Background: Community-acquired pneumonia (CAP) is a leading cause of hospitalization and mortality in children. Diagnosis remains challenging and there are no reliable tools to objectively risk stratify patients or predict clinical outcomes. Molecular distance to health (MDTH) is a genomic score that measures the global perturbation of the transcriptional profile and may help classify patients by disease severity. We evaluated the value of MDTH to assess disease severity in children hospitalized with CAP. Methods: Children hospitalized with CAP and matched healthy controls were enrolled in a prospective observational study. Blood samples were obtained for transcriptome analyses within 24 h of hospitalization. MDTH scores were calculated to assess disease severity and correlated with laboratory markers, such as white blood cell count, c-reactive protein (CRP), and procalcitonin (PCT), and clinical outcomes, including duration of fever and duration of hospitalization (LOS). Univariate and multivariable logistic regression were applied to assess factors associated with LOS and duration of fever after hospitalization. Results: Among children hospitalized with CAP (n = 152), pyogenic bacteria (PB) were detected in 16 (11%), Mycoplasma pneumoniae was detected in 41 (28%), respiratory viruses (RV) alone were detected in 78 (51%), and no pathogen was detected in 17 (11%) children. Statistical group comparisons identified 6,726 genes differentially expressed in patients with CAP vs. healthy controls (n = 39). Children with confirmed PB had higher MDTH scores than those with RV (p < 0.05) or M. pneumoniae (p < 0.01) detected alone. CRP (r = 0.39, p < 0.0001), PCT (r = 0.39, p < 0.0001), and MDTHs (r = 0.24, p < 0.01) correlated with duration of fever, while only MDTHs correlated with LOS (r = 0.33, p < 0.0001). Unadjusted analyses showed that both higher CRP and MDTHs were associated with longer LOS (OR 1.04 [1-1.07] and 1.12 [1.04-1.20], respectively), however, only MDTH remained significant when adjusting for other covariates (aOR 1.11 [1.01-1.22]). Conclusions: In children hospitalized with CAP MDTH score measured within 24 h of admission was independently associated with longer duration of hospitalization, regardless of the pathogen detected. This suggests that transcriptional biomarkers may represent a promising approach to assess disease severity in children with CAP.

Objectives: Animal studies suggest that RSV increases nasopharyngeal (NP) bacterial colonization facilitating bacterial infections. We investigated the influence of antibiotic treatment and colonization with potentially pathogenic bacteria on inflammatory markers and disease severity in RSV-infected in infants.Methods: Healthy young infants hospitalized with RSV bronchiolitis (n = 136) and age-matched healthy controls (n = 23) were enrolled and NP samples cultured for potentially pathogenic bacteria including: Gram-positive bacteria (GPB): Staphylococcus aureus, Streptococcus pneumoniae, beta-hemolytic Streptococcus; and Gram-negative bacteria (GNB): Moraxella catarrhalis and Haemophilus influenzae. Clinical parameters and plasma IL-8, IL-6 and TNF-alpha concentrations were compared according to the bacterial class and antibiotic treatment.Results: Antibiotic treatment decreased by 10-fold NP bacterial recovery. Eighty-one percent of RSV infants who did not receive antibiotics before sample collection were colonized with pathogenic bacteria. Overall, GNB were identified in 21% of patients versus 4% of controls who were mostly colonized with GPB. Additionally, in RSV patients NP white blood cell counts (p = 0.026), and blood neutrophils (p = 0.02) were higher in those colonized with potentially pathogenic bacteria versus respiratory flora. RSV patients colonized with GNB had higher plasma IL-8 (p = 0.01) and IL-6 (p < 0.01) concentrations than controls, and required longer duration of oxygen (p = 0.049).Conclusions: Infants with RSV bronchiolitis colonized with potentially pathogenic bacteria had increased numbers of mucosal and systemic inflammatory cells. Specifically, colonization with GNB was associated with higher concentrations of proinflammatory cytokines and a trend towards increased disease severity. (C) 2015 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

Bordetella parapertussis is widely recognized as a cause of a pertussis-like respiratory illness in children, but reports of invasive infection are rare. We review the literature and describe the clinical presentation and treatment of 2 children with B. parapertussis bacteremia, as well as the techniques used to isolate the organism.

Bordetella parapertussis is widely recognized as a cause of a pertussis-like respiratory illness in children, but reports of invasive infection are rare. We review the literature and describe the clinical presentation and treatment of 2 children with B. parapertussis bacteremia, as well as the techniques used to isolate the organism.

Background. During 9 May 2010-7 May 2011, an outbreak of pertussis-like illness (incidence, 80 cases per 100 000 persons) occurred in Franklin County, Ohio. The majority of cases were identified by IS481-directed polymerase chain reaction (PCR), which does not differentiate among Bordetella species. We sought to determine outbreak etiology and epidemiologic characteristics.Methods. We obtained demographic, clinical, and vaccination-related data from the Ohio Disease Reporting System and Impact Statewide Immunization Information System. We tested sera from 14 patients for anti-pertussis toxin (PT) antibodies and used species-specific PCR on 298 nasopharyngeal specimens.Results. Reported cases totaled 918. IS481 results were available for 10 serologically tested patients; 5 of 10 had discordant anti-PT antibody and IS481 results, suggestive of Bordetella holmesii, which lacks PT and harbors IS481. We identified specific Bordetella species in 164 of 298 specimens tested with multitarget PCR; B. holmesii and Bordetella pertussis were exclusively detected among 48 (29%) and 112 (68%), respectively; both were detected in 4 (2%). Among 48 patients with B. holmesii infections, 63% were aged 11-18 years, compared with 35% of 112 patients with B. pertussis infections (P =.001). Symptoms were similar among B. holmesii- and B. pertussis-infected patients. Adolescent pertussis ("Tdap") booster vaccinations were more effective against B. pertussis than B. holmesii (effectiveness: 67% and 36%, respectively; 95% confidence intervals, 38%-82% and -33% to 69%, respectively).Conclusions. We report the first documented mixed outbreak of B. pertussis and B. holmesii infections. Bordetella holmesii particularly affected adolescents. Although laboratory capacity limitations might inhibit routine use of multitarget PCR for clinical diagnosis, focused testing and enhanced surveillance might improve understanding the burden of B. holmesii infection.

The use of herpes simplex virus (HSV) polymerase chain reaction for diagnosis of HSV disease involving the central nervous system has not translated into widespread use for the detection of DNAemia. We report our 6-year experience using blood polymerase chain reaction testing for HSV infection in neonates and older children with HSV disease. (J Pediatr 2012;161:357-61)

A 2.5-month-old white female with no significant prenatal history presented to the emergency department (ED) with diarrhea that was frequent, foul-smelling, and nonbloody. She was born at full-term by C-section due to late decelerations and had been bottle-fed with premixed formula since birth. The patient was diagnosed by her primary care physician with bronchiolitis 4 weeks before admission, and had cough, congestion, and post-tussive emesis that were improved since then, but not completely resolved. Six days before admission, she developed diarrhea associated with nonbilious, nonbloody emesis, and fever to up to 38.8°C. At that time, she was evaluated in the ED where her white blood cell (WBC) count was 7300/mm3, with a differential count of 28% neutrophils, 57% lymphocytes, and 15% monocytes. The hemoglobin was 9.2 mg/dL and platelet count was 406,000/mm3. A urinalysis was normal, and a blood culture was obtained. A chest radiograph showed bilateral upper lobe atelectasis without consolidation and a normal thymic silhouette. The patient was diagnosed with viral gastroenteritis and was discharged home; however, she continued having approximately 6 loose, foul-smelling, nonbloody stools each day. Six days later, after a Gram-negative bacterium was identified in the blood culture obtained during the ED visit, the patient was directly admitted to the Infectious Diseases service. On admission, her weight was 4.79 kg (25th–50th percentile), and her height was 54.3 cm (3rd–5th percentile). Her axillary temperature was 36.6°C, heart rate was 144 beats/min, respiratory rate was 40 breaths/min, blood pressure was 80/48 mm Hg, and oxygen saturation was 98% in room air. Physical examination revealed an alert, well-developed and well-nourished, extremely pale infant in no apparent distress. She appeared well-hydrated with moist mucous membranes and negative skin turgor test; however, she had mild erythema and edema around her eyes. Breathing was slightly labored with mild subcostal retractions and bilateral wheezing. Abdomen was soft but distended with normal bowel sounds. The liver edge was palpable at 2 cm below the right costal margin and the spleen was not palpable. Her peripheral pulses were present and symmetric, but capillary refill was delayed from 3 to 5 seconds. The remainder of the physical examination was normal. The peripheral WBC count on the day of admission was 14,700/mm3, with a differential count of 3% neutrophils, 8% bands, 69% lymphocytes, 12% monocytes, 3% myelocytes, 3% eosinophils, and 2% basophils. The hemoglobin was 9.1 mg/dL, hematocrit was 27.2%, and platelet count was 560/mm3. Except for an albumin level of 2.4 g/dL, her metabolic panel was normal. A lumbar puncture was performed and showed no evidence of meningitis (3 WBC; glucose of 46 mg/dL and protein of 31 mg/dL). Blood culture, urine culture, and stool studies, including adenovirus 40/41 antigen, rotavirus antigen, Escherichia coli and Shiga toxin by EIA, Clostridiumdifficile toxin, ova and parasite and stool cultures, were obtained. Empiric therapy with piperacillin/tazobactam was initiated for broad Gram-negative coverage. A basic immunologic workup, including quantitative immunoglobulins (IgA, IgE, IgM, and IgG) as well as T-cell, B-cell, CD45 RA, and natural killer cell immunophenotyping, was normal. What is your diagnosis? Denouement A diagnosis of Campylobacter jejuni bacteremia was made on the basis of a positive anaerobic blood culture after 6 days of incubation. Growth of an anaerobic blood agar plate subculture showed small, round, glistening colonies, which stained poorly as gram-negative spiral and curved rods. Carbol fuchsin counterstaining stained the organism significantly better. Confirmatory identification was accomplished by sequencing a portion of the 16S ribosomal RNA gene. In addition, a stool culture collected on the day of admission was also positive for C. jejuni. Our patient was initially treated with piperacillin–tazobactam but then was switched to azithromycin based on the final identification of the bacterium and susceptibility test results by the E-test method, which demonstrated susceptibility to ciprofloxacin and azithromycin, but resistance to ceftriaxone (minimum inhibitory concentration [MIC] = 32). Of note, cerebrospinal fluid and repeat blood and urine cultures collected on admission before the initiation of antibiotics were negative. During the course of her hospitalization, the patient remained afebrile, and the diarrhea resolved. Stool cultures repeated after 48 hours of antibiotic therapy were negative for Campylobacter spp. The patient was discharged after 72 hours, and completed a 7-day course of oral azithromycin. At follow-up, 3 weeks after discharge, the patient was in excellent health. C. jejuni is a curved, small, motile gram-negative rod that commonly causes inflammatory, bloody diarrhea among people of all ages.1 Population-based studies in Europe and the United States have shown a bimodal distribution, with a peak of illness in children <5 years of age and in people 15 to 29 years of age.1–3 There are currently 3 named genera and over 25 species in the family Campylobacteraceae; however, only 13 are considered pathogens in humans.4 Perinatal infections caused by Campylobacter are more commonly associated with C. fetus than C. jejuni, which is probably related to the increased tropism of the bacteria for fetal tissue. In addition, perinatal infections caused by C. jejuni are less severe than those caused by C. fetus.4 The clinical manifestations of Campylobacter infections in the prenatal period include: abortion or stillbirth, premature labor, and neonatal septicemia and meningitis, which are rarely caused by C. jejuni. In neonates C. jejuni can cause gastroenteritis and asymptomatic bloody diarrhea. The source of the bacteria is generally the mother who can be symptomatic or asymptomatic at delivery. In general, bacteremia caused by these organisms is rare and occurs mostly in immunocompromised patients, especially those with humoral immunodeficiencies, the elderly, or patients with chronic conditions. In the developing world, C. jejuni bacteremia has been described in children with severe malnutrition.2,5–7 Most cases of bacteremia are due to C. fetus, which is less likely to cause enteritis.4 Conversely, C. jejuni predominately causes gastroenteritis, and is responsible for bacteremia in <1% of cases.4 Extraintestinal infections related to C. jejuni other than bacteremia are rare and include cholecystitis, urinary tract infection, and meningitis.8 Bloodstream infection in children <14 years of age with intestinal infection occurs in approximately 1 in 3000 compared with 1 in 170 in patients older than 65 years with enteritis. In infants less than 6 months of age, a recent study identified risk factors for Campylobacter infection including drinking well water and riding in a shopping cart next to meat or poultry. Infected infants were also less likely to be breast-fed than control infants.9 Our patient was bottle fed and had not been exposed to well water. Whether she had been exposed to meat or poultry in a shopping cart was unknown. Three patterns of bacteremia associated with Campylobacter spp. have been described: (1) self-limited bacteremia as an isolated event following acute enteritis and due primarily to C. jejuni; (2) secondary bacteremia associated with a focal infection such as meningitis, pneumonia, or endocarditis and due more commonly to C. fetus; and (3) chronic bacteremia with relapses occurring mostly in immunocompromised patients in the absence of enteritis.6 Taken together, our patient's disease onset at 2 1/2 months of age, the identification of C. jejuni instead of C. fetus in blood and stool cultures, the lack of C. jejuni growth in repeated cultures even before antibiotics were initiated, and the lack of involvement of other anatomic sites (negative urine/cerebrospinal fluid cultures and absence of pneumonia on chest radiograph) suggest that most likely she had transient, self-limited bacteremia associated with enteritis rather than other more severe forms of the disease. Stool culture for the isolation of C. jejuni and C. coli, also commonly associated with gastroenteritis, requires the use of selective media and incubation under microaerophilic conditions at 42°C. Recovery of the organism from blood cultures is more problematic for several reasons: (1) relatively slow growth of Campylobacter spp., (2) poor growth under aerobic conditions, (3) poor staining in anaerobic blood cultures by routine methods, which delays the identification of the organism even if blood culture bottles are “flagged” as positive, and (4) delay in subculturing the anaerobic blood culture bottle to optimal anaerobic conditions. In our case, the organism was never recovered from the aerobic blood culture and only the subculture of the initially flagged anaerobic blood culture yielded the organism. Thus, laboratories should have procedures, such as acridine orange staining (which intercalates into DNA and thus nonspecifically stains all bacteria), available for situations where a blood culture is “flagged” as positive in a semiautomated blood culture system but no organisms are seen on Gram stain. For patients with systemic infections, parenteral therapy should be considered. Campylobacter spp. are generally resistant to the penicillins and cephalosporins with variable susceptibility to the macrolides and fluoroquinolones. C. jejuni is generally susceptible to the macrolides but rates of macrolide resistance in C. coli vary considerably. There are a number of reports of increased fluoroquinolone resistance in most countries including the emergence of resistance during therapy.10,11 In our case, the bacterium was susceptible to azithromycin and ciprofloxacin and resistant to ceftriaxone. She received azithromycin for 7 days with excellent response.

The aim of this study was to elucidate the role of two novel membrane fusion proteins (MFPs) in the susceptibility of Acinetobacter baumannii to antimicrobial agents.The genome sequence of A. baumannii ATCC 17978 contains two open reading frames (ORFs) annotated as AdeT in the NCBI genome database. Both the putative efflux genes display > 30% similarity to known MFPs. The antimicrobial susceptibility profiles of Escherichia coli KAM32 cells carrying the genes were monitored by the broth dilution method. Different efflux pump inhibitors were used for fluorimetric efflux assays. The functions of putative ORFs were confirmed in A. baumannii by insertional inactivation and complementation.E. coli cells carrying the ORFs had decreased susceptibility to antibiotics, disinfectants, dyes and detergents, with enhanced efflux activity. Inactivation of the ORFs and further characterization in A. baumannii confirmed its role in antimicrobial resistance by active efflux.This report describes the functions of novel resistance determinants, members of the MFP family, for the first time in A. baumannii.

This multicenter study evaluated the clinical performance of the 3M Rapid Detection RSV test (3MRSV) compared to a composite reference standard of R-Mix culture and direct specimen immunofluorescence for detection of respiratory syncytial virus (RSV). The performance of the BinaxNOW RSV test was also evaluated using this reference standard. In a secondary analysis, discordant results were arbitrated using the Gen-Probe/Prodesse ProFlu+ reverse transcription-PCR (RT-PCR) assay. Subjects were stratified into three groups as follows: group 1 (G1), all ages; G2, subjects <22 years old (FDA-cleared ages for 3MRSV testing); and G3, subjects <5 years old (FDA-cleared ages for BinaxNOW RSV testing). A total of 1,306 specimens (G1, n = 1,306; G2, n = 1,140; G3, n = 953) from subjects of all ages presenting with respiratory symptoms met study criteria for analysis. Sensitivities, specificities, positive predictive values, and negative predictive values of 3MRSV for G1 were 86.5%, 95.8%, 91.4%, and 93.2%, respectively, and those for G2 were 87.3%, 95.6%, 92.4%, and 92.5%, respectively. For those samples analyzed by both 3MRSV and BinaxNOW, the 3MRSV was more sensitive (G1, 86.3%; G2, 87.2%; and G3, 89.9%) than was BinaxNOW (G1, 70.84%; G2, 72.0%; and G3, 72.4%) (P < 0.05). Specificities for RSV detection from nasopharyngeal (NP) aspirates and NP swabs for all groups were comparable for 3MRSV and BinaxNOW, but 3MRSV was less specific than BinaxNOW when nasal washes/aspirates were tested (P < 0.05). The 3MRSV assay performed well for the detection of RSV, and the overall assay performance was superior to that of BinaxNOW. The 3MRSV reader eliminated user misinterpretation and provided test result and quality control documentation.

ABSTRACT To determine the prevalence of serotypes of Streptococcus pneumoniae responsible for pneumonia with pleural effusion, we determined the capsular polysaccharide (PS) type directly on 49 pleural fluid specimens collected from pediatric patients during 2007 to 2009 with laboratory-confirmed pneumococcal pneumonia by using monoclonal antibodies and a multiplex, bead array immunoassay. Because the fluids had to be heated to remove nonspecific reactivity before being tested in the immunoassay and type 19A PS is heat labile, the pleural fluid samples were also tested for serotype 19A capsule gene locus by PCR. Use of the multiplex immunoassay combined with type-specific 19A PCR allowed for serotype determination on 40 of 49 pleural fluids. Pneumococcal pneumonia with pleural effusion was associated with a limited number of serotypes, with types 1, 3, 7F/A, and 19A accounting for 75% of the typeable cases. The concentration of capsular PS in the pleural fluids was often greater than 1 μg/ml and sufficient to inhibit the opsonic capacity of sera from individuals who had received the 23-valent pneumococcal PS vaccine. Based on the serotypes observed before and after introduction of the 7-valent pneumococcal conjugate vaccine, the recently licensed 13-valent pneumococcal conjugate vaccine may reduce the incidence of pneumonia with pleural effusions.

Pneumococcal serotypes causing pneumonia with pleural effusion in 1 pediatric patients 2 3 Running title: Pneumococcal serotypes and pleural effusions 4 5 Authors: Jigui Yu, Douglas Salamon , Mario Marcon , Moon H. Nahm 1* 6 7 1 Department of Pathology, University of Alabama at Birmingham, Birmingham, 8 AL, 9 2 Department of Laboratory Medicine, Nationwide Children’s Hospital, Columbus, 10 OH. 11 3 Departments of Pediatrics and Pathology, Ohio State University, College of 12 Medicine and Public Health, Columbus, OH. 13 14 Corresponding Author: 15 Moon H. Nahm, M.D. 16 University of Alabama at Birmingham 17 BBRB 614 18 1530 Third Avenue South 19 Birmingham, AL 35294-2170 20 Phone: 205-934-0163 21 Fax: 205-975-5479 22 Email: nahm@uab.edu 23 Copyright © 2010, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved. J. Clin. Microbiol. doi:10.1128/JCM.01827-10 JCM Accepts, published online ahead of print on 1 December 2010

Background Over the last decade, nosocomial infections due to Acinetobacter baumannii have been described with an increasing trend towards multidrug resistance, mostly in intensive care units. The aim of the present study was to determine the clonal relatedness of clinical isolates and to elucidate the genetic basis of imipenem resistance. Methods A. baumannii isolates (n = 83) originated from two hospital settings in central Ohio were used in this study. Pulsed-field gel electrophoresis genotyping and antimicrobial susceptibility testing for clinically relevant antimicrobials were performed. Resistance determinants were characterized by using different phenotypic (accumulation assay for efflux) and genotypic (PCR, DNA sequencing, plasmid analysis and electroporation) approaches. Results The isolates were predominantly multidrug resistant (>79.5%) and comprised of thirteen unique pulsotypes, with genotype VII circulating in both hospitals. The presence of bla OXA-23 in 13% (11/83) and IS Aba1 linked bla OXA-66 in 79.5% (66/83) of clinical isolates was associated with high level imipenem resistance. In this set of OXA producing isolates, multidrug resistance was bestowed by bla ADC-25 , class 1 integron-borne aminoglycoside modifying enzymes, presence of sense mutations in gyrA / parC and involvement of active efflux (with evidence for the presence of adeB efflux gene). Conclusion This study underscores the major role of carbapenem-hydrolyzing class D β-lactamases, and in particular the acquired OXA-23, in the dissemination of imipenem-resistant A. baumannii . The co-occurrence of additional resistance determinant could also be a significant threat.