceptibility testing with disk diffusion and E-test for cefoxitin, oxacillin, cefuroxime and cefotaxime showed that this type of MRSA can’t be distinguished from regular MRSA. The results showed that cefoxitin-resistantS. aureus isolates carrying mecALGA251 exist among patients in Skåne County. One cefoxitin-resistant S. aureus isolate lacked both classic mecA and mecALGA251, which indicates that other mechanisms may exist, however these results has not been further analysed in this study.