Development and validation of a multiplex real-time PCR assay for detection and quantification of Streptococcus pneumoniae in pediatric respiratory samples.

IMPORTANCE: Streptococcus pneumoniae (Spn) is the world's leading cause of lower respiratory tract infection morbidity and mortality in children. However, current clinical microbiological methods have disadvantages. Spn can be difficult to grow in laboratory conditions if a patient is pre-treated, and Spn antigen testing has unclear clinical utility in children. Syndromic panel testing is less cost-effective than targeted PCR if clinical suspicion is high for a single pathogen. Also, such testing entails a full, expensive validation for each panel target if used for multiple respiratory sources. Therefore, better diagnostic modalities are needed. Our study validates a multiplex PCR assay with three genomic targets for semi-quantitative and quantitative Spn molecular detection from lower respiratory sources for clinical testing and from upper respiratory sources for research investigation.

Pediatric Swine Model of Methicillin-Resistant Staphylococcus aureus Sepsis-Induced Coagulopathy, Disseminated Microvascular Thrombosis, and Organ Injuries.

Sepsis-induced coagulopathy leading to disseminated microvascular thrombosis is associated with high mortality and has no existing therapy. Despite the high prevalence of Gram-positive bacterial sepsis, especially methicillin-resistant Staphylococcus aureus (MRSA), there is a paucity of published Gram-positive pediatric sepsis models. Large animal models replicating sepsis-induced coagulopathy are needed to test new therapeutics before human clinical trials. HYPOTHESIS: Our objective is to develop a pediatric sepsis-induced coagulopathy swine model that last 70 hours. METHODS AND MODELS: Ten 3 weeks old piglets, implanted with telemetry devices for continuous hemodynamic monitoring, were IV injected with MRSA (n = 6) (USA300, Texas Children's Hospital 1516 strain) at 1 × 109 colony forming units/kg or saline (n = 4). Fluid resuscitation was given for heart rate greater than 50% or mean arterial blood pressure less than 30% from baseline. Acetaminophen and dextrose were provided as indicated. Point-of-care complete blood count, prothrombin time (PT), activated thromboplastin time, d-dimer, fibrinogen, and specialized coagulation assays were performed at pre- and post-injection, at 0, 24, 48, 60, and 70 hours. Piglets were euthanized and necropsies performed. RESULTS: Compared with the saline treated piglets (control), the septic piglets within 24 hours had significantly lower neurologic and respiratory scores. Over time, PT, d-dimer, and fibrinogen increased, while platelet counts and activities of factors V, VII, protein C, antithrombin, and a disintegrin and metalloproteinase with thrombospondin-1 motifs (13th member of the family) (ADAMTS-13) decreased significantly in septic piglets compared with control. Histopathologic examination showed minor focal organ injuries including microvascular thrombi and necrosis in the kidney and liver of septic piglets. INTERPRETATIONS AND CONCLUSIONS: We established a 70-hour swine model of MRSA sepsis-induced coagulopathy with signs of consumptive coagulopathy, disseminated microvascular thrombosis, and early organ injuries with histological minor focal organ injuries. This model is clinically relevant to pediatric sepsis and can be used to study dysregulated host immune response and coagulopathy to infection, identify potential early biomarkers, and to test new therapeutics.

A point-of-need platform for rapid measurement of a host-protein score that differentiates bacterial from viral infection: Analytical evaluation.

The objective was to evaluate the analytical performance of a new point-of-need platform for rapid and accurate measurement of a host-protein score that differentiates between bacterial and viral infection. The system comprises a dedicated test cartridge (MeMed BV®) and an analyzer (MeMed Key®). In each run, three host proteins (TRAIL, IP-10 and CRP) are measured quantitatively and a combinational score (0-100) computed that indicates the likelihood of Bacterial versus Viral infection (BV score). Serum samples collected from patients with acute infection representing viral (0 ≤ score < 35), equivocal (35 ≤ score ≤ 65), or bacterial (65 < score ≤ 100) scores based on pre-defined score cutoffs were employed for the analytical evaluation studies as well as samples from healthy individuals. To assess reproducibility, triplicate runs were conducted at 3 different sites, on 2 analyzers per site over 5 non-consecutive days. Lower limit of quantitation (LLoQ) and analytical measurement range were established utilizing recombinant proteins. Sample stability was evaluated using patient samples representative of BV score range (0-100). MeMed Key® and MeMed BV® passed the acceptance criteria for each study. In the reproducibility study, TRAIL, IP-10 and CRP measurements ranged with coefficient of variation from 9.7 to 12.7%, 4.6 to 6.2% and 5.0 to 11.6%, respectively. LLoQ concentrations were established as 15 pg/mL, 100 pg/mL and 1 mg/L for TRAIL, IP-10 and CRP, respectively. In summary, the analytical performance reported here, along with diagnostic accuracy established in the Apollo clinical validation study (NCT04690569), supports that MeMed BV® run on MeMed Key® can serve as a tool to assist clinicians in differentiating between bacterial and viral infection.

Adhesin genes and biofilm formation among pediatric Staphylococcus aureus isolates from implant-associated infections.

BACKGROUND: Microbial surface component recognizing adhesive matrix molecules (MSCRAMMs) facilitate Staphylococcus aureus adherence to host tissue. We hypothesized that S. aureus isolates from implant-associated infections (IAIs) would differ in MSCRAMM profile and biofilm formation in vitro compared to skin and soft tissue infection (SSTI) isolates. METHODS: Pediatric patients and their isolates were identified retrospectively. IAI and SSTI isolates were matched (1:4). Pulsed field gel electrophoresis was performed to group isolates as USA300 vs. non-USA300. Whole genome sequencing was performed and raw sequence data were interrogated for presence of MSCRAMMs (clfA, clfB, cna, ebh, efb, fnbpA, fnbpB, isdA, isdB, sdrC, sdrD, sdrE), biofilm-associated (icaA,D,B,C), and Panton-Valentine leukocidin (lukSF-PV) genes, accessory gene regulator group, and multilocus sequence types. In vitro biofilm formation was assessed for 47 IAI and 47 SSTI isolates using a microtiter plate assay. Conditional logistic regression was performed for analysis of matched data (STATA11, College Station, TX). RESULTS: Forty-seven IAI and 188 SSTI isolates were studied. IAI isolates were more often methicillin susceptible S. aureus and non-USA300 vs. SSTI isolates [34 (72%) vs. 79 (42%), p = 0.001 and 38 (81%) vs. 57 (30%) p <0.001, respectively]. Greater than 98% of isolates carried clfA, clfB, efb, isdA, isdB, and icaA,D,B,C while cna was more frequently found among IAI vs. SSTI isolates (p = 0.003). Most isolates were strong biofilm producers. CONCLUSIONS: S. aureus IAI isolates were significantly more likely to be MSSA and non-USA300 than SSTI isolates. Carriage of MSCRAMMs and biofilm formation did not differ significantly between isolates. Evaluation of genetic polymorphisms and gene expression profiles are needed to further delineate the role of adhesins in the pathogenesis of IAIs.

Increasing Numbers of Staphylococcal Scalded Skin Syndrome Cases Caused by ST121 in Houston, Texas.

BACKGROUND: The molecular epidemiology of Staphylococcus aureus strains causing staphylococcal scalded skin syndrome (SSSS) in the United States has not been described. We analyzed patient and S. aureus isolate characteristics associated with SSSS in children at Texas Children's Hospital. METHODS: Patients with SSSS were identified by ICD9/10 codes and available S. aureus isolates were identified from an ongoing S. aureus surveillance study. Medical records were reviewed for 58 patients with available S. aureus isolates. Isolate analyses included PCR for agr group, pvl (lukSF-PV), tst, eta and etb, pulsed-field gel electrophoresis, multi-locus sequence typing and antimicrobial susceptibilities. RESULTS: Cases of SSSS increased from 2.3/10,000 admissions in 2008 to 52.6/10,000 admissions in 2017 (P < 0.0001). The 58 study cases (57 methicillin-susceptible S. aureus, 1 MRSA) with isolates were from 2013 to 2017. The majority (88%) of isolates was of clonal cluster (CC) 121, agr group IV, pvl, tst and carried eta and/or etb and 26% were clindamycin resistant. Twelve ST121 isolates had high level resistance to mupirocin. Patients were treated with standard supportive care plus systemic antibiotics [clindamycin alone or in combination with another antibiotic (n = 44)]. One patient had a recurrent SSSS and one patient was transferred to a burn unit on day 3. CONCLUSIONS: Cases of SSSS are increasing at our hospital. Most S. aureus strains isolated were of one CC, CC121 and carried eta and etb. Supportive care plus clindamycin was effective treatment. We speculate that CC121 was recently introduced to our region and is responsible for the increasing numbers of SSSS cases observed at Texas Children's Hospital.

Clinical Characteristics and Outcomes of Staphylococcus aureus Implant-associated Infections in Children.

BACKGROUND: Staphylococcus aureus is a significant cause of implant-associated infections (IAIs). Data detailing the optimal treatment of IAIs are lacking in children. We describe the clinical features and outcomes of pediatric patients with S. aureus IAIs seen at Texas Children's Hospital. METHODS: Patients and their isolates were identified from a S. aureus surveillance database from 2008 to 2016 in Houston, TX. Demographic and clinical data were collected retrospectively. Fisher's exact was used for statistical analysis. RESULTS: Forty-five patients with 47 IAIs were identified. Most patients had an infected orthopedic implant: 22 (47%) spinal rods and 19 (40%) with other orthopedic hardware. Thirty (64%) IAIs developed within 90 days of implant placement. Six patients had polymicrobial infections and 3 patients were bacteremic. Of the 47 IAI isolates, 34 (72%) were methicillin-susceptible S. aureus (MSSA) and 13 (28%) were methicillin-resistant S. aureus. All children underwent surgical irrigation, debridement and antibiotic therapy. Of the 47 IAI episodes, 22 of the implants were removed at time of initial presentation, 7 implants had delayed removal, and 18 implants remained in place. Successful treatment was achieved in all patients with immediate implant removal (22/22) and in 83% of patients with implant retention (15/18), including 10 patients with early postoperative infections (<3 months) and 5 patients with late postoperative infections (>3 months). Four patients had recurrence of infection. CONCLUSIONS: The majority of S. aureus IAIs were methicillin-susceptible S. aureus. All children with immediate implant removal and most children with retained implants were treated successfully with surgery and antibiotic therapy.

Analysis of Invasive Community-Acquired Methicillin-Susceptible Staphylococcus aureus Infections During a Period of Declining Community Acquired Methicillin-Resistant Staphylococcus aureus Infections at a Large Children's Hospital.

BACKGROUND: The epidemiology of community acquired (CA) Staphylococcus aureus infections is changing in the United States. We investigated the current epidemiology of S. aureus infections at Texas Children's Hospital. METHODS: Patients with CA-S. aureus skin and soft tissue and invasive infections were retrospectively identified from January 1, 2007 to December 31, 2014. Invasive CA-MSSA isolates were characterized by pulsed field gel electrophoresis, Spa typing, agr type and presence of lukSF-PV (pvl) genes. Medical records were reviewed. Statistical analyses included Fisher exact, χ for trend and Wilcoxon tests. RESULTS: CA-MRSA infections decreased by 60.4% (1461-578 infections) from 2007 to 2014 (P < 0.0001), while CA-MSSA infections averaged 550 infections annually. Invasive CA-MRSA infections decreased by 67.2% from 61 to 20 infections (P < 0.0001); invasive CA-MSSA averaged 44 infections annually. Among 296 invasive CA-MSSA isolates, 74 (25%) isolates were USA300 and 88 (30%) were pvl+. USA300 declined among invasive CA-MSSA over time (P < 0.008). Musculoskeletal infections were most common (242/296, 82%); 52/242 (21.5%) isolates were USA300 and 62/242 (25.6%) pvl+. All 18 isolates from musculoskeletal infections with deep venous thrombosis and/or septic shock were pvl+ and 16/18 (88.9%) were USA300. Pneumonia isolates were mainly USA300 (8, 66.7%) and pvl+ (11, 91.7%). CONCLUSIONS: MSSA now cause the majority of invasive CA-S. aureus infections at our institution. Molecular analysis of invasive CA-MSSA isolates suggests strain diversity with USA300 on the decline and that disease presentations are to some extent strain specific. Changes in the CA-S. aureus epidemiology may, in part, be related to changes in immunity to the USA300 clone in the general population.

Healthcare-associated Staphylococcus aureus Bacteremia in Children: Evidence for Reverse Vancomycin Creep and Impact of Vancomycin Trough Values on Outcome.

INTRODUCTION: Elevated vancomycin minimum inhibitory concentrations (MICs) in Staphylococcus aureus have been associated with worse clinical outcomes in adults. For invasive meticillin-resistant S. aureus (MRSA) infections in adults, the Infectious Diseases Society of America recommends targeting vancomycin serum trough concentrations between 15 and 20 µg/mL. We evaluated trends in vancomycin MICs from healthcare-associated (HCA) S. aureus bacteremia isolates in children in addition to correlating vancomycin serum trough levels with clinical outcomes. METHODS: Patients and isolates were identified from a prospective S. aureus surveillance study at Texas Children's Hospital (TCH). HCA S. aureus bacteremia isolates from 2003 to 2013 were selected. Vancomycin MICs by E-test were determined and medical records were reviewed. Acute kidney injury (AKI) was defined as doubling of the baseline serum creatinine. RESULTS: Three hundred forty-one isolates met inclusion criteria. We observed a reverse vancomycin creep among MRSA isolates in the study period with a decline in the proportion of isolates with vancomycin MIC ≥ 2 µg/mL (from 32.7% to 5.6%; P < 0.001). However, the proportion of MSSA isolates with MIC ≥ 2 µg/mL increased (from 2.9% to 9%; P = 0.04). Among patients who had vancomycin troughs performed, there was no difference in duration of bacteremia or fever with vancomycin trough >15 versus <15 µg/mL. A vancomycin trough >15 µg/mL was, however, an independent risk factor for AKI. CONCLUSIONS: Vancomycin MICs are shifting among HCA S. aureus bacteremia isolates with significant differences between MRSA and MSSA at TCH. Higher vancomycin troughs did not improve outcomes in pediatric HCA S. aureus bacteremia but were associated with increased nephrotoxicity. Further studies are needed to better understand optimal management of children with S. aureus bacteremia.

Impact of the 13-valent pneumococcal conjugate vaccine on chronic sinusitis associated with Streptococcus pneumoniae in children.

BACKGROUND: The widespread use of the 7-valent pneumococcal conjugate vaccine has been associated with epidemiologic changes of mucosal and invasive pneumococcal disease. No study describes the impact of 13-valent pneumococcal conjugate vaccine (PCV13) on chronic sinusitis in children. We describe changes in epidemiology of Streptococcus pneumoniae chronic sinusitis after the introduction of PCV13 at Texas Children's Hospital. METHODS: We identified patients <18 years with positive sinus culture for S. pneumoniae who underwent endoscopic sinus surgery because of chronic sinusitis from August 2008 to December 2013 at Texas Children's Hospital. Isolates were serotyped by the capsular swelling method. Demographic and clinical information was collected retrospectively. The χ test and Fisher's exact test were used to analyze dichotomous variables. RESULTS: We identified 91 cases of chronic sinusitis with positive sinus culture for S. pneumoniae. Sixty-one (67%) isolates were non-PCV13 serotypes. PCV13 cases decreased 31% in the post-PCV13 period (P = 0.003). Serotype 19A decreased 27% in the post-PCV13 period (P = 0.007), but accounted for all the isolates with penicillin minimal inhibitory concentration ≥ 4 µg/mL and ceftriaxone minimal inhibitory concentration ≥ 2 µg/mL. Serotypes 19A (38%) and 15C (17%) were the most common in the pre- and post-PCV13 periods, respectively. The most common organism co-isolated was Haemophilus influenzae (52%). Isolation of Prevotella spp. increased in the post-PCV13 period (P = 0.02). CONCLUSIONS: S. pneumoniae continues to represent an important pathogen in chronic sinusitis in children <5 years of age. After the introduction of PCV13, S. pneumoniae isolation declined in children with chronic sinusitis at Texas Children's Hospital. We also observed a substantial reduction of PCV13 serotypes, predominantly serotype 19A.

Randomized trial of "bleach baths" plus routine hygienic measures vs. routine hygienic measures alone for prevention of recurrent infections.

Children with probable community-associated Staphylococcus aureus skin and soft tissue or invasive infections were randomized to routine daily hygienic measures with or without "bleach baths" twice a week for 3 months. Within 12 months, a medically attended recurrence occurred in 84 of 495 (17%) children using bleach baths compared to 103 of 492 (21%) of control participants (P = .15).

Changes in Streptococcus pneumoniae serotype 19A invasive infections in children from 1993 to 2011.

Among 594 Streptococcus pneumoniae serotype 19A invasive pneumococcal disease (IPD) isolates collected from 1993 to 2011, we identified 85 sequence types by multilocus sequence typing. CC320 was associated with multidrug resistance and reduced susceptibility to penicillin and ceftriaxone and still predominated among declining serotype 19A IPD isolates following PCV13 introduction.

Staphylococcus aureus infections in pediatric patients with diabetes mellitus.

OBJECTIVES: To describe Staphylococcus aureus infections in children with diabetes mellitus (DM). METHODS: Children with DM (cases) and S. aureus infections (2/02-6/10) were identified from a surveillance database. Patient charts were reviewed, and S. aureus isolates were characterized by molecular methods. Cases were compared to age-matched controls without DM but with CA-S. aureus skin and soft tissue infections (SSTI) using conditional logistic regression. RESULTS: Forty-seven cases were identified; 41 were matched with 123 controls. Four cases had osteomyelitis and 43 had SSTI. Mean age was 14.2 years and 63% of cases had hemoglobin (Hb) A1c levels above 10%. Cases and controls differed by gender (85% vs. 45% female, P < 0.001), BMI% (median 87% vs. 72%, P = 0.04), methicillin-resistant S. aureus (MRSA) infection (49% vs. 68%, P = 0.04), and recurrent infections (22% vs. 4%, P = 0.001). Among cases, 88% of recurrences were caused by MRSA. CONCLUSIONS: The majority of cases had poor glycemic control, more recurrences, fewer primary MRSA infections and were more likely to be female compared to a control group. Improved glycemic control may reduce the risk for infection, and decrease hospitalizations due to S. aureus infections.

Characterization of nontypeable and atypical Streptococcus pneumoniae pediatric isolates from 1994 to 2010.

Streptococcus pneumoniae is a major cause of bacteremia, meningitis, pneumonia, sinusitis, and acute otitis media in children. Although optochin susceptibility, bile solubility, and Quellung testing are the standards for identifying and differentiating pneumococci, there are several reports of nontypeable pneumococci that give inconsistent results with one or more of these tests. We characterized 52 isolates previously labeled as nontypeable pneumococci. Microbiological methods included repeating the Quellung reaction using a new and expanded group of antisera, optochin susceptibility and bile solubility tests, and automated Vitek 2 identification. Molecular methods included PCR detection of ply and psaA genes, multilocus sequence typing (MLST), 16S rRNA gene sequencing, and pyrosequencing. Of the 52 isolates, 38 (73%) were optochin susceptible, were psaA and ply positive, and could be serotyped by the Quellung reaction. The remaining 14 isolates, isolated from patients with otitis media (n = 6), bacteremia (n = 6), meningitis (n = 1), and pneumonia (n = 1), underwent further analysis. Three of these 14 isolates were nontypeable due to autoagglutination but were pneumococci by all tests and represented pneumococcal sequence types previously recognized by MLST. The 11 remaining isolates were optochin resistant, and 6 of these were bile soluble. Three of 11 were both psaA and ply positive and clustered with pneumococci by MLST (2 were bile soluble); 8 lacked psaA (5 ply positive, 4 bile soluble) and likely belonged to other Streptococcus species. In conclusion, few isolates were truly nontypeable by Quellung reaction, and MLST and the presence of psaA proved useful in distinguishing between atypical pneumococci and other streptococcal species.

Increase in prevalence of Streptococcus pneumoniae serotype 6C at Eight Children's Hospitals in the United States from 1993 to 2009.

Streptococcus pneumoniae serotype 6C, which was described in 2007, causes invasive disease in adults and children. We investigated the prevalence of 6C among pediatric isolates obtained from eight children's hospitals in the United States. S. pneumoniae isolates were identified from a prospective multicenter study (1993 to 2009). Fifty-seven serotype 6C isolates were identified by multiplex PCR and/or Quellung reaction. Five were isolated before 2000, and the prevalence increased over time (P < 0.000001). The median patient age was 2.1 years (range, 0.2 to 22.5 years). Clinical presentations included bacteremia (n = 24), meningitis (n = 7), pneumonia (n = 4), abscess/wound (n = 3), mastoiditis (n = 2), cellulitis (n = 2), peritonitis (n = 1), septic arthritis (n = 1), otitis media (n = 10), and sinusitis (n = 3). By broth microdilution, 43/44 invasive serotype 6C isolates were susceptible to penicillin (median MIC, 0.015 µg/ml; range, 0.008 to 2 µg/ml); all were susceptible to ceftriaxone (median MIC, 0.015 µg/ml; range, 0.008 to 1 µg/ml). By disk diffusion, 16/44 invasive isolates (36%) were nonsusceptible to erythromycin, 19 isolates (43%) were nonsusceptible to trimethoprim-sulfamethoxazole (TMP-SMX), and all isolates were clindamycin susceptible. Multilocus sequence typing (MLST) revealed 24 sequence types (STs); 9 were new to the MLST database. The two main clonal clusters (CCs) were ST473 and single-locus variants (SLVs) (n = 13) and ST1292 and SLVs (n = 23). ST1292 and SLVs had decreased antibiotic susceptibility. Serotype 6C causes disease in children in the United States. Emerging CC1292 expressed TMP-SMX resistance and decreased susceptibility to penicillin and ceftriaxone. Continued surveillance is needed to monitor changes in serotype prevalence and possible emergence of antibiotic resistance in pediatric pneumococcal disease.

Staphylococcus aureus pneumonia in children in the era of community-acquired methicillin-resistance at Texas Children's Hospital.

BACKGROUND: Community-acquired Staphylococcus aureus (SA) pneumonia has increased in children, yet few studies have focused on this infection. METHODS: Patients with SA pneumonia (not ventilator-associated) were identified from our surveillance database. Medical records were reviewed; isolates were genotyped by PFGE and Panton-Valentine leukocidin genes detected by polymerase chain reaction. RESULTS: From August 2001 to April 2009, 117 patients had SA pneumonia. The rate of SA pneumonia per 10,000 admissions increased from 4.81 hospitalizations in year 1 to 9.75 in year 7 (P = 0.04). Methicillin-resistant SA (MRSA) caused 74% and methicillin-susceptible SA (MSSA) caused 26% of the infections. USA300 represented 75/82 (92%) of the MRSA and 14/28 (50%) of the MSSA isolates (P < 0.01). Patients with MRSA were younger (median [range], 0.8 years [0.1-16.9 years]) than patients with MSSA infections (2.5 years [0.2-20.9 years]) (P = 0.008). Clinical presentation was pneumonia with or without effusion in 30, empyema in 72, or lung abscess in 15 cases. Viral coinfections in 18/68 patients tested were associated with respiratory failure (72% vs. 24% [P < 0.001]). Thirty-five children were intubated and 68 had intensive care unit care; 89, 25, and 3 had video-assisted thoracoscopy (VATS), thoracentesis, and lobectomy, respectively. VATS was used more for USA300 than non-USA300 infections (80% vs. 57% [P = 0.03]). In all, 88 children received clindamycin. Improvement or cure occurred in 103 patients (88%), unscheduled visit or readmission related to the same problem in 6, respiratory sequelae in 7, and death in 1 patient. CONCLUSIONS: SA pneumonia increased in frequency over the study years and most were caused by community-acquired MRSA and USA300 isolates. Viral coinfection in 15% of the cases was associated with respiratory failure. Clindamycin is an effective treatment for susceptible-SA pneumonia; VATS was more common in patients with USA300 infections.

Nasal Staphylococcus aureus colonization among mothers of term and late preterm previously healthy neonates with community-acquired Staphylococcus aureus infections.

We enrolled 35 case neonates with community-acquired Staphylococcus aureus infection and their mothers and 19 control mother-neonate pairs. We obtained neonatal and maternal anterior nasal cultures, and clinical isolates. S. aureus nasal colonization was greater in case than control pairs. Neonates were more often infected with their nasal strain than their mother's nasal strain.

Staphylococcus aureus sinus infections in children.

OBJECTIVE: Staphylococcus aureus can cause sinusitis in children. The predominant MRSA clone in the United States, USA300, has been associated with skin and soft tissue as well as invasive diseases. USA300 has increased among CA methicillin-susceptible S. aureus (CA-MSSA) isolates. We describe the clinical characteristics of pediatric patients with S. aureus cultured from sinus specimens, treated at Texas Children's Hospital (TCH), and characterized their isolates by molecular methods. METHODS: This was a retrospective study of children with endoscopic sinus surgery (ESS) cultures positive for S. aureus between 01/2005 and 12/2008 at TCH. Medical records were reviewed and associated S. aureus isolates were characterized by pulsed field gel electrophoresis (PFGE). Data were analyzed by Mann-Whitney U, Chi-square, Fisher's exact test, and Chi-square for trend. RESULTS: We identified 56 patients with S. aureus sinus infections; 12 (21%) were MRSA. Seven of 12 (58%) MRSA vs. 5/44 (11%) MSSA were USA300 (p<0.01). All MRSA isolates were non-susceptible to erythromycin compared to 30% of MSSA (p<0.01); 75% of the USA300 strains were non-susceptible to erythromycin compared to 36% of the non-USA300 strains (p<0.04). Co-pathogens were isolated from 77% (43/56) of the patient specimens. Both MRSA and USA300 isolates were associated with Haemophilus influenzae co-isolation (p<0.05). Patients with USA300 strains were significantly younger (p=0.02) and more likely to experience snoring as a symptom associated with their sinusitis (p=0.03) than those infected with non-USA300 strains. Children with MRSA (4/12) tended to have a greater recurrence rate than children with MSSA isolates (5/44) (p=0.09). No significant differences were observed between groups for fever or complications such as neck cellulitis, nasal abscess, meningitis, subdural empyema, and orbital cellulitis. CONCLUSION: MSSA was more commonly isolated than MRSA from sinus cultures of children who underwent ESS at TCH. The majority of ESS cultures positive for S. aureus, were mixed with other respiratory pathogens, principally H. influenzae. USA300 was the major clone among the MRSA sinusitis isolates, but was not associated with more complications than other S. aureus isolates.

Hospital-acquired Staphylococcus aureus infections at Texas Children's Hospital, 2001-2007.

OBJECTIVE: To document the introduction of the methicillin-resistant Staphylococcus aureus (MRSA) USA300 clone into a children's hospital. Current molecular epidemiology of infections due to the USA300 strain of MRSA in the pediatric healthcare setting remains obscure. DESIGN: Retrospective study of patients with hospital-acquired S. aureus infection during the period from August 1, 2001, through July 31, 2007, at Texas Children's Hospital in Houston. METHODS: Patients with hospital-acquired S. aureus infection from whom an isolate was available for molecular analysis were included. Clinical information was obtained from patient medical records and the electronic hospital information system. S. aureus isolates underwent antimicrobial susceptibility testing, pulsed-field gel electrophoresis, and polymerase chain reaction testing for staphylococcal cassette chromosome (SCC) mec, agr, the diamine N-acetyltransferase gene, and the Panton-Valentine leukocidin genes (pvl). RESULTS: Of 242 patients with hospital-acquired S. aureus infection, 147 (61%) had methicillin-susceptible S. aureus infection. Of the 95 MRSA isolates causing hospital-acquired infection, 69 (73%) were USA300 isolates, and that rate did not increase over time. Skin and soft tissue infection (P < .001), onset of infection less than 10 days after admission (P = .007), and lack of comorbidities (P < .001) were associated with hospital-acquired MRSA infection caused by the USA300 strain, compared with other isolates (hereafter referred to as non-USA300 isolates). Nine of 10 patients with a S. aureus infection at the time of death were infected with a non-USA300 strain. USA300 carried SCCmec IV, agr I, the diamine N-acetyl transferase gene, and pvl. USA300 isolates were more susceptible to clindamycin, gentamicin, and trimethoprim-sulfamethoxazole than were other non-USA300 isolates (P < .01). CONCLUSIONS: In our patient population, the annual numbers of observed cases of hospital-acquired S. aureus infection have remained constant. USA300 was the most common clone and, compared with other non-USA300 MRSA isolates, was associated with skin and soft tissue infection, early onset of infection after admission, and greater susceptibility to antimicrobial agents.

Vancomycin MICs for Staphylococcus aureus vary by detection method and have subtly increased in a pediatric population since 2005.

Vancomycin MICs for Staphylococcus aureus isolates in a pediatric hospital with a high rate of staphylococcal infections were examined for any increase over a 7-year period. A broth microdilution scheme allowed direct comparison of the MICs generated by this method to MICs generated by Etest. MICs generated by both methods were determined with the same inoculum suspension. One hundred sixty-five S. aureus isolates were selected on the basis of the patients having been bacteremic or having received vancomycin as the definitive therapy for their infections. Of the 165 isolates, 117 were methicillin-resistant S. aureus and 48 were methicillin-susceptible S. aureus. Forty-seven were acquired in the hospital (nosocomial), 56 were community acquired, and 62 were community onset-health care associated. All but one isolate tested by broth microdilution had MICs of < 1.0 microg/ml, while 96% of these same isolates tested by Etest had MICs of > or = 1 microg/ml. A significant increase in MICs that occurred after study year 4 (2004 to 2005) was demonstrated by the Etest (P < 0.00007) but not by broth microdilution. MICs were not different for isolates of community or health care origin, regardless of methodology. The proportion of isolates with Etest MICs of < 1 and > or = 1 microg/ml between children with bacteremia for < or = 5 days and > 5 days (P = 0.3) was not different. We conclude that MICs for pediatric isolates have increased slightly since 2005 and therapeutic decisions based on vancomycin MICs need to be made by considering the methodology used.