Incidence, Risk Factors, and Outcomes of Panton-Valentine

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Incidence, Risk Factors, and Outcomes of Panton-Valentine

     JOURNAL OF CLINICAL MICROBIOLOGY, Oct. 2010, p. 34703474 Vol. 48, No. 10 0095-1137/10/$12.00 doi:10.1128/JCM.00911-10 Copyright ? 2010, American Society for Microbiology. All Rights Reserved.

Incidence, Risk Factors, and Outcomes of Panton-Valentine

    Leukocidin-Positive Methicillin-Susceptible

    Staphylococcus aureus Infections in

    Auckland, New Zealand

    1213451S. Muttaiyah,* G. Coombs,S. Pandey,P. Reed,S. Ritchie,D. Lennon,and S. Roberts

    1Department of Microbiology, Auckland City Hospital, Auckland, New Zealand; Department of Microbiology and Infectious Diseases,

    PathWest Laboratory Medicine-WA, Royal Perth Hospital, and School of Pathology and Laboratory Medicine, University of 2Western Australia, Perth, Australia; Childrens Research Centre, Starship Childrens Health, Auckland City Hospital, 3Auckland, New Zealand; Department of Molecular Medicine and Pathology, University of Auckland, Auckland, 45New Zealand; and Community Paediatrics, University of Auckland, Auckland, New Zealand

    Received 10 May 2010/Returned for modication 1 June 2010/Accepted 28 July 2010

Panton-Valentine leukocidin (PVL) has been linked to invasive community-acquired methicillin-resistant Staph-

    ylococcus aureusinfections. However, the association between disease and PVL-positive methicillin-susceptible Staphylococcus aureus (MSSA) has not been widely reported. We aimed to examine the epidemiology of PVL in

    clinical MSSA isolates from patients presenting to Auckland City Hospital. Four hundred eleven MSSA clinical

    isolates and 93 nasal carriage isolates were collected and tested for the presence of the lukSF-PV genes using PCR. The results were examined in light of host and disease factors. Multilocus sequence typing (MLST) was performed

    on a random subset of isolates to ensure that there was no single PVL-positive MSSA clone responsible for disease

    in Auckland. The prevalence of the lukSF-PV genes in MSSA isolates associated with disease (124/335; 37%) was not signicantly different from the prevalence of the lukSF-PV genes in MSSA nasal carriage isolates (29/93; 31% [P

    0.33]). PVL-positive MSSA isolates in Auckland are genetically diverse and come from a number of different clonal

    complexes. PVL-positive infections peaked at between 10 and 20 years of age, with a subsequent decline. Pacic ethnicity, age, diagnosis of skin and soft tissue infection (SSTI), community-onset infection, and the need for

    surgical intervention were found by multivariate analysis to be independently associated with PVL-positive MSSA

    infection. More than one-third of MSSA infections in our patient population are caused by PVL-positive strains. Those patients with PVL-positive MSSA infection were more likely to be of Pacic ethnicity, be younger in age, have

    community-onset infection, have SSTI, and need surgical intervention.

     Staphylococcus aureus is a nasal commensal that can be de- type IV and also SCCmec type V (4, 25). The epidemiology of PVL-positive methicillin-susceptible S. aureus (MSSA) has not tected in up to 20 to 30% of the general population, one-third of whom are persistently colonized (28). S. aureus produces a been reported as extensively, and the lukSF-PV genes are not exclusively linked to the presence of the SCCmec element. In wide variety of virulence factors that contribute to its ability to the 1950s MSSA ST80 strains, which were associated with colonize, invade, and evade the immune system, which includes outbreaks of SSTI, harbored the lukSF-PV genes (22). There Panton-Valentine leukocidin (PVL), a bicomponent, pore- forming toxin encoded by two contiguous genes, lukF-PV and have also been recent reports of PVL-positive MSSA causing lukS-PV. PVL can cause either neutrophil lysis or apoptosis clusters of SSTI and necrotizing pneumonia (5, 15). and contributes to tissue necrosis (25). PVL has been linked to The vast majority of S. aureus strains in New Zealand are skin and soft tissue infections (SSTIs), necrotizing pneumonia, methicillin susceptible (MSSA); the prevalence of methicillin- and bone and joint infections in humans (3, 11, 17). Rabbit and resistant S. aureus (MRSA) remains low, at about 5% (12). human leukocytes are highly sensitive to PVL-mediated leukocy- New Zealand has a high incidence of S. aureus disease; the tosis (18), and animal studies have shown that PVL causes more- incidence of S. aureus bacteremia in the late 1990s was 41 cases severe disease in dermonecrosis (7, 27), osteomyelitis (6), and per 100,000 adults per year (12). We aimed to examine the

    necrotizing pneumonia models (B. A. Diep, L. Chan, and P. prevalence of the lukSF-PV genes in MSSA isolates responsi-

    Tattevin, presented at the 49th Interscience Conference on An- ble for disease and asymptomatic nasal carriage, to determine

     timicrobial Agents and Chemotherapy, San Francisco, CA, 2009). risk factors for infection with PVL-positive MSSA, and to

     The presence of PVL has been extensively described for examine the association between PVL and severity of disease.

     methicillin-resistant S. aureus (MRSA), specically in associa- tion with staphylococcal cassette chromosome mec (SCCmec) MATERIALS AND METHODS Clinical MSSA isolates. All MSSA isolates isolated from diagnostic specimens * Corresponding author. Mailing address: Department of Microbi- submitted to the Microbiology Laboratory of Auckland City Hospital from Feb- ology, Auckland City Hospital, Private Bag 92026, Grafton, Auckland, ruary to April 2008 were collected. These specimens were generally obtained at New Zealand. Phone: 3074949, ext. 25985. Fax: 3074940. E-mail: the request of the attending physician, midwife, or district nurse for clinical reasons. Duplicate isolates from the same patient were excluded. The isolates Published ahead of print on 4 August 2010. were stocked onto nutrient agar (Difco Laboratories, Detroit, MI).



Nasal carriage isolates. MSSA isolates from nasal carriers were obtained from and Fishers exact tests, and the results were expressed in terms of odds ratios healthy population volunteers in the Auckland community during a separate (ORs) and associated 95% condence intervals (CIs). For continuous variables, study performed to examine the demographic features of nasal carriers in Auck- the differences between patient groups were investigated by the parametric t test land. These volunteers were recruited in public places spread across the Auck- or the nonparametric Wilcoxon rank sum test as appropriate. Multivariate anal- land region between February and November 2008; people with hospital contact yses were undertaken by using nominal logistic regression and effect likelihood in the previous 3 months were excluded. ratio tests, with signicance set at a P value of 0.05. Detection of the lukSF-PV genes. All isolates were cultured onto tryptic soy Ethical approval. The regional ethics committee of the New Zealand Ministry agar with 5% sheep blood (Difco Laboratories, Detroit, MI) and incubated of Health provided ethical approval for the study. aerobically overnight at 35?C. Nucleic acid was extracted from MSSA isolates as previously described (20). The PVL and nuc probes were synthesized with the nonuorescent quencher BHQ1 (Biosearch Technologies, CA). Cal Fluor Gold RESULTS 540 and FAM (6-carboxyuorescein) were utilized as reporter dyes. Primers were synthesized based on oligonucleotide sequences described previously (20). MSSA isolates were obtained from 411 patients for whom Data for the duplex PCR (for the simultaneous amplication of the lukSF-PV clinical details were available. The majority of the patients, genes and the nuc gene) were collected with an ABI 7500 real-time PCR system 335/411 (81.5%), had a clearly identiable focus of infection; (Applied Biosystems, CA). Reactions were performed with 1 universal master 76/411 (18.5%) patients were not considered to have S. aureus mix (Applied Biosystems, CA) under the following thermal cycling conditions: 15 min at 95?C, followed by 40 cycles of 95?C for 15 s and 55?C for 1 min. Ampli- infection and were excluded from analysis. We also obtained cation was conrmed by the generation of a sigmoid amplication plot. A 93 MSSA isolates from nasal carriers for comparison. The negative control was provided by use of negative blanks and PCR blanks. prevalence of the lukSF-PV genes among the clinical MSSA MLST. In order to ensure that there was no single dominant PVL-positive isolates, 124/335 (37%), was not signicantly different from the MSSA clone that caused disease in Auckland, all of the lukSF-PV MSSA carriage prevalence of the lukSF-PV genes among the nasal carriage isolates were genotyped by using multilocus sequence typing (MLST) (9). A similar number of randomly selected lukSF-PV-positive clinical MSSA isolates isolates, 29/93 (31%) (P 0.33). MLST conrmed that a were also genotyped. To assign a clonal complex (CC), sequences were com- diverse range of MSSA genotypes was PVL positive in Auck- pared with the sequences in the S. aureus MLST database (http://saureus.mlst land: the 29 lukSF-PV-positive nasal carriage isolates belonged .net/) (10). to eight clonal complexes (CC5, 9/29 isolates [31%]; CC30, Clinical data. Electronic clinical records were reviewed to ascertain the fol- lowing demographic data: patient age, gender, ethnicity, and place of domicile. 7/29 [24%]; CC1, 6/29 [21%]; CC22, 2/29 [7%]; CC78, 2/29 The NZDep2006 (New Zealand Deprivation 2006) score was calculated for each [7%]; CC15, 1/29 [3%]; CC97, 1/29 [3%]; CC121, 1/29 [3%]), patient; the NZDep2006 score is derived from population data for each neigh- and the 24 randomly selected lukSF-PV-positive clinical iso- borhood (23). This score ranges from 1 to 10, where 1 represents the least lates belonged to six clonal complexes (CC1, 12/24 isolates deprived areas of Auckland and 10 represents the most deprived areas of Auck- [50%]; CC121, 6/24 [25%]; CC30, 3/24 [13%]; CC5, 1/24 [4%]; land. The source of infection was also recorded. In order to evaluate potential risk factors associated with S. aureus disease, the following data were ascertained: CC8, 1/24 [4%]; CC78, 1/24 [4%]). Several predominant clonal community versus hospital onset, Charlson weighted index of comorbidity (16), complexes contained the majority of the lukSF-PV-positive the presence of an implanted medical device, recent trauma with breach of skin MSSA isolates: 88% of the clinical isolates belonged to three within 30 days, the presence of a noninfective chronic skin condition, or a predominant clonal complexes (CC1, CC30, and CC121), and history of alcoholism or intravenous drug use. Disease severity was measured by means of the following data: requirement for surgical intervention, num- 76% of the nasal carriage isolates belonged to three predom- ber of surgical procedures, bloodstream invasion, duration of admission, inant clonal complexes (CC1, CC5, and CC30). 30-day readmission rate, and 30-day mortality rate. The prevalences of PVL-positive MSSA for patients within Denitions used. Skin and soft tissue infection was dened as the presence of each infection group were 112/235 (48%) with skin and soft a cutaneous abscess or other signs of inammation and a culture positive for tissue infection, 4/13 (31%) with bone and joint disease, 2/11 MSSA. Bone infection was dened as radiologic or surgical evidence of osteomyelitis (18%) with pneumonia, 2/36 (2%) with surgical-site infection, and MSSA-positive blood or aspirate culture. 1/13 (1%) with catheter-related infection, and 0/6 (0%) with Joint infection was dened as pain with, or limitation of, joint movement and primary bacteremia and endocarditis. Univariate analysis was MSSA-positive aspirate or blood cultures. performed to compare features of the PVL-positive MSSA Pneumonia was dened as the presence of fever with radiographic changes consistent with pneumonia and MSSA-positive sputum, tracheal aspirate, bron- infections (n 124) to the features of the PVL-negative MSSA chial washing, or blood cultures. infections (n 211) (Table 1). Histories of alcoholism and Surgical-site infection and catheter-related infection were dened in accor- intravenous drug use were excluded from Table 1, as they were dance with the criteria of the Australian Council of Healthcare Standards (1). not routinely documented in clinical records. Factors associ- Primary bacteremia was dened as the presence of a blood culture positive for ated with PVL-positive infections in the univariate analysis MSSA with no identiable focus of infection. The diagnosis of endocarditis was based on modied Dukes criteria (8). were investigated further by employing multivariate analysis. A Community-onset, not health care-associated, infections were dened as those diagnosis of SSTI, Pacic ethnicity, younger age, community- episodes with an MSSA-positive culture within 48 h of admission without a <