Higher incidence of perineal community acquired MRSA infections among toddlers
© McCullough et al; licensee BioMed Central Ltd. 2011
Received: 31 January 2011
Accepted: 27 October 2011
Published: 27 October 2011
A six-fold increase in pediatric MRSA infections, prompted us to examine the clinical profile of children with MRSA infections seen at Mercy Children's Hospital, Toledo, Ohio and to characterize the responsible strains.
Records were reviewed of pediatric patients who cultured positive for MRSA from June 1 to December 31, 2007. Strain typing by pulsed field gel electrophoresis (PFT) and DiversiLab, SCCmec typing, and PCR-based lukSF-PV gene (encodes Panton-Valentine leukocidin), arginine catabolic mobile element (ACME) and cap5 gene detection was performed.
Chart review of 63 patients with MRSA infections revealed that 58(92%) were community acquired MRSA (CAMRSA). All CAMRSA were skin and soft tissue infections (SSTI). Twenty five (43%) patients were aged < 3 yrs, 19(33%) aged 4-12 and 14(24%) aged 13-18. Nineteen (76%) of those aged < 3 yrs had higher incidence of perineal infections compared to only 2(11%) of the 4-12 yrs and none of the 13-18 yrs of age. Infections in the extremities were more common in the older youth compared to the youngest children. Overall, there was a significant association between site of the infection and age group (Fisher's Exact p-value < 0.001). All CAMRSA were USA300 PFT, clindamycin susceptible, SCCmec type IVa and lukSF-PV gene positive. Nearly all contained ACME and about 80% were cap5 positive. Of the 58 USA300 strains by PFT, 55(95%) were also identified as USA300 via the automated repetitive sequence-based PCR method from DiversiLab.
CAMRSA SSTI of the perineum was significantly more common among toddlers and that of the extremities in older children. The infecting strains were all USA300 PFT. Further studies are needed to identify the unique virulence and colonization characteristics of USA300 strains in these infections.
Staphylococcus aureus (S. aureus) is a common human commensal organism and a clinically important invasive pathogen. Methicillin resistant S. aureus (MRSA) remains one of the most prevalent pathogens isolated from hospital patients. However, MRSA infections are increasingly arising outside of healthcare settings among individuals in the community with no established risk factors. Furthermore, the incidence of invasive community acquired MRSA (CAMRSA) disease in previously-healthy children has been increasing [1–3].
MRSA isolate collection and patient medical record review
This is a retrospective, descriptive, single-cohort study conducted at Mercy Children's Hospital, Toledo, Ohio and was approved by the institutional review board. Pediatric patients (< 18 yrs of age), who were culture positive for MRSA between June 1 to December 31, 2007, were identified by the clinical microbiology laboratory (Mercy Integrated Laboratories, Toledo, Ohio). This facility also provided us with a sample of the clinical isolate that had been frozen at -80°C in a solution of 50% brain-heart infusion and 50% glycerol for later analysis.
Medical charts of these patients were reviewed for social and demographic information including age, gender, race, socio-economic status based on insurance, pre-existing history of skin and soft tissue infection (SSTI); respiratory, cardiovascular, gastrointestinal, and nervous system diseases, type of care (outpatient vs. emergency room vs. inpatient care) and site of infection. Antimicrobial susceptibilities were determined based on Clinical and laboratory Standard Institute (CLSI) guidelines at the clinical laboratory using the Vitek system for oxacillin, erythromycin, clindamycin, vancomycin, ciprofloxacin, tetracycline, trimethoprim/sulfamethoxasole, rifampin and linezolid. All erythromycin-resistant strains were tested for inducible clindamycin resistance using the D-test  before final susceptibilities were reported.
Pulsed Field typing (PFT): Strain typing was done by PFT as described by Chang et al , using SmaI as the restriction enzyme, at the Children's Memorial Hospital, Chicago, Illinois. The relatedness of isolates was based on visual comparison of band patterns by the use of criteria described by Tenover et al .
DiversiLab typing: Strain typing was also performed using the DiversiLab System at Mercy Integrated Laboratories. This is an automated method using repetitive sequence-based PCR that targets multiple noncoding repetitive sequences in the genomic DNA. Band patterns were subsequently analyzed using the web-based DiversiLab software .
DNA Extraction and PCR
Clinical isolates were grown on 5% blood agar plates overnight at 37°C. DNA was extracted from the isolates using the Wizard® Genomic DNA Purification Kit. PCR amplification was then performed on all patient isolates for the presence of mecA, SCCmec typing, arcA, ACME, lukSF-PV and cap5 genes (Additional file 1) [14–17].
Fisher's Exact test was used to explore for associations of site of infection, pre-existing SSTI and respiratory disease with age group and PFGE type. A Fisher's Exact p-value of < 0.05 was considered statistically significant. Data were analyzed using SAS (SAS Cary, NC, version 9).
From June 1, 2007 to December 31, 2007, 63 pediatric patients with MRSA infections were seen at the emergency room, outpatient clinics and the inpatient ward at Mercy Children's Hospital. Of these, only 5 patients did not meet the CDC clinical criteria for CAMRSA . Thus, 58 (92%) of all pediatric MRSA infections were community-acquired. All of these CAMRSA infections were SSTIs. During this time period there were no other culture positive invasive pediatric MRSA infections like bacteremia, sepsis, endocarditis, pneumonia, or osteomyelitis.
Age group (years)
Type of Visit
Preexisting Conditions (patients may have more than 1)
Site of Infection
Associations with Age Group
Age 0-3 N = 25
Age 4-12 N = 19
Age 13-18 N = 14
Fisher's Exact Two-tailed P-value
Site of Infection
Respiratory disease pre-existing condition
SSTI pre-existing condition
PFT Sub-type 1 (compared to types 2-7 combined)
Molecular epidemiology of USA300 PFT strains (n = 58)
USA300 PFT subtypes n(%)
Diversilab type* n(%)
1 - 42(72%)
2 - 6(10%)
3 - 4(7%)
4 - 2(3%)
5 - 2(3%)
6 - 1(2%)
7 - 1(2%)
Associations with PFT Sub-type
PFT Sub-type 1 N = 42
PFT Sub-types 2-7 N = 16
Fisher's Exact Two-tailed P-value
Site of Infection
Respiratory disease pre-existing condition
SSTI pre-existing condition
All 58 CAMRSA strains were mecA and lukSF-PV positive, with SCCmec type IVa; 56 (97%) contained ACME and the cap5 gene was present in 46 (79%) (Table 3). We observed that all the cap5 negative strains belonged to a single USA300 PFT pattern (Table 3 and Figure 2).
Figure 1 clearly demonstrates a six fold increase in the incidence of pediatric MRSA infections from 2002 to 2007 (p < 0.0001). A clear majority of our pediatric MRSA infections were CAMRSA, which is consistent with the trend reported for the United States  and Europe [19, 20].
A significant association of age with site of infection has for the first time been demonstrated in our study. Perineal MRSA colonization in children has been recorded in the daycare setting . Koski et al indicate that pediatric perineal infections with MRSA are increasing . We found that such infections were more common to the 0-3 y-old cohort. This age preference could be due to increased rates of perineal CAMRSA colonization in this age group, but may also reflect use of diapers and possible dermabrasion caused by vigorous wiping of the area during diaper changes or both. Transfer of vancomycin resistance gene vanA from vancomycin resistant Enterococcus to MRSA has been shown to occur in vitro and in vivo . Perineal CAMRSA infection in children could contribute to the emergence of vancomycin resistant S.aureus strains when co-colonized with vancomycin resistant Enterococcus .
Strain typing of our patients' MRSA isolates supports the observation that USA300 PFT is the most common causative strain and is clonal . The molecular characteristics of our isolates were similar to the USA300 strains from other published reports in that they were SCCmec type IVa and lukSF-PV positive [27, 28]. 56/58 of the strains were also positive for ACME, a novel mobile genetic region predominantly reported in USA300 MRSA strains that potentially enhance colonization and virulence . We observed that all the 12 strains that were cap5 gene negative belonged to the predominant USA300 PFT subtype (Table 3 and Figure 2). cap5 gene encodes for a capsule that enhances the virulence of Staphylococcus aureus. It has been used as vaccine target. The clinical significance of cap5 negative strains in our study is unclear at this time.
Typing using the DiversiLab system is rapid and user friendly compared to PFT. However, differentiating PFT USA300 from USA500 is tricky using DiversiLab . Of interest, 3/3 isolates identified by PFT as USA300 were mis-identified by DiversiLab as USA500. These isolates were also positive for ACME which is a mobile genetic element that is thought to be acquired by USA300 as it evolved from its progenitor USA500 .
The incidence of CAMRSA infection is increasing in the pediatric population of Northwest Ohio. SSTIs are the most common type of infection and among children < 3 years of age with perineal SSTIs being the dominant form caused by strain USA300 PFT that carries the SCCmec type IVa, lukSF-PV gene and ACME. The automated rapid strain typing method, the DiversiLab system, is not as discriminative as PFT.
Further investigations are needed to assess the extent of USA300 perineal colonization in toddlers and to identify unique virulence characteristics to develop strategies for prevention and treatment of these infections.
Arginine catabolic mobile element
Community-acquired methicillin resistant Staphylococcus aureus
Methicillin resistant Staphylococcus aureus
Polymerase chain reaction
Pulsed field type (or) pulsed field gel electrophoresis
- S.aureus :
Skin and soft tissue infection.
I would like to thank Jan Tucker of Mercy Integrated Laboratories and Dr. John Schaeufele, President and CEO of Mercy Children's Hospital for their support. Nancy Buderer helped with the statistical analysis.
- Chambers HF: The changing epidemiology of S. aureus?. Emerg Infect Dis. 2001, 7: 178-82. 10.3201/eid0702.010204.View ArticlePubMedPubMed CentralGoogle Scholar
- Zetola N, Francis JS, Nuermberger EL, Bishai WR: CAMRSA: An emerging threat. Lancet Infect Dis. 2005, 5: 275-86. 10.1016/S1473-3099(05)70112-2.View ArticlePubMedGoogle Scholar
- Shapiro A, Raman S, Johnson M, Piehl M: CAMRSA infections in North Carolina children: prevalence, antibiotic sensitivities, and risk factors. N C Med J. 2009, 70: 102-7.PubMedGoogle Scholar
- Naimi TS, LeDell KH, Como-Sabetti K, Borchardt SM, Boxrud DJ, Etienne J, et al: Comparison of community- and health care-associated MRSA infection. JAMA. 2003, 290: 2976-84. 10.1001/jama.290.22.2976.View ArticlePubMedGoogle Scholar
- Goering RV, McDougal LK, Fosheim GE, Bonnstetter KK, Wolter DJ, Tenover FC: Epidemiologic distribution of the ACME among selected methicillin-resistant and methicillin-susceptible S.aureus isolates. J Clin Microbiol. 2007, 45: 1981-4. 10.1128/JCM.00273-07.View ArticlePubMedPubMed CentralGoogle Scholar
- King MD, Humphrey BJ, Wang YF, Kourbatova EV, Ray SM, Blumberg HM: Emergence of CAMRSA USA 300 clone as the predominant cause of SSTI. Ann Intern Med. 2006, 144: 309-17.View ArticlePubMedGoogle Scholar
- Chua K, Laurent F, Coombs G, Grayson ML, Howden BP: Antimicrobial resistance: Not community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA)! A clinician's guide to community MRSA - its evolving antimicrobial resistance and implications for therapy. Clin Infect Dis [Research Support, Non-U.S. Gov't Review]. 2011, 52 (1): 99-114.Google Scholar
- Reygaert W: Methicillin-resistant Staphylococcus aureus (MRSA): prevalence and epidemiology issues. Clin Lab Sci. 2009, 22 (2): 111-4.PubMedGoogle Scholar
- Moore CL, Hingwe A, Donabedian SM, Perri MB, Davis SL, Haque NZ, et al: Comparative evaluation of epidemiology and outcomes of methicillin-resistant Staphylococcus aureus (MRSA) USA300 infections causing community- and healthcare-associated infections. Int J Antimicrob Agents. [Comparative Study]. 2009, 34 (2): 148-55. 10.1016/j.ijantimicag.2009.03.004.View ArticleGoogle Scholar
- Shutt CK, Pounder JI, Page SR, Schaecher BJ, Woods GL: Clinical evaluation of the DiversiLab microbial typing system using repetitive-sequence-based PCR for characterization of S.aureus strains. J Clin Microbiol. 2005, 43: 1187-92. 10.1128/JCM.43.3.1187-1192.2005.View ArticlePubMedPubMed CentralGoogle Scholar
- Lewis JS, Jorgensen JH: Inducible clindamycin resistance in Staphylococci: should clinicians and microbiologists be concerned?. Clin Infect Dis. 200 (40): 280-5.
- Chang N, Chui L: A standardized protocol for the rapid preparation of bacterial DNA for pulsed field electrophoresis. Diagn Microbiol Infect Dis. 1998, 31: 275-9. 10.1016/S0732-8893(98)00007-8.View ArticlePubMedGoogle Scholar
- Tenover FC, Arbeit RD, Goering RV, Mickelsen PA, Murray BE, Persing DH, et al: Interpreting chromosomal DNA restriction patterns produced by pulsed-field gel electrophoresis: criteria for bacterial strain typing. J Clin Microbiol. 1995, 33: 2233-9.PubMedPubMed CentralGoogle Scholar
- Oliveira DC, de Lencastre H: Multiplex PCR strategy for rapid identification of structural types and variants of the mec element in MRSA. Antimicrob Agents Chemother. 2002, 46: 2155-61. 10.1128/AAC.46.7.2155-2161.2002.View ArticlePubMedPubMed CentralGoogle Scholar
- Zhang K, McClure JA, Elsayed S, Louie T, Conly JM: Novel multiplex PCR assay for characterization and concomitant subtyping of SCC mec types I to V in MRSA. J Clin Microbiol. 2005, 43: 5026-33. 10.1128/JCM.43.10.5026-5033.2005.View ArticlePubMedPubMed CentralGoogle Scholar
- Lina G, Piemont Y, Godail-Gamot F, Bes M, Peter MO, Gauduchon V, et al: Involvement of PVL-producing S.aureus in primary skin infections and pneumonia. Clin Infect Dis. 1999, 29: 1128-32. 10.1086/313461.View ArticlePubMedGoogle Scholar
- Moore PC, Lindsay JA: Genetic variation among hospital isolates of methicillin-sensitive S.aureus: evidence for horizontal transfer of virulence genes. J Clin Microbiol. 2001, 39: 2760-7. 10.1128/JCM.39.8.2760-2767.2001.View ArticlePubMedPubMed CentralGoogle Scholar
- Como-Sabetti K, Harriman KH, Buck JM, Glennen A, Boxrud DJ, Lynfield R: CAMRSA: trends in case and isolate characteristics from six years of prospective surveillance. Public Health Rep. 2009, 124: 427-35.PubMedPubMed CentralGoogle Scholar
- Faria NA, Oliveira DC, Westh H, Monnet DL, Larsen AR, Skov R, et al: Epidemiology of emerging MRSA in Denmark: A nationwide study in a country with low prevalence of MRSA infection. J Clin Microbiol. 2005, 43: 1836-42. 10.1128/JCM.43.4.1836-1842.2005.View ArticlePubMedPubMed CentralGoogle Scholar
- Vourli S, Perimeni D, Makri A, Polemis M, Voyiatzi A, Vatopoulos A: CAMRSA infections in a paediatric population in Greece. Euro Surveill. 2005, 10: 78-9.PubMedGoogle Scholar
- Shahin R, Johnson IL, Jamieson F, McGeer A, Tolkin J, Ford-Jones EL: MRSA carriage in a child care center following a case of disease. Toronto Child Care Center Study Group. Arch Pediatr Adolesc Med. 1999, 153: 864-8.View ArticlePubMedGoogle Scholar
- Koski ME, DeMarco RT, Brock JW, Pope JC, Adams MC, Thomas JC: Community associated methicillin resistant staphylococcal infections in a pediatric urology practice. J Urol. 2008, 179: 1098-101. 10.1016/j.juro.2007.10.086.View ArticlePubMedGoogle Scholar
- Perichon B, Courvalin P: VanA-type vancomycin-resistant S.aureus. Antimicrob Agents Chemother. 2009, 53: 4580-7. 10.1128/AAC.00346-09.View ArticlePubMedPubMed CentralGoogle Scholar
- Warren DK, Nitin A, Hill C, Fraser VJ, Kollef MH: Occurrence of co-colonization or co-infection with vancomycin-resistant enterococci and MRSA in a medical intensive care unit. Infect Control Hosp Epidemiol. 2004, 25: 99-104. 10.1086/502357.View ArticlePubMedGoogle Scholar
- Diep BA, Chambers HF, Graber CJ, Szumowski JD, Miller LG, Han LL, et al: Emergence of multidrug-resistant, CAMRSA clone USA300 in men who have sex with men. Ann Intern Med. 2008, 148: 249-57.View ArticlePubMedGoogle Scholar
- Kaplan SL, Hulten KG, Gonzalez BE, Hammerman WA, Lamberth L, Versalovic J, et al: Three-year surveillance of CAMRSA infections in children. Clin Infect Dis. 2005, 40: 1785-91. 10.1086/430312.View ArticlePubMedGoogle Scholar
- David MZ, Rudolph KM, Hennessy TW, Boyle-Vavra S, Daum RS: Molecular epidemiology of MRSA, rural southwestern Alaska. Emerg Infect Dis. 2008, 14: 1693-9. 10.3201/eid1411.080381.View ArticlePubMedPubMed CentralGoogle Scholar
- Jones RN, Nilius AM, Akinlade BK, Deshpande LM, Notario GF: Molecular characterization of S.aureus isolates from a 2005 clinical trial of uncomplicated skin and skin structure infections. Antimicrob Agents Chemother. 2007, 51: 3381-4. 10.1128/AAC.01588-06.View ArticlePubMedPubMed CentralGoogle Scholar
- Diep BA, Stone GG, Basuino L, Graber CJ, Miller A, des Etages SA, et al: The ACME and SCCmec linkage: convergence of virulence and resistance in the USA300 clone of MRSA. J Infect Dis. 2008, 197: 1523-30. 10.1086/587907.View ArticlePubMedGoogle Scholar
- Library Stats Sheet: MRSA. 2008 BioMerieux Inc. BBI-019-08. Accessed April 6, 2010, [http://www.biomerieux-usa.com/upload/BBI-019-08%20LSS%20MRSA%20v3-1.pdf]
- Li M, Diep BA, Villaruz AE, Braughton KR, Jiang X, DeLeo FR, et al: Evolution of virulence in epidemic community-associated methicillin-resistant Staphylococcus aureus. Proc Natl Acad Sci USA. 2009, 106 (14): 5883-8. 10.1073/pnas.0900743106.View ArticlePubMedPubMed CentralGoogle Scholar
- The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2431/11/96/prepub
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.