Methods
MRSA isolates (n=156) from 142 patients were collected in five Latvian hospitals in Riga and Liepaja from April 2003 to February 2004. Antimicrobial susceptibility testing on these strains was performed according to National Committee for Clinical Laboratory Standards (NCCLS) standards by the disc-diffusion method and the presence of the mecA gene was verified by PCR [7]. Presence of PVL genes (lukS-lukF) and SCCmec type were tested by PCR as described earlier [8,9] in all strains. PVL-positive MRSA isolates (S-5408 and S-5690) were genotyped by multilocus restriction fragment (MLRF) [10]. In addition, the S-5408 strain was typed by PFGE [11] and multilocus sequence typing (MLST) [12]. Information about the clinical features of the disease in the patient was obtained retrospectively.

Results
Screening of 156 MRSA strains revealed two isolates harbouring genes required for the synthesis of PVL. These two isolates, S-5408 and S-5690, were cultured from catheter and nasal swab, respectively, of the same patient.

This patient, a forty six year old male with no previous clinical predisposition (immunosuppression, chronic illness, previous hospital admission), had a traumatic injury of the upper limb during construction works. Three days later he developed fatigue, swelling of the limb and fever. On the next day he was admitted to the ICU with bullous eruptions around the lips, necrotising pneumonia with pleuritic effusion, hypotension and renal failure. He reported some possible inhalation of industrial disinfectant and poisoning was suspected. Elevated WBC count and CRP levels were recorded at the time of admission. Edematous swelling of the limbs persisted during the whole treatment period within the hospital. Blood cultures were not taken but treatment with ciprofloxacin was initiated on admission. The patient gradually improved in ICU and was transferred to the nephrology unit where cultures from the tip of the central venous catheter and nasal swab were taken as a routine MRSA screening procedure. MRSA was isolated from both cultures and treatment was changed to vancomycin. The patient was discharged from hospital in a stable condition.

S-5408 and S-5690 were resistant only to oxacillin and susceptible to all other antibiotics tested (erythromycin, gentamicin, ciprofloxacin, trimethoprim/sulfamethoxazole, fusidic acid, kanamycin, vancomycin and rifampicin). It should be noted that both isolates showed low level resistance to oxacillin (MIC = 2 mg/L). In addition to the lukS-lukF genes encoding the PVL these strains carried SCCmec of type IV. Molecular analysis showed that MLRF pattern was identical in both strains but markedly different from the pattern of other MRSA isolated at the same time. The PFGE pattern of S-5408 was identical to pattern SE00-3 of MRSA isolated in Sweden from 29 patients during 2000 -2003 [13] [FIGURE]. The allelic profile (2-2-2-2-6-3-2) of two Swedish isolates typed so far and of Latvian strain S-5408 defined them as ST30 (http://www.mlst.net). This was in agreement with our analysis of the PFGE pattern (related to the pattern of strain UK EMRSA-16).

Discussion
A PVL-positive ST30-MRSA-IV was isolated from a nasal swab and the central venous catheter of a patient with fever and multiple organ failure three days after admission into ICU. This is the first MRSA with features of a community acquired strain to be isolated in Latvia.

Only nasal swab and central venous catheter cultures were available. Therefore causal relationship between the clinical symptoms and isolated bacteria has not been proven. Due to the clinical presentation the patient was suspected to have some kind of industrial poisoning and blood cultures were not taken. Retrospective analysis of the patient's clinical history and improvement on treatment with ciprofloxacin made S. aureus sepsis the most likely explanation. Colonisation of the patient by this particular MRSA strain during his brief stay in ICU seemed unlikely because the PFGE and MLRF patterns of other strains isolated from ICU patients at this time were different.

The PVL -positive CA-MRSA strain was isolated soon after the first hospital acquired MRSA strains were detected in early 2003 in Latvia. Although no country-wide surveillance existed, several hospitals had been actively testing for MRSA in previous years, with no MRSA isolate reported. This was a rather different scenario compared with what has been observed in other European countries, where hospital acquired strains appeared much earlier. There is no clear explanation as to why MRSA has emerged in Latvian hospitals so late. Most likely, epidemic strains were not imported from abroad earlier because transfer of hospitalised patients between countries was uncommon. In addition, the use of third generation cephalosporins and fluoroquinolones increased significantly only after 2001, when cheaper generic drugs became available on the market. The use of these broad-spectrum antibiotics could have facilitated the spread of MRSA strains as was suggested by other investigators [14,15].

Multilocus sequence typing attributed S-5408 and Swedish isolates with the same PFGE pattern to ST30. This is in agreement with our interpretation of the PFGE pattern as being related to that of strain UK EMRSA-16 [10,13]. Even though in the MLST database EMRSA-16 isolates are of a different sequence type, ST36, they belong to the same clonal cluster, CC30, as ST30 strains. In Europe many CA-MRSA are of ST80 [6,14] while ST30 strains are believed to be related to the South Pacific area [6]. The epidemiology of the Swedish cases is under investigation and preliminary information links at least some of them to this area. The Latvian patient had not travelled abroad but epidemiological investigation of his household contacts was not performed.

In conclusion, the PVL-positive ST30-MRSA-IV strain in Latvia is an important finding which strengthens the hypothesis of global spread of this pathogen.