Coagulase-negative staphylococci (CNS) belong to the normal microbial flora of the skin and mucous membranes of humans. The most frequently encountered CNS species in humans, in decreasing order of occurrence, are Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus saprophyticus, and Staphylococcus lugdunensis (8). CNS are an important cause of nosocomial infections, particularly causing foreign device-related infections and infections among immunocompromised patients. In a recent prospective laboratorybased surveillance in four Finnish acute-care hospitals, 76% of the blood culture CNS isolates were resistant to methicillin (meticillin) (23

In order to study the clonality of clinical methicillin-resistant Staphylococcus epidermidis (MRSE) strains and their staphylococcal cassette chromosome mec (SCCmec) elements, 60 isolates of MRSE from bacteraemic patients in three units of the Helsinki University Hospital, Finland were selected, covering the periods 1990-1993 and 1997-1998. The MRSE strains were analysed by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing and SCCmec typing. Eleven PFGE types (FIN-SE-1-11) with sequence type ST2 (clonal complex 2; CC2) were identified. The previously established methicillin-resistant Staphylococcus aureus SCCmec criteria were applied to name the MRSE SCCmec complexes, and it was found that 7% of the isolates carried SCCmec type IA (ccrA1, class B), whereas the majority (93%) yielded six non-typeable SCCmec PCR patterns (P1-P6). Within each SCCmec PCR pattern, two ccr recombinase genes (ccrA2 and ccrA3) and two mec gene complexes (class A and class B) were detected. In addition, the ccrC gene was associated with three of the six patterns. In conclusion, the MRSE strains were genetically related to each other (ST2) but their SCCmec complexes were unique combinations of elements previously recognized among SCCmec types III and IV.

Clostridium difficile infection is most often induced by antibiotic treatment. Recently, morbidity and mortality resulting especially from C. difficile PCR ribotype 027 have increased significantly. In addition, more severe disease has been associated with C. difficile PCR ribotype 078 strains. Thus, reliable typing methods for epidemic control are needed. In the present study, we compared an automated repetitive extragenic palindromic sequence-based PCR (rep-PCR) method (DiversiLab; Bacterial Barcodes, Inc., Athens, GA, USA) to PCR ribotyping and pulsed-field gel electrophoresis (PFGE) typing using 205 isolates of C. difficile (including 24 previously characterized isolates). Among the 181 clinical isolates, a total of 31 different PCR ribotypes, 38 different PFGE types and subtypes and 28 different rep-PCR types were found. Six major rep-PCR groups (DL1-DL6) harboured 86% of the clinical isolates. All isolates belonging to PCR ribotypes 027 and 001 clustered in their own rep-PCR groups, enabling us to screen out the hypervirulent ribotype 027 strain. Within the PCR ribotype 001, four subgroups were found using rep-PCR. Overall, in 75% (135/181) of the isolates, the classification attributed following rep-PCR and PCR ribotyping was comparable. In conclusion, the automated rep-PCR-based typing method represents an option for first-line molecular typing in local clinical microbiology laboratories. The method was easy to use as well as rapid, requiring less hands-on time than PCR ribotyping or PFGE typing. The conventional PCR ribotyping or PFGE, however, are needed for confirmatory molecular epidemiology. In addition, more epidemiology-oriented studies are needed to examine the discriminatory power of automated rep-PCR with isolates collected from a larger geographical area and during a longer period of time.

In Finland, the incidence of methicillin-resistant Staphylococcus aureus (MRSA) strains has increased ten fold within the last decade. In order to follow the changing epidemiology of MRSA, accurate typing of S. aureus strains is important. The purpose of this study was to reanalyse 44 previously recognised Finnish epidemic MRSA strains (EMRSA) by several molecular typing methods and to revise their nomenclature. The 44 EMRSA strains were grouped into 26 pulsed-field gel electrophoresis (PFGE) clusters, 20 multi locus sequence typing (MLST) sequence types (ST) belonging to 12 clonal complexes (CC) of which CC8 was the most prevalent, and 27 spa types belonging to four clonal complexes. The staphylococcal cassette chromosome mec (SCCmec) type IV was predominant, and 48% of the strains were nonmultiresistant to antibiotics. The discriminatory power of PFGE clusters, MLST, and spa typing was high. The overall concordance values of typing methods differed when assessed by two different methods. Adjusted Rand coefficient provided fairly low correlations for all comparisons. However, spa type was able to efficiently predict types and clonal complexes of most of the other methods with high probability (> or =80%).

BackgroundIn 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.ResultsThe 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.ConclusionThe predominant MRSA strains seem to change over time, which encourages us to continue implementing active control measures with each new MRSA case.

Our point-prevalence survey followed an outbreak of methicillin-resistant Staphylococcus aureus (MRSA) in a long-term care facility and identified five MRSA strains, of which two possessed an outbreak genotype not encountered previously and three had another profile. All of them possessed SCCmec type V. Six methicillinsensitive S. aureus strains were genotypically related to the epidemic strains.In long-term care facilities (LTCFs) methicillin-resistant Staphylococcus aureus (MRSA) strains are more often associated with colonization than clinical infection. LTCFs may also represent a reservoir of MRSA, and therefore create a two-way flow of MRSA between hospitals and LTCFs. Only a few studies have previously described the molecular epidemiology of MRSA in nursing homes (2,4,12,15,16). In all of them, MRSA strains have been identical or closely related to the strains found in hospitals in the region.An outbreak of MRSA in a health care ward and an associated nursing home of a 5,000-inhabitant municipality in northern Finland during autumn 2003 was caused by an outbreak MRSA strain of a new pulsed-field gel electrophoresis (PFGE) (FIN-22) profile and a multilocus sequence type (MLST) (ST-27) not encountered previously in Finland. With this intriguing genotype knowledge and the epidemiological fact that the LTCF is located in a remote area with limited patient transfer, we performed a point-prevalence survey 6 months later. We also assessed the molecular epidemiology of MRSA and methicillin-susceptible Staphylococcus aureus (MSSA) strains, comparing the results to the national strain collection.Setting and outbreak. A 34-bed health care ward (HCW) mainly takes care of elderly patients with multiple underlying diseases but also provides primary care, and an associated 46-bed nursing home (NH) is only for the elderly. Patient transfer from HCW to the nearby secondary-care hospital occurs approximately 0 to 2 times per week, and from the NH to the HCW and to the secondary-care hospital approximately 40 and 2 to 5 times per year, respectively.The first isolate of MRSA was found in a urine culture in the HCW in August 2003. The index patient was placed in a single room and cared for according to contact precautions. All three roommates were screened for MRSA. The following data were collected for each patient: age, sex, use of antimicrobials, foreign devices, and length of nursing stay. The screening swabs (Probact transport swab; Schofield St-Heywood, United Kingdom) were cultivated on nonselective sheep blood agar (SBA), and on selective oxacillin resistance screening agar (ORSAB CM1008, Oxoid, United Kingdom) plates. The SBA plates were incubated for 48 h and ORSAB for 96 h and inspected daily. S. aureus-like colonies were picked and subcultured onto SBA. The colonies were identified by conventional biochemical tests (8).Antimicrobial resistance to cefoxitin, cephalexin, gentamicin, tobramycin, erythromycin, clindamycin, chloramphenicol, ciprofloxacin, levofloxacin, rifampin, fusidic acid, trimethoprim-sulfamethoxazo...

Multilocus variable-number tandem repeat analysis (MLVA) is a rapid and reproducible typing method that is an important tool for investigation, as well as detection, of national and multinational outbreaks of a range of food-borne pathogens. Salmonella enterica serovar Enteritidis is the most common Salmonella serovar associated with human salmonellosis in the European Union/European Economic Area and North America. Fourteen laboratories from 13 countries in Europe and North America participated in a validation study for MLVA of S. Enteritidis targeting five loci. Following normalisation of fragment sizes using a set of reference strains, a blinded set of 24 strains with known allele sizes was analysed by each participant. The S. Enteritidis 5-loci MLVA protocol was shown to produce internationally comparable results as more than 90% of the participants reported less than 5% discrepant MLVA profiles. All 14 participating laboratories performed well, even those where experience with this typing method was limited. The raw fragment length data were consistent throughout, and the inter-laboratory validation helped to standardise the conversion of raw data to repeat numbers with at least two countries updating their internal procedures. However, differences in assigned MLVA profiles remain between well-established protocols and should be taken into account when exchanging data.

Methicillin-resistant Staphylococcus aureus (MRSA) strains from Finland covering years 1997-1999 were studied for the presence of Panton-Valentine leukocidin (PVL) gene loci, and the clinically well-defined community-acquired MRSA (CA-MRSA) strains (n = 108) also for staphylococcal chromosomal cassette mec (SCCmec) and multilocus sequence types (MLST). Only a minority (12%) of the CA-MRSA strains contained the PVL gene loci and possessed genotypes formerly described as typical to CA-MRSA strains. The majority of these strains were heterogenous by MLST and pulsed-field gel electrophoresis (PFGE) analysis but, however, harboured the SCCmec cassette type IV. In conclusion, it seems doubtful to consider only molecular characteristics such as the presence of PVL genes as definite markers for CA-MRSA strains.