Judy A. Daly

Genome sequence and comparative microarray analysis of serotype M18 group A Streptococcus strains associated with acute rheumatic fever outbreaks

Acute rheumatic fever (ARF), a sequelae of group A Streptococcus (GAS) infection, is the most common cause of preventable childhood heart disease worldwide. The molecular basis of ARF and the subsequent rheumatic heart disease are poorly understood. Serotype M18 GAS strains have been associated for decades with ARF outbreaks in the U.S. As a first step toward gaining new insight into ARF pathogenesis, we sequenced the genome of strain MGAS8232, a serotype M18 organism isolated from a patient with ARF. The genome is a circular chromosome of 1,895,017 bp, and it shares 1.7 Mb of closely related genetic material with strain SF370 (a sequenced serotype M1 strain). Strain MGAS8232 has 178 ORFs absent in SF370. Phages, phage-like elements, and insertion sequences are the major sources of variation between the genomes. The genomes of strain MGAS8232 and SF370 encode many of the same proven or putative virulence factors. Importantly, strain MGAS8232 has genes encoding many additional secreted proteins involved in human–GAS interactions, including streptococcal pyrogenic exotoxin A (scarlet fever toxin) and two uncharacterized pyrogenic exotoxin homologues, all phage-associated. DNA microarray analysis of 36 serotype M18 strains from diverse localities showed that most regions of variation were phages or phage-like elements. Two epidemics of ARF occurring 12 years apart in Salt Lake City, UT, were caused by serotype M18 strains that were genetically identical, or nearly so. Our analysis provides a critical foundation for accelerated research into ARF pathogenesis and a molecular framework to study the plasticity of GAS genomes.

An Epidemiological Investigation of a Sustained High Rate of Pediatric Parapneumonic Empyema: Risk Factors and Microbiological Associations

We investigated the increasing incidence of pediatric empyema during the 1990s at Primary Childrens Medical Center in Salt Lake City. Of 540 children hospitalized with community-acquired bacterial pneumonia (CAP) who were discharged from 1 July 1993 through 1 July 1999, 153 (28.3%) had empyema. The annual population incidence of empyema increased during the study period from 1 to 5 cases per 100,000 population aged <19 years. Streptococcus pneumoniae was identified as the most common cause of CAP with or without empyema; serotype 1 accounted for 50% of the cases of pneumococcal empyema. Patients with empyema were more likely to be >3 years old, to have > or =7 days of fever, to have varicella, and to have received antibiotics and ibuprofen before admission to the hospital, compared with patients without empyema (P<.0001 for each factor). The increasing incidence of empyema was associated with infection due to S. pneumoniae serotype 1, outpatient treatment with certain antibiotics, ibuprofen use, and varicella.

Impact of the pneumococcal conjugate vaccine on pneumococcal parapneumonic empyema.

Background: Pediatric pneumococcal parapneumonic empyema (PPE) has become increasingly common. In the last decade, Utah has had one of the highest rates of PPE in the United States, 14/100,000 children, attributed primarily to Streptococcus pneumoniae serotype 1. Our objective was to describe the temporal trends in PPE in Utah before and after the availability of the 7-valent pneumococcal conjugate vaccine (PCV-7). Methods: The Intermountain Health Care (IHC) data warehouse was queried for all cases of empyema in children younger than 18 years, defined as International Classification of Diseases, 9th revision, Clinical Modification code 510.9, for the study period March 1996–June 2005. We also retrieved and serotyped all blood and pleural fluid isolates of S. pneumoniae from children younger than 18 years with a diagnosis of PPE at Primary Childrens Medical Center (PCMC) between March 1996 and June 2005. The pre-PCV-7 period (PRE) included 57 months (March 1996–December 2000) and the post-PCV-7 period (POST) included 54 months (January 2001–June 2005). Results: We identified 776 cases of pediatric empyema in the IHC system, and 478 (62%) were managed at PCMC. In the years 1996–2000, we managed a mean of 38 cases of empyema per year compared with 71.5 cases per year between 2001 and 2004 (P = 0.006). At PCMC, there were 295 cases of invasive pneumococcal disease (IPD), and 74 (25%) were PPE. During the PRE period, PPE represented 24 of 137 (17.5%) cases of IPD compared with 50 of 158 (32%) in the POST period (P = 0.008). One-half of the children with PPE required intensive care and 4 died. During the PRE and POST periods, PPE was most often caused by serotype 1 (46 and 34%, respectively), but in the POST period serogroups 3 (20%), and 19A (14%) were also prevalent. PPE in PCV-7-immunized children was caused exclusively by nonvaccine serotypes. Conclusions: PPE in the post-PCV-7 era is more common, representing one-third of the IPD in children in UT. PPE is associated with significant morbidity and mortality. Serotype 1 remains the most common cause of PPE, but serotypes 3 and 19A are emerging.

Temporal trends of invasive disease due to Streptococcus pneumoniae among children in the intermountain west : Emergence of nonvaccine serogroups

BACKGROUND Use of the heptavalent pneumococcal conjugate vaccine (PCV-7 [Prevnar]) has been associated with decreased a incidence of invasive pneumococcal disease (IPD) among children in the United States. METHODS Cases of IPD in children < 18 years of age insured by or receiving health care from Intermountain Health Care during 1996-2003 were identified. Isolates of S. pneumoniae from children with IPD treated at Primary Childrens Medical Center (PCMC; Salt Lake City, UT) during 1997-2003 were serogrouped. Temporal trends of IPD, serogroup distribution of pneumococci, and antibiotic resistance among pneumococci were analyzed. RESULTS A total of 1535 cases of IPD were identified. The rate of IPD decreased 27% after the introduction of PCV7. Among children with IPD who were cared for at PCMC, disease in 73% was caused by PCV7 serogroups in 1997-2000, compared with 50% in 2001-2003 (P < .001), and the percentage of isolates resistant to penicillin decreased from 34% in 1997-2000 to 22% in 2001-2003 (P = .04). The percentage of IPD cases that were empyema increased from 16% to 30% (P = .015), and the percentage of severe cases of IPD increased from 57% to 71% (P = .026). Children with IPD due to non-PCV7 serogroups were older, were more likely to have parapneumonic empyema, and had longer hospital stays. CONCLUSIONS The incidence of IPD in the IMW decreased by 27% after the introduction of the PCV7 vaccine. During the postvaccine period (2001-2003), there were significant decreases in the proportion of cases of IPD caused by PCV7 and antibiotic-resistant serogroups. These benefits were accompanied by a significant increase in the proportion of IPD cases due to non-PCV7 serogroups, with increases in the incidence of empyema and severe IPD.

FilmArray, an Automated Nested Multiplex PCR System for Multi-Pathogen Detection: Development and Application to Respiratory Tract Infection

The ideal clinical diagnostic system should deliver rapid, sensitive, specific and reproducible results while minimizing the requirements for specialized laboratory facilities and skilled technicians. We describe an integrated diagnostic platform, the “FilmArray”, which fully automates the detection and identification of multiple organisms from a single sample in about one hour. An unprocessed biologic/clinical sample is subjected to nucleic acid purification, reverse transcription, a high-order nested multiplex polymerase chain reaction and amplicon melt curve analysis. Biochemical reactions are enclosed in a disposable pouch, minimizing the PCR contamination risk. FilmArray has the potential to detect greater than 100 different nucleic acid targets at one time. These features make the system well-suited for molecular detection of infectious agents. Validation of the FilmArray technology was achieved through development of a panel of assays capable of identifying 21 common viral and bacterial respiratory pathogens. Initial testing of the system using both cultured organisms and clinical nasal aspirates obtained from children demonstrated an analytical and clinical sensitivity and specificity comparable to existing diagnostic platforms. We demonstrate that automated identification of pathogens from their corresponding target amplicon(s) can be accomplished by analysis of the DNA melting curve of the amplicon.

Epidemiology, complications, and cost of hospitalization in children with laboratory-confirmed influenza infection

BACKGROUND. Influenza causes significant morbidity among children. Previous studies used indirect case ascertainment methods with little cost data. We sought to measure the burden of laboratory-confirmed influenza from hospitalized children. METHODS. We conducted a retrospective cohort study during 3 viral seasons at Primary Childrens Medical Center (Salt Lake City, UT). Children ≤18 years of age who were hospitalized with laboratory-confirmed influenza infection were included. Outcomes included hospitalization rates, complications including intensive care unit stays, mechanical ventilation, length of stay, and total hospital costs. RESULTS. A total of 325 children had hospitalizations attributable to influenza over 3 viral seasons: 28% <6 months of age, 33% between 6 and 23 months of age; and 39% >2 years of age; 37% had high-risk medical conditions. Population-based rates of hospitalization for Salt Lake County residents ranged from 6.3 to 252.7 per 100000 children. The highest rates were in children younger than 6 months, and rates decreased with increasing age. Forty-nine (15%) children had an ICU stay; 27 required mechanical ventilation, and half of these patients were >2 years of age. Total hospital cost for the cohort was

Rapid identification of pathogens from positive blood cultures by multiplex polymerase chain reaction using the FilmArray system.

2 million; 55% was accounted for by children >2 years of age. Length of stay and total hospital costs were significantly higher in all children >2 years of age compared with children <6 months of age and were comparable to all children 6 to 23 months of age. CONCLUSIONS. Proven influenza infection in children results in substantial hospital resource utilization and morbidity. Nationwide, the median hospital costs may total

Seasonal Invasive Pneumococcal Disease in Children: Role of Preceding Respiratory Viral Infection

55 million. Our data support the Advisory Committee on Immunizations recommendations to expand the use of influenza vaccine to children >2 years of age.

Pneumococcal Necrotizing Pneumonia in Utah: Does Serotype Matter?

Sepsis is a leading cause of death. Rapid and accurate identification of pathogens and antimicrobial resistance directly from blood culture could improve patient outcomes. The FilmArray® (FA; Idaho Technology, Salt Lake City, UT, USA) Blood Culture (BC) panel can identify >25 pathogens and 4 antibiotic resistance genes from positive blood cultures in 1 h. We compared a development version of the panel to conventional culture and susceptibility testing on 102 archived blood cultures from adults and children with bacteremia. Of 109 pathogens identified by culture, 95% were identified by FA. Among 111 prospectively collected blood cultures, the FA identified 84 (91%) of 92 pathogens covered by the panel. Among 25 Staphylococcus aureus and 21 Enterococcus species detected, FA identified all culture-proven methicillin-resistant S. aureus and vancomycin-resistant enterococci. The FA BC panel is an accurate method for the rapid identification of pathogens and resistance genes from blood culture.

Molecular Epidemiology of Pediatric Pneumococcal Empyema from 2001 to 2007 in Utah

OBJECTIVE. Our objective was to demonstrate correlations between invasive pneumococcal disease in children and circulating respiratory viruses. METHODS. This retrospective study included 6 winter respiratory viral seasons (2001–2007) in Intermountain Healthcare, an integrated health system in the Intermountain West, including Primary Childrens Medical Center in Salt Lake City, Utah. Children <18 years of age who were hospitalized with either invasive pneumococcal disease in any Intermountain Healthcare facility or culture-confirmed invasive pneumococcal disease at Primary Childrens Medical Center were included. We analyzed the correlation between invasive pneumococcal disease and circulating respiratory viruses. RESULTS. A total of 435 children with invasive pneumococcal disease and 203 with culture-confirmed invasive pneumococcal disease were hospitalized in an Intermountain Healthcare facility or Primary Childrens Medical Center during the study period. During the same period, 6963 children with respiratory syncytial virus, 1860 with influenza virus, 1459 with parainfluenza virus, and 818 with adenoviruses were evaluated at Primary Childrens Medical Center. A total of 253 children with human metapneumovirus were identified during the last 5 months of the study. There were correlations between invasive pneumococcal disease and seasonal respiratory syncytial virus, influenza virus, and human metapneumovirus activity. The correlation with invasive pneumococcal disease was strong up to 4 weeks after respiratory syncytial virus activity. For influenza virus and human metapneumovirus, the correlations were strong at 2 weeks after activity of these viruses. Pneumonia was the most common clinical disease associated with culture-confirmed invasive pneumococcal disease, mostly attributable to serotypes 1, 19A, 3, and 7F. CONCLUSIONS. In the post–pneumococcal conjugate vaccine era, seasonal increases in respiratory syncytial virus, influenza virus, and human metapneumovirus infections in children were associated with increased pediatric admissions with invasive pneumococcal disease, especially pneumonia caused by nonvaccine serotypes.