Livestock-associated methicillin-resistant Staphylococcus aureus in humans, Europe.

To estimate the proportion of methicillin-resistant Staphylococcus aureus (MRSA) isolates from humans that were sequence type (ST) 398, we surveyed 24 laboratories in 17 countries in Europe in 2007. Livestock-associated MRSA ST398 accounted for only a small proportion of MRSA isolates from humans; most were from the Netherlands, Belgium, Denmark, and Austria.

Streptococcus dysgalactiae subsp. equisimilis Bacteremia, Finland, 1995-2004.

We conducted a retrospective population-based study of 140 episodes of Streptococcus dysgalactiae subsp. equisimilis bacteremia occurring in Finland during 1995-2004. Rare emm types were associated with more severe disease and increased mortality rates. Skin and soft tissue infections were more frequent clinical signs among cases caused by common emm types.

Predictors of death after severe Streptococcus pyogenes infection.

An evaluation of the relative importance of host and pathogen factors on the survival rate of patients with invasive Streptococcus pyogenes infection found a number of clinical and demographic factors to be associated with risk for death. Some evidence suggested a seasonal pattern to patient survival rate.

Complement factor H allotype 402H is associated with increased C3b opsonization and phagocytosis of Streptococcus pyogenes.

The main virulence factor of group A streptococcus (GAS), M protein, binds plasma complement regulators factor H (FH) and FH-like protein 1 (FHL-1) leading to decreased opsonization. The M protein binding site on FH is within domain 7 in which also the age-related macular degeneration (AMD)-associated polymorphism Y402H is located. We studied if FH allotypes 402H and 402Y have different binding affinities to GAS. Plasma-derived FH allotype 402H and its recombinant fragment FH5-7(402H) showed decreased binding to several GAS strains. Growth of GAS in human blood taken from FH(402H) homozygous individuals was decreased when compared with blood taken from FH(402Y) homozygous individuals. The effect of the allotype 402H can be explained by combining the previous M protein mutagenesis data and the recently published crystal structure of FH6-8. In conclusion the data indicate that the AMD-associated allotype 402H leads to diminished binding of FH to GAS and increased opsonophagocytosis of the bacteria in blood. These results suggest that the homozygous presence of the allele 402H could be associated with decreased risk for severe GAS infections offering an explanation for the high frequency of the allele despite its association with visual impairment.

Methicillin-resistant Staphylococcus aureus screening by online immunometric monitoring of bacterial growth under selective pressure.

Rapid, high-throughput screening tools are needed to contain the spread of hospital-acquired methicillin (meticillin)-resistant Staphylococcus aureus (MRSA) strains. Most techniques used in current clinical practice still require time-consuming culture for primary isolation of the microbe. We present a new phenotypic assay for MRSA screening. The technique employs a two-photon excited fluorescence (TPX) detection technology with S. aureus-specific antibodies that allows the online monitoring of bacterial growth in a single separation-free process. Different progressions of fluorescence signals are recorded for methicillin-susceptible and -resistant strains when the growth of S. aureus is monitored in the presence of cefoxitin. The performance of the new technique was evaluated with 20 MRSA strains, 6 methicillin-susceptible S. aureus strains, and 7 coagulase-negative staphylococcal strains and two different monoclonal S. aureus-specific antibodies. When either of these antibodies was used, the sensitivity and the specificity of the TPX assay were 100%. All strains were correctly classified within 8 to 12 h, and up to 70 samples were simultaneously analyzed on a single 96-well microtiter plate. As a phenotypic method, the TPX assay is suited for screening purposes. The final definition of methicillin resistance in any S. aureus strain should be based on the presence of the mecA gene. The main benefit afforded by the initial use of the TPX methodology lies in its low cost and applicability to high-throughput analysis.

Rapid emergence of emm84 among invasive Streptococcus pyogenes infections in Finland.

From 2005 to 2007, in Finland, the incidence of invasive Streptococcus pyogenes disease increased sharply, partly due to the uncommon emm84 gene becoming more prevalent from 2006 onwards. The overall case fatality rate of infections caused by strains carrying emm84 was not significantly different than that of infections caused by other types (7% versus 10%, respectively; P = 0.50).

Carriage of methicillin-resistant Staphylococci and their SCCmec types in a long-term-care facility.

Following an outbreak caused by staphylococcal cassette chromosome mec (SCCmec) type V methicillin (meticillin)-resistant Staphylococcus aureus (MRSA), a point-prevalence survey of the nasal carriage of staphylococci was conducted in a long-term-care facility in northern Finland in 2004. The focus was directed at methicillin-resistant coagulase-negative staphylococci (MR-CNS) and their SCCmec elements. A nasal swab was taken from 76 of the 80 residents 6 months after the onset of the outbreak. Staphylococcal isolates were identified by conventional methods and the GenoType Staphylococcus test, and their SCCmec elements were analyzed. Of the 76 individuals, 24 (32%) carried S. aureus and 67 (88%) CNS in their nostrils. Of the CNS carriers, 41 (61%) had at least one mecA-positive MR-CNS, and two individuals (3%) had both MRSA and methicillin-resistant Staphylococcus epidermidis (MRSE). Among the 61 MR-CNS isolates identified, 49 (80%) were MRSE. The distribution of the SCCmec types was diverse: 20 (33%) were of type IV, 11 (18%) of type V, 4 (6%) of type I or IA, 3 (4%) of type II, and 23 (38%) of new types (with six different combinations of ccr and other mec genes or only mecA). Both of the individuals with MRSA and MRSE shared SCCmec type V among their isolates. Nasal MR-CNS carriage was common among the residents of this long-term-care facility. A variety of SCCmec types, including many new types, were identified among the MR-CNS strains. The horizontal transfer of SCCmec elements is speculated based on the sharing of SCCmec type V between MRSA and MRSE.

Community-associated methicillin-resistant Staphylococcus aureus isolated in Finland in 2004 to 2006.

A nationwide population-based study on community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) in Finland during 2004 to 2006 showed that both incidence (1.9/100,000 population) and strain variation increased in comparison to years 1997 to 1999. There were 7 community-associated epidemic and 25 sporadic MRSA strain types. Half of these had Panton-Valentine leukocidin genes.

Clinical and microbiological characteristics of severe Streptococcus pyogenes disease in Europe.

In an attempt to compare the epidemiology of severe Streptococcus pyogenes infection within Europe, prospective data were collected through the Strep-EURO program. Surveillance for severe cases of S. pyogenes infection diagnosed during 2003 and 2004 was undertaken in 11 countries across Europe by using a standardized case definition and questionnaire. Patient data as well as bacterial isolates were collected and characterized by T and M/emm typing, and selected strains were analyzed for the presence of superantigen genes. Data were analyzed to compare the clinical and microbiological patterns of the infections across the participating countries. A total of 4,353 isolates were collected from 5,521 cases with severe S. pyogenes infections who were identified. A wide diversity of M/emm types (n = 104) was found among the S. pyogenes clinical isolates, but the M/emm type distribution varied broadly between participating countries. The 10 most predominant M/emm types were M/emm type 1 (M/emm1), M/emm28, M/emm3, M/emm89, M/emm87, M/emm12, M/emm4, M/emm83, M/emm81, and M/emm5, in descending order. A correlation was found between some specific disease manifestations, the age of the patients, and the emm types. Although streptococcal toxic shock syndrome and necrotizing fasciitis were caused by a large number of types, they were particularly associated with M/emm1 and M/emm3. The emm types included in the 26-valent vaccine under development were generally well represented in the present material; 16 of the vaccine types accounted for 69% of isolates. The Strep-EURO collaborative program has contributed to enhancement of the knowledge of the spread of invasive disease caused by S. pyogenes within Europe and encourages future surveillance by the notification of cases and the characterization of strains, which are important for vaccination strategies and other health care issues.

Acute bacterial, nonnecrotizing cellulitis in Finland: microbiological findings.

BACKGROUND: Bacterial, nonnecrotizing cellulitis is a localized and often recurrent infection of the skin. The aim of this study was to identify the beta-hemolytic streptococci that cause acute nonnecrotizing cellulitis infection in Finland. METHODS: A case-control study of 90 patients hospitalized for acute cellulitis and 90 control subjects was conducted during the period of April 2004-March 2005. Bacterial swab samples were obtained from skin lesions or any abrasion or fissured toe web. Blood culture samples were taken for detection of bacteremia. The patients, their household members, and control subjects were assessed for pharyngeal carrier status. beta-Hemolytic streptococci and Staphylococcus aureus were isolated and identified, and group A and G streptococcal isolates were further analyzed by T serotyping and emm and pulsed-field gel electrophoresis typing. RESULTS: beta-Hemolytic streptococci were isolated from 26 (29%) of 90 patients, 2 isolates of which were blood-culture positive for group G streptococci, and 24 patients had culture-positive skin lesions. Group G Streptococcus (Streptococcus dysgalactiae subsp. equisimilis) was found most often and was isolated from 22% of patient samples of either skin lesions or blood, followed by group A Streptococcus, which was found in 7% of patients. Group G streptococci were also carried in the pharynx of 7% of patients and 13% of household members but was missing from control subjects. Several emm and pulsed-field gel electrophoresis types were present among the isolates. Six patients (7%) had recurrent infections during the study. In 2 patients, the group G streptococcal isolates recovered from skin lesions during 2 consecutive episodes had identical emm and pulsed-field gel electrophoresis types. CONCLUSIONS: Group G streptococci, instead of group A streptococci, predominated in bacterial cellulitis. No clear predominance of a specific emm type was seen. The recurrent nature of cellulitis became evident during this study.

Binding of complement regulators factor H and C4b binding protein to group A streptococcal strains isolated from tonsillar tissue and blood.

Group A streptococcus (GAS) is the most common pathogen causing bacterial pharyngitis. We isolated streptococcal strains from tonsils removed from patients with tonsillar disease (n=202) and studied their ability to bind the complement regulators factor H (FH) and C4b binding protein (C4BP) using 125 I-labeled proteins. Blood isolates of GAS (n=10) were obtained from patients with bacteraemia. Streptococci were isolated from 21% of the tonsillitis patients. The emm and T types of the GAS strains were determined. Of the 26 GAS strains studied, only six could bind FH and/or C4BP above the threshold levels. The fraction of the offered radioactive protein bound ranged between 6-12% for FH and 19-56% for C4BP. The clinical course of the tonsillar disease was not related to the binding of FH or C4BP by GAS. The binding strains were mostly of the T4M4 or T28M28 type. From the invasive strains (n=10), three bound FH (binding level: 8-11%) and two C4BP (36-39%). The binding correlated only partially to M-protein (emm) type suggesting that the binding was not exclusively due to M-protein. The results indicate that complement regulator binding by GAS is only partially related to pathogenicity and not a universal property of all group A streptococci.

Epidemiology of severe Streptococcus pyogenes disease in Europe.

The past 2 decades have brought worrying increases in severe Streptococcus pyogenes diseases globally. To investigate and compare the epidemiological patterns of these diseases within Europe, data were collected through a European Union FP-5-funded program (Strep-EURO). Prospective population-based surveillance of severe S. pyogenes infection diagnosed during 2003 and 2004 was undertaken in 11 countries across Europe (Cyprus, the Czech Republic, Denmark, Finland, France, Germany, Greece, Italy, Romania, Sweden, and the United Kingdom) using a standardized case definition. A total of 5,522 cases were identified across the 11 countries during this period. Rates of reported infection varied, reaching 3/100,000 population in the northern European countries. Seasonal patterns of infection showed remarkable congruence between countries. The risk of infection was highest among the elderly, and rates were higher in males than in females in most countries. Skin lesions/wounds were the most common predisposing factor, reported in 25% of cases; 21% had no predisposing factors reported. Skin and soft tissue were the most common foci of infection, with 32% of patients having cellulitis and 8% necrotizing fasciitis. The overall 7-day case fatality rate was 19%; it was 44% among patients who developed streptococcal toxic shock syndrome. The findings from Strep-EURO confirm a high incidence of severe S. pyogenes disease in Europe. Furthermore, these results have identified targets for public health intervention, as well as raising awareness of severe S. pyogenes disease across Europe.

Nationwide trends in molecular epidemiology of methicillin-resistant Staphylococcus aureus, Finland, 1997-2004.

BACKGROUND: In Finland, the annual number of MRSA notifications to the National Infectious Disease Register (NIDR) has constantly increased since 1995, and molecular typing has revealed numerous outbreak isolates of MRSA. We analyzed the data on MRSA notifications of the NIDR, and MRSA isolates were identified mainly by pulsed-field gel electrophoresis (PFGE) at the National Reference Laboratory (NRL) in Finland during 1997-2004. One isolate representative of each major PFGE type was further characterized by multilocus sequence (MLST)-, staphylococcal cassette chromosome mec (SCCmec)-, and Panton-Valentine leukocidin (PVL)-typing. RESULTS: The annual number of MRSA notifications to the NIDR rose over ten-fold, from 120 in 1997 to 1458 in 2004, and the proportion of MRSA among S. aureus blood isolates tripled, from <1% during 1997-2003 to 2.8% in 2004. During the same period of time, 253 different strains among 4091 MRSA isolates were identified by PFGE: 215 were sporadic and 38 outbreak/epidemic strains, including 24 new strains. Two epidemic strains resembling internationally recognized MRSA clones accounted for most of the increase: FIN-16 (ST125:IA) from <1% in 1997 to 25% in 2004, and FIN-21 (ST228:I) from 6% in 2002 to 28% in 2004. Half of the ten most common strains carried SCCmec IV or V. CONCLUSION: The predominant MRSA strains seem to change over time, which encourages us to continue implementing active control measures with each new MRSA case.

A cluster of Candida krusei infections in a haematological unit.

BACKGROUND: Candida krusei infections are associated with high mortality. In order to explore ways to prevent these infections, we investigated potential routes for nosocomial spread and possible clonality of C. krusei in a haematological unit which had experienced an unusually high incidence of cases. METHODS: We searched for C. krusei contamination of the hospital environment and determined the level of colonization in patients and health care workers. We also analyzed the possible association between exposure to prophylactic antifungals or chemotherapeutic agents and occurrence of C. krusei. The C. krusei isolates found were genotyped by pulsed-field electrophoresis method in order to determine possible relatedness of the cases. RESULTS: Twelve patients with invasive C. krusei infection and ten patients with potentially significant infection or mucosal colonization were documented within nine months. We were unable to identify any exogenic source of infection or colonization. Genetic analysis of the isolates showed little evidence of clonal transmission of C. krusei strains between the patients. Instead, each patient was colonized or infected by several different closely related genotypes. No association between medications and occurrence of C. krusei was found. CONCLUSION: Little evidence of nosocomial spread of a single C. krusei clone was found. The outbreak may have been controlled by cessation of prophylactic antifungals and by intensifying infection control measures, e.g. hand hygiene and cohorting of the patients, although no clear association with these factors was demonstrated.

emm typing of invasive T28 group A streptococci, 1995-2004, Finland.

A total of 985 group A streptococcus (GAS) bacteraemia isolates collected in Finland during 1995-2004 were T-serotyped, and of these, 336 isolates of serotype T28 were subjected to further emm typing. The total number of isolates referred per year showed an increase within the study period, from 43 in 1995 to 130 in 2004. The annual incidence of invasive GAS (iGAS) bacteraemia showed a general increase during the study period, from 1.1 to 2.5 per 100 000 population. Serotype T28 remained among the most common serotypes, in addition to serotypes TB3264 and T1. The serotype T28 isolates were found to be distributed across six distinct emm types: emm28, emm77, emm53 (including subtypes 53.2 and 53.4), emm87, emm2 and emm4. The serotype distribution and the emm type distribution of serotype T28 fluctuated over time. Within the study period, the proportion of T28/emm28 isolates became the most prominent. During periods of low emm28 incidence, emm types 77 and 53 seemed to show a resurgence. emm typing revealed T28 isolates to be a genetically heterogeneous group harbouring a variety of distinct M proteins. This study confirms that T serotyping alone is not a sufficient method for epidemiological surveillance of iGAS.

An outbreak of Streptococcus equi subspecies zooepidemicus associated with consumption of fresh goat cheese.

BACKGROUND: Streptococcus equi subspecies zooepidemicus is a rare infection in humans associated with contact with horses or consumption of unpasteurized milk products. On October 23, 2003, the National Public Health Institute was alerted that within one week three persons had been admitted to Tampere University Central Hospital (TaYS) because of S. equi subsp. zooepidemicus septicaemia. All had consumed fresh goat cheese produced in a small-scale dairy located on a farm. We conducted an investigation to determine the source and the extent of the outbreak. METHODS: Cases were identified from the National Infectious Disease Register. Cases were persons with S. equi subsp. zooepidemicus isolated from a normally sterile site who had illness onset 15.9-31.10.2003. All cases were telephone interviewed by using a standard questionnaire and clinical information was extracted from patient charts. Environmental and food specimens included throat swabs from two persons working in the dairy, milk from goats and raw milk tank, cheeses made of unpasteurized milk, vaginal samples of goats, and borehole well water. The isolates were characterized by ribotyping and pulsed-field gel electrophoresis (PFGE). RESULTS: Seven persons met the case definition; six had septicaemia and one had purulent arthritis. Five were women; the median age was 70 years (range 54-93). None of the cases were immunocompromized and none died. Six cases were identified in TaYS, and one in another university hospital in southern Finland. All had eaten goat cheese produced on the implicated farm. S. equi subsp. zooepidemicus was isolated from throat swabs, fresh goat cheese, milk tank, and vaginal samples of one goat. All human and environmental strains were indistinguishable by ribotyping and PFGE. CONCLUSION: The outbreak was caused by goat cheese produced from unpasteurized milk. Outbreaks caused by S. equi subsp. zooepidemicus may not be detected if streptococcal strains are only typed to the group level. S. equi subsp. zooepidemicus may be a re-emerging disease if unpasteurized milk is increasingly used for food production. Facilities using unpasteurized milk should be carefully monitored to prevent this type of outbreaks.

Clustering of Serratia marcescens infections in a neonatal intensive care unit.

OBJECTIVES: To study clusters of infections caused by Serratia marcescens in a neonatal intensive care unit (NICU) and to determine risk factors for S. marcescens infection or colonization. DESIGN: Genotyping of S. marcescens isolates was performed by pulsed-field gel electrophoresis (PFGE). A retrospective case-control study was conducted. SETTING: A tertiary-care pediatric hospital with a 16-bed NICU. PATIENTS: All neonates with at least one culture positive for S. marcescens in the NICU during December 1999 to July 2002. Case-patients (n = 11) treated in the NICU during December 1999 to February 2000 were included in the case-control study. Neonates treated in the NICU for at least 72 hours during the same period with cultures negative for S. marcescens were used as control-patients (n = 27). RESULTS: S. marcescens was cultured from 19 neonates; 9 were infected and 10 were colonized. PFGE analysis identified three epidemic strains; each cluster consisted of identical isolates, except one isolate in the first cluster that was different. The risk factors identified were low birth weight, prematurity, prolonged respiratory therapy, prolonged use of antibiotics, and maternal infection prior to delivery. Overcrowding and understaffing were recorded simultaneously with the clusters. CONCLUSIONS: PFGE analysis showed three independent clusters. Several factors contributed to spread of the epidemic strains: (1) there were many severely premature and susceptible neonates, (2) the NICU was overcrowded during the clusters, and (3) transmission was likely to occur via the hands of staff. Cohorting and improvement of routine infection control measures led to the cessation of each cluster.

Origin and evolution of European community-acquired methicillin-resistant Staphylococcus aureus.

UNLABELLED: Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) was recognized in Europe and worldwide in the late 1990s. Within a decade, several genetically and geographically distinct CA-MRSA lineages carrying the small SCCmec type IV and V genetic elements and the Panton-Valentine leukocidin (PVL) emerged around the world. In Europe, the predominant CA-MRSA strain belongs to clonal complex 80 (CC80) and is resistant to kanamycin/amikacin and fusidic acid. CC80 was first reported in 1993 but was relatively rare until the late 1990s. It has since been identified throughout North Africa, the Middle East, and Europe, with recent sporadic reports in sub-Saharan Africa. While strongly associated with skin and soft tissue infections, it is rarely found among asymptomatic carriers. Methicillin-sensitive S. aureus (MSSA) CC80 strains are extremely rare except in sub-Saharan Africa. In the current study, we applied whole-genome sequencing to a global collection of both MSSA and MRSA CC80 isolates. Phylogenetic analyses strongly suggest that the European epidemic CA-MRSA lineage is derived from a PVL-positive MSSA ancestor from sub-Saharan Africa. Moreover, the tree topology suggests a single acquisition of both the SCCmec element and a plasmid encoding the fusidic acid resistance determinant. Four canonical SNPs distinguish the derived CA-MRSA lineage and include a nonsynonymous mutation in accessory gene regulator C (agrC). These changes were associated with a star-like expansion into Europe, the Middle East, and North Africa in the early 1990s, including multiple cases of cross-continent imports likely driven by human migrations. IMPORTANCE: With increasing levels of CA-MRSA reported from most parts of the Western world, there is a great interest in understanding the origin and factors associated with the emergence of these epidemic lineages. To trace the origin, evolution, and dissemination pattern of the European CA-MRSA clone (CC80), we sequenced a global collection of strains of the S. aureus CC80 lineage. Our study determined that a single descendant of a PVL-positive methicillin-sensitive ancestor circulating in sub-Saharan Africa rose to become the dominant CA-MRSA clone in Europe, the Middle East, and North Africa. In the transition from a methicillin-susceptible lineage to a successful CA-MRSA clone, it simultaneously became resistant to fusidic acid, a widely used antibiotic for skin and soft tissue infections, thus demonstrating the importance of antibiotic selection in the success of this clone. This finding furthermore highlights the significance of horizontal gene acquisitions and underscores the combined importance of these factors for the success of CA-MRSA.

A selective broth enrichment combined with real-time nuc-mecA-PCR in the exclusion of MRSA.

We analyzed the performance of a selective enrichment broth combined with Taqman-based real-time duplex nuc-mecA-PCR to expedite the screening of methicillin-resistant Staphylococcus aureus (MRSA). We found the broth to be able to select MRSA strains (oxacillin MIC range 4-256 microg/ml) from MSSA strains. A total of 31 MRSA strains were found from 1250 clinical samples screened. The nuc-mecA-PCR was positive from all enrichment broths containing MRSA. From the remaining 1219 samples negative for MRSA on culture/subculture, 138 samples were nuc+/mecA+ in PCR. The sensitivity of the test was 93.5%, specificity 88.6%, positive predictive value 17.3%, and negative predictive value 99.8% as compared to culture. Thus, with this method, the negative MRSA results can be reliably reported within 24-48 h from sampling. The method is a practical additional alternative to those already described for the same purpose.