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Natural history of meticillin-resistant Staphylococcus aureus colonisation among residents in community long term care facilities in Spain
Journal of Hospital Infection 76 (2010) 215e219
Available online at www.sciencedirect.com
Journal of Hospital Infection journal homepage: www.elsevierhealth.com/journals/jhin
Natural history of meticillin-resistant Staphylococcus aureus colonisation among residents in community long term care facilities in Spain A. Manzur a, *, M.A. Dominguez b, E. Ruiz de Gopegui c, D. Mariscal d, L. Gavalda e, F. Segura f, J.L. Perez c, M. Pujol a and the Spanish Network for Research in Infectious Diseasesy a
Infectious Diseases Service, Hospital Universitari de Bellvitge, Barcelona, Spain Microbiology Service, Hospital Universitari de Bellvitge, Barcelona, Spain Microbiology Service, Hospital Universitari Son Dureta, Balearic Islands, Spain d Microbiology Service, Corporació Sanitària Parc Tauli, Barcelona, Spain e Preventive Medicine, Hospital Universitari de Bellvitge, Barcelona, Spain f Infectious Diseases Service, Corporació Sanitària Parc Tauli, Barcelona, Spain b c
a r t i c l e i n f o
s u m m a r y
Article history: Received 2 November 2009 Accepted 20 May 2010 Available online 6 August 2010
The spread of meticillin-resistant Staphylococcus aureus (MRSA) is a major problem for both acute care hospitals and among residents in long term care facilities (LTCFs). We performed a cohort study to assess the natural history of MRSA colonisation in LTCF residents. Two cohorts of residents (231 MRSA carriers and 196 non-carriers) were followed up for an 18 month period, with cultures of nasal and decubitus ulcers performed every six months. In the MRSA carrier cohort, 110 (47.8%) residents had persistent MRSA colonisation for six months or longer, 44 (19.0%) had transient colonisation and nine (3.9%) were intermittently colonised. No risk factors for persistent MRSA colonisation could be determined. The annual incidence of MRSA acquisition was around 20% [95% confidence interval (CI): 14.3e25.5]. Antibiotic treatment was independently associated with MRSA acquisition (odds ratio: 2.27; 95% CI: 1.05e4.88; P ¼ 0.03). Just two clones were distinguishable by pulsed-field gel electrophor esis and multilocus sequence typing: CC5-MRSA IV, which is widely disseminated in Spanish hospitals, and ST22-MRSA IV. This study adds to the knowledge of the epidemiology of MRSA in community LTCFs, which are important components of long term care in Spain. Ó 2010 The Hospital Infection Society. Published by Elsevier Ltd. All rights reserved.
Keywords: Epidemiology Geriatrics Long term care Meticillin-resistant Staphylococcus aureus
Introduction Meticillin-resistant Staphylococcus aureus (MRSA) is endemic in most acute care hospitals and long term care facilities (LTCFs) in Spain and is a major problem for the healthcare system.1e4 Recently we found a 17% prevalence of MRSA colonisation in Spanish community LTCFs.3 In this cross-sectional study of S. aureus colonisation, we found comorbidity and size of LTCFs to be associated with meticillin-susceptible S. aureus (MSSA) colonisation; and comorbidity, age, hospital admissions, medical devices, decubitus
ulcers and antibiotic use associated with MRSA colonisation.3 Other risk factors for nasal S. aureus colonisation among nursing home residents are indwelling devices, functional dependence, low nursing staff ratio, location of LTCF in a deprived area, and prolonged hospitalisation in the previous two years.5e7 There are limited data on the natural history of MRSA colonisation in LTCFs.6,8,9 We conducted a multicentre cohort study among residents of community LTCFs for the elderly to further define the natural history of MRSA colonisation.
Methods * Corresponding author. Address: Infectious Disease Service, Hospital Universitari de Bellvitge, Feixa Llarga s/n, L’Hospitalet de Llobregat, Barcelona 08907, Spain. E-mail address: [email protected] (A. Manzur). y R. Fernández, D. Herrero, R. Casas, E. Fontseca, M. Bota, R. Iniesta, J. Alburquerque, C. Andreu, E. Campos, M. Vaqueiro, E. Antón, J. Trelis, A. Esteve, M. Canals, A. Diaz, E. Penelo, A. Oliver, J. Ariza, F. Gudiol.
Study population and characteristics of community LTCFs The characteristics of this population have been described.3 Nine community LTCFs for the elderly, located in two
0195-6701/$ e see front matter Ó 2010 The Hospital Infection Society. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.jhin.2010.05.022
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A. Manzur et al. / Journal of Hospital Infection 76 (2010) 215e219
communities in Spain (Catalonia and Balearic Islands) with 1586 beds (median: 120; range: 72e552) were included. Five were located in the catchment area of a 900 bed acute-care hospital (Hospital Universitari de Bellvitge, HUB), three in that of a 490 bed hospital (Corporació Sanitària Parc Taulí) and one in that of an 800 bed hospital (Hospital Universitari Son Dureta, HUSD). These facilities provide care for elderly long term residents, who may be disabled or infirm. Each LTCF has a dementia ward and its own medical staff. Residents are accommodated in rooms with one, two or three beds. Surveillance for MRSA and decolonisation procedures are not routine. In addition to standard precautions for all patient care, contact precautions are applied for residents colonised or infected with MRSA. Known MRSA carriers are not denied admission.10 Study design This was a multicentre prospective cohort study conducted from November 2005 to May 2007. The study population consisted of all the residents in the LTCFs at baseline (N ¼ 1377), and was identical to that of our previously published cross-sectional study.3 A total of 231 residents were found to be colonised with MRSA at baseline (MRSA carrier cohort). A representative sample of non-MRSA carriers was selected from the 1146 residents without MRSA colonisation at baseline as follows: for each MRSA carrier identified at baseline, one non-MRSA carrier was randomly selected from the same ward and screened six months later. Those with two consecutive negative cultures were included in the non-MRSA carrier cohort (N ¼ 196). Subjects were visited by the investigators every six months for an 18 month period (Figure 1). During this
MRSA CARRIER COHORT
Baseline population N = 1377
period no changes were made to infection control practices in the LTCFs and data from the study were not available to clinical staff. Data collected and definitions Complete clinical data were obtained for all residents at baseline and medical charts were reviewed thereafter at six-monthly intervals. Decubitus ulcers, hospital admissions, antibiotic consumption and clinical infections were recorded. In the MRSA carrier cohort, persistent colonisation was defined as at least two MRSA-positive cultures separated by fewer than two negative cultures. Intermittent colonisation was defined as a second positive culture separated from the first by at least two negative cultures. Transient colonisation was defined as two or more negative cultures after a single positive culture for MRSA. The prevalence of MRSA colonisation in each LTCF at baseline (17%; range: 6.7e35.8%) was used as surrogate for colonisation pressure.3 High colonisation pressure was defined as prevalence of MRSA colonisation of >20%.11,12 Microbiological methods Every six months we obtained nasal and where applicable decubitus ulcer swabs for culture. Areas were swabbed with sterile cotton-tipped applicator sticks and placed into Stuart transport medium. Swabs were plated on to coagulase mannitol agar and selective MRSA agar (MRSA Select, Bio-Rad Laboratories, Madrid, Spain), then inoculated into staphylococcal enrichment broth composed of braineheart infusion plus 7% NaCl. After 24 h of incubation at 35 C, broths were subcultured on to coagulase
NON-MRSA CARRIER COHORT
Non-MRSA carriers N = 1146
0 months
MRSA carriers N = 231
6 months
N = 178
MRSA (–) N = 196
12 months
N = 150
N = 164
18 months
End follow-up N = 127
End follow-up N = 153
Random sample N = 231
MRSA (+) N = 24
Lost N = 11
Figure 1. Residents considered and finally included at baseline and follow-up. MRSA, meticillin-resistant Staphylococcus aureus.
A. Manzur et al. / Journal of Hospital Infection 76 (2010) 215e219
mannitol agar and selective MRSA agar. Plates were incubated for 48 h and inspected daily. Putative S. aureus colonies were identified by a latex agglutination test (PastorexÒ Staph-plus, Bio-Rad) and DNase production (DNase Test Agar, Biomedics, Madrid, Spain). Testing for antimicrobial susceptibility was performed by the discdiffusion method. Meticillin resistance was determined by cefoxitin disc diffusion following Clinical and Laboratory Standards Institute recommendations.13
217
colonisation in the MRSA carrier cohort. We identified 110 (47.8%) persistent carriers, 44 (19.0%) transient carriers and nine (3.9%) intermittent carriers. The remaining 68 (29.4%) could not be classified under the definitions used. In order to identify risk factors for persistence, persistent carriers were compared with transient and intermittent carriers (Table II). Only residents with decubitus ulcers were significantly more likely to be persistent MRSA carriers, but this was not statistically significant in multivariate analysis.
Molecular epidemiology methods Macrorestriction of chromosomal DNA and pulsed-field gel electrophoresis (PFGE) analysis were performed on MRSA isolates identified in the microbiology laboratories of HUB and HUSD. Genomic DNA was digested with SmaI and restriction fragments were separated using a CHEF DR III apparatus (Bio-Rad) following published methodology, and interpreted according to Van Belkum.14,15 In a collection of representative isolates selected by their diverse PFGE patterns the following molecular studies were performed: staphylococcal cassette chromosome mec (SCCmec) polymorphism, genes encoding class S (lukS-PV) and class F (lukF-PV) proteins for PantoneValentine leucocidin (PVL), and multilocus sequence typing (MLST).16e18
Incidence of MRSA colonisation in the cohort not colonised at baseline Among 196 residents with negative cultures for MRSA at baseline and at 6 months, 43 were colonised with MSSA and 153 were not colonised with S. aureus. Table III compares characteristics of residents in the non-MRSA carrier cohort who were lost to followup with those who were followed to completion of the study. Thirty-nine residents acquired MRSA while in the LTCF. The incidence of MRSA colonisation during LTCF stay was 19.9% [95% confidence interval (CI): 14.3e25.5] per year. Antibiotic use was independently associated with acquisition of MRSA (odds ratio: 2.12; 95% CI: 1.05e4.45; P ¼ 0.05) (Table IV).
Statistical analysis
Molecular epidemiology
Categorical variables were analysed with c2-test or Fisher’s test as appropriate and continuous variables by Student’s t-test or nonparametric tests. Logistic regression models were used to analyse risk factors for persistence in the MRSA carrier cohort and risk factors for acquisition in the non-MRSA carrier cohort. Variables with P < 0.15 were included in the model. All statistical tests were two-tailed and P < 0.05 was deemed significant. SPSS package version 12.0 was used. Approval for study was obtained from the Research Ethics Committee of HUB. No written informed consent was obtained since the study met criteria for a waiver of this requirement.
Among 491 residents of five LTCFs related to HUB, MRSA was isolated from 144 (276 MRSA isolates), 108 of whom were identified at baseline. All 276 MRSA isolates were studied by PFGE and 88% belonged to a single clone. Representative strains of this dominant clone belonged to ST146, single locus variant by MLST typing of ST5, founder of clonal complex 5. All belonged to SCCmec type IV and were PVL negative. Among 466 residents of one LTCF related to HUSD, 39 MRSA isolates were identified at baseline. Two different strains, clone A (three subtypes) and clone C (five subtypes) were recognised by PFGE. Clone A belongs to ST125 MRSA-IV and clone B to ST22-MRSA IV. All strains were PVL negative and carried SCCmecIV. Subsequent MRSA isolates had the same PFGE profiles as the initial isolates.
Results
Discussion
Natural history of MRSA colonisation in the cohort colonised at baseline A total of 231 residents colonised with MRSA at baseline were included. Of these, 104 were lost to follow-up; 70 died, 21 were discharged and 13 lost for other reasons. Table I compares residents who were lost to follow-up with those in this cohort followed to completion. Figure 2 shows the natural history of MRSA
Table I Baseline characteristics of patients who did and did not complete the follow-up period in the MRSA carrier cohort Characteristic Female sex Age (years)a Charlson index 2 Barthel index <30 Medical devices Decubitus ulcers LTCFs with <150 beds LTCFs with prevalence of MRSA >20% Two-site colonisation
Completed follow-up N ¼ 127
Lost to follow-up N ¼ 104
P
88 (69.3%) 81.6 (12.6) 50 (39.4%) 49 (38.6%) 10 (7.9%) 30 (23.6%) 56 (44.1%) 82 (64.6%)
71 (68.3%) 83.3 (10.1) 53 (50.9%) 50 (48.1%) 21 (20.2%) 28 (26.9%) 37 (35.6%) 55 (52.9%)
0.87 0.52 0.08 0.15 0.006 0.56 0.19 0.07
17 (13.4%)
17 (16.3%)
0.53
MRSA, meticillin-resistant Staphylococcus aureus; LTCF, long term care facility. a Mean (SD).
We have previously reported a prevalence of MRSA colonisation of w17% among residents of community LTCFs in Spain.3 This is similar to reports from other European countries, but is lower than the prevalence reported in Veterans Affairs facilities in the USA.7,19e22 There is little information on the dynamics of MRSA colonisation in residents of community LTCFs. Most of the carriers we identified had persistent MRSA colonisation. This agrees with studies gathered in Veterans Affairs facilities, albeit with different epidemiological features.6 We did not identify any factor that distinguished persistent MRSA carriers from transient and intermittent carriers. Stone et al. found that only bacterial burden could distinguish the groups.23 Defining the duration of colonisation is problematic, because low bacterial burdens lead to false-negative results and the limited number of samples collected from residents could contribute towards the misclassification of colonisation status. There was a high incidence of MRSA colonisation in this study, w20% per annum. Only antibiotic use was independently associated with MRSA acquisition in LTCFs. We used the prevalence of MRSA colonisation in each LTCF at baseline as a surrogate for colonisation pressure. This is regarded as an important determinant of the probability of acquisition when organisms spread mainly via cross-colonisation.11,12 In our population, the probability of MRSA
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MRSA colonised patients
62
Negative cultures
44
MRSA colonisation for 18 months
9
MRSA colonisation for 12 months
10
MRSA colonisation for 18 months
18
MRSA colonisation for 6 months
5
MRSA colonisation for 18 months
6
Intermittent MRSA colonisation
6
Transient MRSA colonisation Recolonisation after 18 months
105 30 231
14
73
44
29 Transient MRSA colonisation
0 month N = 231
6 months N = 178
12 months N = 150
18 months N = 127
Figure 2. Natural history of meticillin-resistant Staphylococcus aureus (MRSA) colonisation in the MRSA carrier cohort. Results of MRSA detection performed at each point of the study. Subjects lost to follow-up are not included.
acquisition was no greater in LTCFs with a high baseline MRSA prevalence, which we defined as >20%. Perhaps the baseline prevalence of MRSA colonisation in an LTCF did not represent adequately the colonisation pressure throughout the study period. The spread of MRSA in community LTCFs increases the reservoir of this organism in the healthcare setting, and residents transferred from LTCFs account for a large part of the burden of hospitalised MRSA carriers. Programmes for detection of carriers at hospital admission should take account of this.24e26 Regarding the molecular epidemiology results, ST146 and ST125 are single locus variants of ST5, and therefore both belong to CC5. Remarkably, only two distinct clones could be distinguished by
Table II Risk factors for persistent MRSA colonisation among residents in community long term care facilities Transient or Persistent Bivariate intermittent MRSA analysis colonisation colonisation P N ¼ 53 N ¼ 110 Aged >85 years Female sex Charlson index 2 Barthel index <30 Decubitus ulcers Two-site colonisation Hospital admissions Antibiotic use LTCFs with MRSA colonisation prevalence >20%
22 (41.5%) 43 (81.1%) 16 (30.8%) 23 (34.8%) 7 (13.7%) 3 (9.7%) 6 (11.5%) 28 (53.8%) 30 (57.7%)
45 73 46 43 30 17 23 67 73
(40.9%) (66.4%) (42.6%) (39.8%) (27.5%) (19.1%) (21.1%) (61.5%) (67.0%)
0.94 0.06 0.15 0.59 0.05 0.23 0.14 0.36 0.25
Logistic regression OR (95% CI) 0.99 (0.96e1.02) 2.02 (0.91e4.50) 0.55 (0.28e1.08) 0.51 (0.22e1.15) 0.61 (0.55e1.44)
MRSA, meticillin-resistant Staphylococcus aureus; OR, odds ratio; CI, confidence interval; LTCF, long term care facility.
PFGE and MLST: CC5-MRSA IV and ST22-MRSA IV. The former has been reported to be widely disseminated in Spanish hospitals.27,28 The dominance of clone ST146-MRSA-IV made it impossible to differentiate between persistent colonisation and recurrent acquisition. Our study has a number of limitations. Apart from nasal swabs, we cultured swabs only from decubitus ulcers, the most frequent skin lesion. High levels of MRSA colonisation in decubitus ulcers have been reported among LTCF residents.9,21,29 However, recent results suggest that most community LTCF residents with MRSA are colonised at multiple sites. Some colonised residents are not detected unless multiple sites are sampled and one-third are missed if only a nasal swab is collected.8 We did not collect data on potential confounders such as nursing staff:bed ratio, invasive devices or functional status of residents. Functional status is likely to have been poorer in those lost to follow-up, because deaths
Table III Baseline characteristics of residents who did and did not complete follow-up in the non-MRSA carrier cohort Characteristic Female sex Age (years)a Charlson index 2 Barthel index <30 Decubitus ulcers LTCFs with <150 beds LTCFs with MRSA prevalence >20%
Completed follow-up N ¼ 153 103 (67.3%) 80.4 (11.1) 49 (32.0%) 52 (34.2%) 10 (6.5%) 53 (34.6%) 90 (58.8%)
Lost to follow-up N ¼ 43
P
31 (72.1%) 83.4 (11.6) 17 (39.5%) 25 (58.1%) 7 (16.3%) 23 (46.5%) 33 (76.7%)
0.55 0.89 0.36 0.005 0.05 0.16 0.03
MRSA, meticillin-resistant Staphylococcus aureus; LTCF, long term care facility. a Mean (SD).
A. Manzur et al. / Journal of Hospital Infection 76 (2010) 215e219 Table IV Risk factors associated with MRSA acquisition in community long term care facilities
Age (years)a Female sex Charlson index 2 Barthel index <30 Decubitus ulcers Hospital admissions Antibiotic use LTCFs with MRSA prevalence >20%
New MRSA carriers N ¼ 39
Non-MRSA carriers N ¼ 157
Bivariate analysis P
Logistic regression OR (95% CI)
78.6 (12.3) 24 (61.5%) 14 (35.9%) 18 (46.2%) 6 (15.4%) 6 (15.4%) 25 (64.1%) 23 (59.0%)
81.6 (10.9) 110 (70.1%) 55 (33.1%) 59 (37.8%) 11 (7.0%) 21 (13.4%) 69 (43.9%) 100 (63.7%)
0.88 0.31 0.73 0.34 0.09 0.75 0.02 0.59
0.98 (0.95e1.01) 0.74 (0.34e1.64)
2.17 (0.73e6.52) 2.12 (1.05e4.45)
MRSA, meticillin-resistant Staphylococcus aureus; OR, odds ratio; CI, confidence interval; LTCF, long term care facility. a Mean (SD).
accounted for most of them, and poor functional status is associated with earlier death in this population.30 Few longitudinal studies have assessed the incidence of MRSA colonisation among residents of LTCFs.6 This multicentre prospective study adds to existing knowledge of natural history of MRSA in community LTCFs and may inform more effective control measures. Conflict of interest statement None declared. Funding source Supported by Ministerio de Sanidad y Consumo, Instituto de Salud Carlos III, Spanish Network for the Research in Infectious Diseases (REIPI RD06/0008). References 1. Cretnik TZ, Vovko P, Retelj M, et al. Prevalence and nosocomial spread of methicillin-resistant Staphylococcus aureus in a long-term-care facility in Slovenia. Infect Control Hosp Epidemiol 2005;26:184e190. 2. Dominguez MA, De Lencastre H, Liñares J, Tomasz A. Spread and maintenance of a dominant methicillin-resistant Staphylococcus aureus (MRSA) clone during an outbreak of MRSA disease in a Spanish hospital. J Clin Microbiol 1994;32:2081e2087. 3. Manzur A, Gavalda L, Ruiz de Gopegui E, et al. Prevalence of methicillinresistant Staphylococcus aureus and factors associated with colonization among residents in community long-term-care facilities in Spain. Clin Microbiol Infect 2008;14:867e872. 4. Olona-Cabases M, Tico-Falguera N, Ramírez-Garceran L, Del Valle-Ortiz O, Castello-Verdu T, García-Fernandez L. Methicillin-resistant Staphylococcus aureus: a four-year experience in a spinal cord injury unit in Spain. Spinal Cord 1996;34:315e319. 5. Mody L, Maheshwari S, Galecki A, et al. Indwelling device use and antibiotic resistance in nursing homes: identifying a high-risk group. J Am Geriatr Soc 2007;55:1921e1926. 6. Bradley SF, Terpenning MS, Ramsey MA, et al. Methicillin-resistant Staphylococcus aureus: colonization and infection in a long-term-care facility. Ann Intern Med 1991;115:417e422. 7. Barr B, Wilcox MH, Brady A, Parnell P, Darby B, Tompkins D. Prevalence of methicillin-resistant Staphylococcus aureus colonization among older residents of care homes in the United Kingdom. Infect Control Hosp Epidemiol 2007;28:853e859. 8. Mody L, Kauffman CA, Donabedian S, Zervos M, Bradley SF. Epidemiology of Staphylococcus aureus colonization in nursing home residents. Clin Infect Dis 2008;46:1368e1373.
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9. Muder RR, Brennen C, Wagener MM, et al. Methicillin-resistant staphylococcal colonization and infection in a long-term care facility. Ann Intern Med 1991;114:107e112. 10. Rodríguez-Baño J, Bischofberger C, Alvarez-Lerma F, et al. Surveillance and control of methicillin-resistant Staphylococcus aureus in Spanish hospitals. A GEIH-SEIMC and SEMPSPH consensus document. Enferm Infecc Microbiol Clin 2008;26:285e298. 11. Bonten M, Slaughter S, Ambergen AW, et al. The role of “colonization pressure” in the spread of vancomycin-resistant enterococci. An important infection control variable. Arch Intern Med 1998;158:1127e1132. 12. Merrer J, Santoli F, Appéré-De Vecchi C, Tran B, De Jonghe B, Outin H. “Colonization pressure” and risk of acquisition of methicillin-resistant Staphylococcus aureus in a medical intensive care unit. Infect Control Hosp Epidemiol 2000;21:718e723. 13. Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing; 16th informational supplement. CLSI document M100-S16. Wayne, PA: Clinical and Laboratory Standards Institute; 2006. 14. Chung M, De Lencastre H, Matthews P, et al. Molecular typing of methicillinresistant Staphylococcus aureus (MRSA) by pulsed field gel electrophoresis: comparison of results obtained in a multilaboratory effort using identical protocols and MRSA strains. Microb Drug Resist 2000;6:189e197. 15. Van Belkum A, Tassios PT, Dijkshoorn L, et al. Guidelines for the validation and application of typing methods for use in bacterial epidemiology. Clin Microbiol Infect 2007;13:1e46. 16. Milheiriço C, Oliveira DC, De Lencastre H. Multiplex PCR strategy for subtyping the staphylococcal cassette chromosome mec type IV in methicillin-resistant Staphylococcus aureus: ‘SCCmec IV multiplex’. J Antimicrob Chemother 2007;60:42e48. 17. Lina GM, Piemont Y, Godail-Gamot F, et al. Involvement of PantoneValentine leukocidin-producing Staphylococcus aureus in primary skin infections and pneumonia. Clin Infect Dis 1999;29:1128e1132. 18. Enright MC, Day NP, Davies CE, Peacock SJ, Spratt BG. Multilocus sequence typing for characterization of methicillin-resistant and methicillin-susceptible clones of Staphylococcus aureus. J Clin Microbiol 2000;38:1008e1015. 19. O’Sullivan NP, Keane CT. The prevalence of methicillin-resistant Staphylococcus aureus among the residents of six nursing homes for the elderly. J Hosp Infect 2000;45:322e329. 20. von Baum H, Schmidt C, Svoboda D, Bock-Hensley O, Wendt C. Risk factors for methicillin-resistant Staphylococcus aureus carriage in residents of German nursing homes. Infect Control Hosp Epidemiol 2002;23:511e515. 21. Bradley SF. Methicillin-resistant Staphylococcus aureus: long-term care concerns. Am J Med 1999;106:2Se10S. 22. Mulhausen PL, Harrell LJ, Weinberger M, Kochersberger GG, Feussner JR. Contrasting methicillin-resistant Staphylococcus aureus colonization in veteran’s affairs and community nursing homes. Am J Med 1996;100:24e31. 23. Stone ND, Lewis DR, Lowery HK, et al. Importance of bacterial burden among methicillin-resistant Staphylococcus aureus carriers in a long-term care facility. Infect Control Hosp Epidemiol 2008;29:143e148. 24. Manzur A, Vidal M, Pujol M, et al. Predictive factors of meticillin resistance among patients with Staphylococcus aureus bloodstream infection at hospital admission. J Hosp Infect 2007;66:135e141. 25. Gavalda L, Masuet C, Beltran J, et al. Comparative cost of selective screening to prevent transmission of methicillin resistant Staphylococcus aureus (MRSA), compared with the attributable costs of MRSA infection. Infect Control Hosp Epidemiol 2006;27:1264e1266. 26. Cooper BS, Medley GF, Stone SP, et al. Methicillin-resistant Staphylococcus aureus in hospitals and the community: stealth dynamics and control catastrophes. Proc Natl Acad Sci USA 2004;101:10223e10228. 27. Vindel A, Cuevas O, Cercenado E, et al. Methicillin-resistant Staphylococcus aureus in Spain: molecular epidemiology and utility of different typing methods. J Clin Microbiol 2009;47:1620e1627. 28. Rodríguez-Baño J, Domínguez M, Millán A, et al. Clinical and molecular epidemiology of community-acquired, healthcare-associated and nosocomial methicillin-resistant Staphylococcus aureus in Spain. Clin Microbiol Infect 2009;15:1111e1118. 29. Ruiz de Gopegui E, Oliver A, Ramírez A, Gutiérrez O, Andreu C, Pérez JL. Epidemiological relatedness of methicillin-resistant Staphylococcus aureus from a tertiary hospital and a geriatric institution in Spain. Clin Microbiol Infect 2004;10:339e342. 30. Mahoney F, Barthel DW. Functional evaluation: the Barthel index. Md State Med J 1965;14:61e65.
Available online at www.sciencedirect.com
Journal of Hospital Infection journal homepage: www.elsevierhealth.com/journals/jhin
Natural history of meticillin-resistant Staphylococcus aureus colonisation among residents in community long term care facilities in Spain A. Manzur a, *, M.A. Dominguez b, E. Ruiz de Gopegui c, D. Mariscal d, L. Gavalda e, F. Segura f, J.L. Perez c, M. Pujol a and the Spanish Network for Research in Infectious Diseasesy a
Infectious Diseases Service, Hospital Universitari de Bellvitge, Barcelona, Spain Microbiology Service, Hospital Universitari de Bellvitge, Barcelona, Spain Microbiology Service, Hospital Universitari Son Dureta, Balearic Islands, Spain d Microbiology Service, Corporació Sanitària Parc Tauli, Barcelona, Spain e Preventive Medicine, Hospital Universitari de Bellvitge, Barcelona, Spain f Infectious Diseases Service, Corporació Sanitària Parc Tauli, Barcelona, Spain b c
a r t i c l e i n f o
s u m m a r y
Article history: Received 2 November 2009 Accepted 20 May 2010 Available online 6 August 2010
The spread of meticillin-resistant Staphylococcus aureus (MRSA) is a major problem for both acute care hospitals and among residents in long term care facilities (LTCFs). We performed a cohort study to assess the natural history of MRSA colonisation in LTCF residents. Two cohorts of residents (231 MRSA carriers and 196 non-carriers) were followed up for an 18 month period, with cultures of nasal and decubitus ulcers performed every six months. In the MRSA carrier cohort, 110 (47.8%) residents had persistent MRSA colonisation for six months or longer, 44 (19.0%) had transient colonisation and nine (3.9%) were intermittently colonised. No risk factors for persistent MRSA colonisation could be determined. The annual incidence of MRSA acquisition was around 20% [95% confidence interval (CI): 14.3e25.5]. Antibiotic treatment was independently associated with MRSA acquisition (odds ratio: 2.27; 95% CI: 1.05e4.88; P ¼ 0.03). Just two clones were distinguishable by pulsed-field gel electrophor esis and multilocus sequence typing: CC5-MRSA IV, which is widely disseminated in Spanish hospitals, and ST22-MRSA IV. This study adds to the knowledge of the epidemiology of MRSA in community LTCFs, which are important components of long term care in Spain. Ó 2010 The Hospital Infection Society. Published by Elsevier Ltd. All rights reserved.
Keywords: Epidemiology Geriatrics Long term care Meticillin-resistant Staphylococcus aureus
Introduction Meticillin-resistant Staphylococcus aureus (MRSA) is endemic in most acute care hospitals and long term care facilities (LTCFs) in Spain and is a major problem for the healthcare system.1e4 Recently we found a 17% prevalence of MRSA colonisation in Spanish community LTCFs.3 In this cross-sectional study of S. aureus colonisation, we found comorbidity and size of LTCFs to be associated with meticillin-susceptible S. aureus (MSSA) colonisation; and comorbidity, age, hospital admissions, medical devices, decubitus
ulcers and antibiotic use associated with MRSA colonisation.3 Other risk factors for nasal S. aureus colonisation among nursing home residents are indwelling devices, functional dependence, low nursing staff ratio, location of LTCF in a deprived area, and prolonged hospitalisation in the previous two years.5e7 There are limited data on the natural history of MRSA colonisation in LTCFs.6,8,9 We conducted a multicentre cohort study among residents of community LTCFs for the elderly to further define the natural history of MRSA colonisation.
Methods * Corresponding author. Address: Infectious Disease Service, Hospital Universitari de Bellvitge, Feixa Llarga s/n, L’Hospitalet de Llobregat, Barcelona 08907, Spain. E-mail address: [email protected] (A. Manzur). y R. Fernández, D. Herrero, R. Casas, E. Fontseca, M. Bota, R. Iniesta, J. Alburquerque, C. Andreu, E. Campos, M. Vaqueiro, E. Antón, J. Trelis, A. Esteve, M. Canals, A. Diaz, E. Penelo, A. Oliver, J. Ariza, F. Gudiol.
Study population and characteristics of community LTCFs The characteristics of this population have been described.3 Nine community LTCFs for the elderly, located in two
0195-6701/$ e see front matter Ó 2010 The Hospital Infection Society. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.jhin.2010.05.022
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communities in Spain (Catalonia and Balearic Islands) with 1586 beds (median: 120; range: 72e552) were included. Five were located in the catchment area of a 900 bed acute-care hospital (Hospital Universitari de Bellvitge, HUB), three in that of a 490 bed hospital (Corporació Sanitària Parc Taulí) and one in that of an 800 bed hospital (Hospital Universitari Son Dureta, HUSD). These facilities provide care for elderly long term residents, who may be disabled or infirm. Each LTCF has a dementia ward and its own medical staff. Residents are accommodated in rooms with one, two or three beds. Surveillance for MRSA and decolonisation procedures are not routine. In addition to standard precautions for all patient care, contact precautions are applied for residents colonised or infected with MRSA. Known MRSA carriers are not denied admission.10 Study design This was a multicentre prospective cohort study conducted from November 2005 to May 2007. The study population consisted of all the residents in the LTCFs at baseline (N ¼ 1377), and was identical to that of our previously published cross-sectional study.3 A total of 231 residents were found to be colonised with MRSA at baseline (MRSA carrier cohort). A representative sample of non-MRSA carriers was selected from the 1146 residents without MRSA colonisation at baseline as follows: for each MRSA carrier identified at baseline, one non-MRSA carrier was randomly selected from the same ward and screened six months later. Those with two consecutive negative cultures were included in the non-MRSA carrier cohort (N ¼ 196). Subjects were visited by the investigators every six months for an 18 month period (Figure 1). During this
MRSA CARRIER COHORT
Baseline population N = 1377
period no changes were made to infection control practices in the LTCFs and data from the study were not available to clinical staff. Data collected and definitions Complete clinical data were obtained for all residents at baseline and medical charts were reviewed thereafter at six-monthly intervals. Decubitus ulcers, hospital admissions, antibiotic consumption and clinical infections were recorded. In the MRSA carrier cohort, persistent colonisation was defined as at least two MRSA-positive cultures separated by fewer than two negative cultures. Intermittent colonisation was defined as a second positive culture separated from the first by at least two negative cultures. Transient colonisation was defined as two or more negative cultures after a single positive culture for MRSA. The prevalence of MRSA colonisation in each LTCF at baseline (17%; range: 6.7e35.8%) was used as surrogate for colonisation pressure.3 High colonisation pressure was defined as prevalence of MRSA colonisation of >20%.11,12 Microbiological methods Every six months we obtained nasal and where applicable decubitus ulcer swabs for culture. Areas were swabbed with sterile cotton-tipped applicator sticks and placed into Stuart transport medium. Swabs were plated on to coagulase mannitol agar and selective MRSA agar (MRSA Select, Bio-Rad Laboratories, Madrid, Spain), then inoculated into staphylococcal enrichment broth composed of braineheart infusion plus 7% NaCl. After 24 h of incubation at 35 C, broths were subcultured on to coagulase
NON-MRSA CARRIER COHORT
Non-MRSA carriers N = 1146
0 months
MRSA carriers N = 231
6 months
N = 178
MRSA (–) N = 196
12 months
N = 150
N = 164
18 months
End follow-up N = 127
End follow-up N = 153
Random sample N = 231
MRSA (+) N = 24
Lost N = 11
Figure 1. Residents considered and finally included at baseline and follow-up. MRSA, meticillin-resistant Staphylococcus aureus.
A. Manzur et al. / Journal of Hospital Infection 76 (2010) 215e219
mannitol agar and selective MRSA agar. Plates were incubated for 48 h and inspected daily. Putative S. aureus colonies were identified by a latex agglutination test (PastorexÒ Staph-plus, Bio-Rad) and DNase production (DNase Test Agar, Biomedics, Madrid, Spain). Testing for antimicrobial susceptibility was performed by the discdiffusion method. Meticillin resistance was determined by cefoxitin disc diffusion following Clinical and Laboratory Standards Institute recommendations.13
217
colonisation in the MRSA carrier cohort. We identified 110 (47.8%) persistent carriers, 44 (19.0%) transient carriers and nine (3.9%) intermittent carriers. The remaining 68 (29.4%) could not be classified under the definitions used. In order to identify risk factors for persistence, persistent carriers were compared with transient and intermittent carriers (Table II). Only residents with decubitus ulcers were significantly more likely to be persistent MRSA carriers, but this was not statistically significant in multivariate analysis.
Molecular epidemiology methods Macrorestriction of chromosomal DNA and pulsed-field gel electrophoresis (PFGE) analysis were performed on MRSA isolates identified in the microbiology laboratories of HUB and HUSD. Genomic DNA was digested with SmaI and restriction fragments were separated using a CHEF DR III apparatus (Bio-Rad) following published methodology, and interpreted according to Van Belkum.14,15 In a collection of representative isolates selected by their diverse PFGE patterns the following molecular studies were performed: staphylococcal cassette chromosome mec (SCCmec) polymorphism, genes encoding class S (lukS-PV) and class F (lukF-PV) proteins for PantoneValentine leucocidin (PVL), and multilocus sequence typing (MLST).16e18
Incidence of MRSA colonisation in the cohort not colonised at baseline Among 196 residents with negative cultures for MRSA at baseline and at 6 months, 43 were colonised with MSSA and 153 were not colonised with S. aureus. Table III compares characteristics of residents in the non-MRSA carrier cohort who were lost to followup with those who were followed to completion of the study. Thirty-nine residents acquired MRSA while in the LTCF. The incidence of MRSA colonisation during LTCF stay was 19.9% [95% confidence interval (CI): 14.3e25.5] per year. Antibiotic use was independently associated with acquisition of MRSA (odds ratio: 2.12; 95% CI: 1.05e4.45; P ¼ 0.05) (Table IV).
Statistical analysis
Molecular epidemiology
Categorical variables were analysed with c2-test or Fisher’s test as appropriate and continuous variables by Student’s t-test or nonparametric tests. Logistic regression models were used to analyse risk factors for persistence in the MRSA carrier cohort and risk factors for acquisition in the non-MRSA carrier cohort. Variables with P < 0.15 were included in the model. All statistical tests were two-tailed and P < 0.05 was deemed significant. SPSS package version 12.0 was used. Approval for study was obtained from the Research Ethics Committee of HUB. No written informed consent was obtained since the study met criteria for a waiver of this requirement.
Among 491 residents of five LTCFs related to HUB, MRSA was isolated from 144 (276 MRSA isolates), 108 of whom were identified at baseline. All 276 MRSA isolates were studied by PFGE and 88% belonged to a single clone. Representative strains of this dominant clone belonged to ST146, single locus variant by MLST typing of ST5, founder of clonal complex 5. All belonged to SCCmec type IV and were PVL negative. Among 466 residents of one LTCF related to HUSD, 39 MRSA isolates were identified at baseline. Two different strains, clone A (three subtypes) and clone C (five subtypes) were recognised by PFGE. Clone A belongs to ST125 MRSA-IV and clone B to ST22-MRSA IV. All strains were PVL negative and carried SCCmecIV. Subsequent MRSA isolates had the same PFGE profiles as the initial isolates.
Results
Discussion
Natural history of MRSA colonisation in the cohort colonised at baseline A total of 231 residents colonised with MRSA at baseline were included. Of these, 104 were lost to follow-up; 70 died, 21 were discharged and 13 lost for other reasons. Table I compares residents who were lost to follow-up with those in this cohort followed to completion. Figure 2 shows the natural history of MRSA
Table I Baseline characteristics of patients who did and did not complete the follow-up period in the MRSA carrier cohort Characteristic Female sex Age (years)a Charlson index 2 Barthel index <30 Medical devices Decubitus ulcers LTCFs with <150 beds LTCFs with prevalence of MRSA >20% Two-site colonisation
Completed follow-up N ¼ 127
Lost to follow-up N ¼ 104
P
88 (69.3%) 81.6 (12.6) 50 (39.4%) 49 (38.6%) 10 (7.9%) 30 (23.6%) 56 (44.1%) 82 (64.6%)
71 (68.3%) 83.3 (10.1) 53 (50.9%) 50 (48.1%) 21 (20.2%) 28 (26.9%) 37 (35.6%) 55 (52.9%)
0.87 0.52 0.08 0.15 0.006 0.56 0.19 0.07
17 (13.4%)
17 (16.3%)
0.53
MRSA, meticillin-resistant Staphylococcus aureus; LTCF, long term care facility. a Mean (SD).
We have previously reported a prevalence of MRSA colonisation of w17% among residents of community LTCFs in Spain.3 This is similar to reports from other European countries, but is lower than the prevalence reported in Veterans Affairs facilities in the USA.7,19e22 There is little information on the dynamics of MRSA colonisation in residents of community LTCFs. Most of the carriers we identified had persistent MRSA colonisation. This agrees with studies gathered in Veterans Affairs facilities, albeit with different epidemiological features.6 We did not identify any factor that distinguished persistent MRSA carriers from transient and intermittent carriers. Stone et al. found that only bacterial burden could distinguish the groups.23 Defining the duration of colonisation is problematic, because low bacterial burdens lead to false-negative results and the limited number of samples collected from residents could contribute towards the misclassification of colonisation status. There was a high incidence of MRSA colonisation in this study, w20% per annum. Only antibiotic use was independently associated with MRSA acquisition in LTCFs. We used the prevalence of MRSA colonisation in each LTCF at baseline as a surrogate for colonisation pressure. This is regarded as an important determinant of the probability of acquisition when organisms spread mainly via cross-colonisation.11,12 In our population, the probability of MRSA
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MRSA colonised patients
62
Negative cultures
44
MRSA colonisation for 18 months
9
MRSA colonisation for 12 months
10
MRSA colonisation for 18 months
18
MRSA colonisation for 6 months
5
MRSA colonisation for 18 months
6
Intermittent MRSA colonisation
6
Transient MRSA colonisation Recolonisation after 18 months
105 30 231
14
73
44
29 Transient MRSA colonisation
0 month N = 231
6 months N = 178
12 months N = 150
18 months N = 127
Figure 2. Natural history of meticillin-resistant Staphylococcus aureus (MRSA) colonisation in the MRSA carrier cohort. Results of MRSA detection performed at each point of the study. Subjects lost to follow-up are not included.
acquisition was no greater in LTCFs with a high baseline MRSA prevalence, which we defined as >20%. Perhaps the baseline prevalence of MRSA colonisation in an LTCF did not represent adequately the colonisation pressure throughout the study period. The spread of MRSA in community LTCFs increases the reservoir of this organism in the healthcare setting, and residents transferred from LTCFs account for a large part of the burden of hospitalised MRSA carriers. Programmes for detection of carriers at hospital admission should take account of this.24e26 Regarding the molecular epidemiology results, ST146 and ST125 are single locus variants of ST5, and therefore both belong to CC5. Remarkably, only two distinct clones could be distinguished by
Table II Risk factors for persistent MRSA colonisation among residents in community long term care facilities Transient or Persistent Bivariate intermittent MRSA analysis colonisation colonisation P N ¼ 53 N ¼ 110 Aged >85 years Female sex Charlson index 2 Barthel index <30 Decubitus ulcers Two-site colonisation Hospital admissions Antibiotic use LTCFs with MRSA colonisation prevalence >20%
22 (41.5%) 43 (81.1%) 16 (30.8%) 23 (34.8%) 7 (13.7%) 3 (9.7%) 6 (11.5%) 28 (53.8%) 30 (57.7%)
45 73 46 43 30 17 23 67 73
(40.9%) (66.4%) (42.6%) (39.8%) (27.5%) (19.1%) (21.1%) (61.5%) (67.0%)
0.94 0.06 0.15 0.59 0.05 0.23 0.14 0.36 0.25
Logistic regression OR (95% CI) 0.99 (0.96e1.02) 2.02 (0.91e4.50) 0.55 (0.28e1.08) 0.51 (0.22e1.15) 0.61 (0.55e1.44)
MRSA, meticillin-resistant Staphylococcus aureus; OR, odds ratio; CI, confidence interval; LTCF, long term care facility.
PFGE and MLST: CC5-MRSA IV and ST22-MRSA IV. The former has been reported to be widely disseminated in Spanish hospitals.27,28 The dominance of clone ST146-MRSA-IV made it impossible to differentiate between persistent colonisation and recurrent acquisition. Our study has a number of limitations. Apart from nasal swabs, we cultured swabs only from decubitus ulcers, the most frequent skin lesion. High levels of MRSA colonisation in decubitus ulcers have been reported among LTCF residents.9,21,29 However, recent results suggest that most community LTCF residents with MRSA are colonised at multiple sites. Some colonised residents are not detected unless multiple sites are sampled and one-third are missed if only a nasal swab is collected.8 We did not collect data on potential confounders such as nursing staff:bed ratio, invasive devices or functional status of residents. Functional status is likely to have been poorer in those lost to follow-up, because deaths
Table III Baseline characteristics of residents who did and did not complete follow-up in the non-MRSA carrier cohort Characteristic Female sex Age (years)a Charlson index 2 Barthel index <30 Decubitus ulcers LTCFs with <150 beds LTCFs with MRSA prevalence >20%
Completed follow-up N ¼ 153 103 (67.3%) 80.4 (11.1) 49 (32.0%) 52 (34.2%) 10 (6.5%) 53 (34.6%) 90 (58.8%)
Lost to follow-up N ¼ 43
P
31 (72.1%) 83.4 (11.6) 17 (39.5%) 25 (58.1%) 7 (16.3%) 23 (46.5%) 33 (76.7%)
0.55 0.89 0.36 0.005 0.05 0.16 0.03
MRSA, meticillin-resistant Staphylococcus aureus; LTCF, long term care facility. a Mean (SD).
A. Manzur et al. / Journal of Hospital Infection 76 (2010) 215e219 Table IV Risk factors associated with MRSA acquisition in community long term care facilities
Age (years)a Female sex Charlson index 2 Barthel index <30 Decubitus ulcers Hospital admissions Antibiotic use LTCFs with MRSA prevalence >20%
New MRSA carriers N ¼ 39
Non-MRSA carriers N ¼ 157
Bivariate analysis P
Logistic regression OR (95% CI)
78.6 (12.3) 24 (61.5%) 14 (35.9%) 18 (46.2%) 6 (15.4%) 6 (15.4%) 25 (64.1%) 23 (59.0%)
81.6 (10.9) 110 (70.1%) 55 (33.1%) 59 (37.8%) 11 (7.0%) 21 (13.4%) 69 (43.9%) 100 (63.7%)
0.88 0.31 0.73 0.34 0.09 0.75 0.02 0.59
0.98 (0.95e1.01) 0.74 (0.34e1.64)
2.17 (0.73e6.52) 2.12 (1.05e4.45)
MRSA, meticillin-resistant Staphylococcus aureus; OR, odds ratio; CI, confidence interval; LTCF, long term care facility. a Mean (SD).
accounted for most of them, and poor functional status is associated with earlier death in this population.30 Few longitudinal studies have assessed the incidence of MRSA colonisation among residents of LTCFs.6 This multicentre prospective study adds to existing knowledge of natural history of MRSA in community LTCFs and may inform more effective control measures. Conflict of interest statement None declared. Funding source Supported by Ministerio de Sanidad y Consumo, Instituto de Salud Carlos III, Spanish Network for the Research in Infectious Diseases (REIPI RD06/0008). References 1. Cretnik TZ, Vovko P, Retelj M, et al. Prevalence and nosocomial spread of methicillin-resistant Staphylococcus aureus in a long-term-care facility in Slovenia. Infect Control Hosp Epidemiol 2005;26:184e190. 2. Dominguez MA, De Lencastre H, Liñares J, Tomasz A. Spread and maintenance of a dominant methicillin-resistant Staphylococcus aureus (MRSA) clone during an outbreak of MRSA disease in a Spanish hospital. J Clin Microbiol 1994;32:2081e2087. 3. Manzur A, Gavalda L, Ruiz de Gopegui E, et al. Prevalence of methicillinresistant Staphylococcus aureus and factors associated with colonization among residents in community long-term-care facilities in Spain. Clin Microbiol Infect 2008;14:867e872. 4. Olona-Cabases M, Tico-Falguera N, Ramírez-Garceran L, Del Valle-Ortiz O, Castello-Verdu T, García-Fernandez L. Methicillin-resistant Staphylococcus aureus: a four-year experience in a spinal cord injury unit in Spain. Spinal Cord 1996;34:315e319. 5. Mody L, Maheshwari S, Galecki A, et al. Indwelling device use and antibiotic resistance in nursing homes: identifying a high-risk group. J Am Geriatr Soc 2007;55:1921e1926. 6. Bradley SF, Terpenning MS, Ramsey MA, et al. Methicillin-resistant Staphylococcus aureus: colonization and infection in a long-term-care facility. Ann Intern Med 1991;115:417e422. 7. Barr B, Wilcox MH, Brady A, Parnell P, Darby B, Tompkins D. Prevalence of methicillin-resistant Staphylococcus aureus colonization among older residents of care homes in the United Kingdom. Infect Control Hosp Epidemiol 2007;28:853e859. 8. Mody L, Kauffman CA, Donabedian S, Zervos M, Bradley SF. Epidemiology of Staphylococcus aureus colonization in nursing home residents. Clin Infect Dis 2008;46:1368e1373.
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9. Muder RR, Brennen C, Wagener MM, et al. Methicillin-resistant staphylococcal colonization and infection in a long-term care facility. Ann Intern Med 1991;114:107e112. 10. Rodríguez-Baño J, Bischofberger C, Alvarez-Lerma F, et al. Surveillance and control of methicillin-resistant Staphylococcus aureus in Spanish hospitals. A GEIH-SEIMC and SEMPSPH consensus document. Enferm Infecc Microbiol Clin 2008;26:285e298. 11. Bonten M, Slaughter S, Ambergen AW, et al. The role of “colonization pressure” in the spread of vancomycin-resistant enterococci. An important infection control variable. Arch Intern Med 1998;158:1127e1132. 12. Merrer J, Santoli F, Appéré-De Vecchi C, Tran B, De Jonghe B, Outin H. “Colonization pressure” and risk of acquisition of methicillin-resistant Staphylococcus aureus in a medical intensive care unit. Infect Control Hosp Epidemiol 2000;21:718e723. 13. Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing; 16th informational supplement. CLSI document M100-S16. Wayne, PA: Clinical and Laboratory Standards Institute; 2006. 14. Chung M, De Lencastre H, Matthews P, et al. Molecular typing of methicillinresistant Staphylococcus aureus (MRSA) by pulsed field gel electrophoresis: comparison of results obtained in a multilaboratory effort using identical protocols and MRSA strains. Microb Drug Resist 2000;6:189e197. 15. Van Belkum A, Tassios PT, Dijkshoorn L, et al. Guidelines for the validation and application of typing methods for use in bacterial epidemiology. Clin Microbiol Infect 2007;13:1e46. 16. Milheiriço C, Oliveira DC, De Lencastre H. Multiplex PCR strategy for subtyping the staphylococcal cassette chromosome mec type IV in methicillin-resistant Staphylococcus aureus: ‘SCCmec IV multiplex’. J Antimicrob Chemother 2007;60:42e48. 17. Lina GM, Piemont Y, Godail-Gamot F, et al. Involvement of PantoneValentine leukocidin-producing Staphylococcus aureus in primary skin infections and pneumonia. Clin Infect Dis 1999;29:1128e1132. 18. Enright MC, Day NP, Davies CE, Peacock SJ, Spratt BG. Multilocus sequence typing for characterization of methicillin-resistant and methicillin-susceptible clones of Staphylococcus aureus. J Clin Microbiol 2000;38:1008e1015. 19. O’Sullivan NP, Keane CT. The prevalence of methicillin-resistant Staphylococcus aureus among the residents of six nursing homes for the elderly. J Hosp Infect 2000;45:322e329. 20. von Baum H, Schmidt C, Svoboda D, Bock-Hensley O, Wendt C. Risk factors for methicillin-resistant Staphylococcus aureus carriage in residents of German nursing homes. Infect Control Hosp Epidemiol 2002;23:511e515. 21. Bradley SF. Methicillin-resistant Staphylococcus aureus: long-term care concerns. Am J Med 1999;106:2Se10S. 22. Mulhausen PL, Harrell LJ, Weinberger M, Kochersberger GG, Feussner JR. Contrasting methicillin-resistant Staphylococcus aureus colonization in veteran’s affairs and community nursing homes. Am J Med 1996;100:24e31. 23. Stone ND, Lewis DR, Lowery HK, et al. Importance of bacterial burden among methicillin-resistant Staphylococcus aureus carriers in a long-term care facility. Infect Control Hosp Epidemiol 2008;29:143e148. 24. Manzur A, Vidal M, Pujol M, et al. Predictive factors of meticillin resistance among patients with Staphylococcus aureus bloodstream infection at hospital admission. J Hosp Infect 2007;66:135e141. 25. Gavalda L, Masuet C, Beltran J, et al. Comparative cost of selective screening to prevent transmission of methicillin resistant Staphylococcus aureus (MRSA), compared with the attributable costs of MRSA infection. Infect Control Hosp Epidemiol 2006;27:1264e1266. 26. Cooper BS, Medley GF, Stone SP, et al. Methicillin-resistant Staphylococcus aureus in hospitals and the community: stealth dynamics and control catastrophes. Proc Natl Acad Sci USA 2004;101:10223e10228. 27. Vindel A, Cuevas O, Cercenado E, et al. Methicillin-resistant Staphylococcus aureus in Spain: molecular epidemiology and utility of different typing methods. J Clin Microbiol 2009;47:1620e1627. 28. Rodríguez-Baño J, Domínguez M, Millán A, et al. Clinical and molecular epidemiology of community-acquired, healthcare-associated and nosocomial methicillin-resistant Staphylococcus aureus in Spain. Clin Microbiol Infect 2009;15:1111e1118. 29. Ruiz de Gopegui E, Oliver A, Ramírez A, Gutiérrez O, Andreu C, Pérez JL. Epidemiological relatedness of methicillin-resistant Staphylococcus aureus from a tertiary hospital and a geriatric institution in Spain. Clin Microbiol Infect 2004;10:339e342. 30. Mahoney F, Barthel DW. Functional evaluation: the Barthel index. Md State Med J 1965;14:61e65.