BackgroundNon-polio human enteroviruses are the leading cause of aseptic meningitis in children. The role of enterovirus PCR for diagnosis and management of aseptic meningitis has not been fully explored.Methodology/Principal FindingsA retrospective study was conducted to determine the epidemiological, clinical, and laboratory characteristics of aseptic meningitis and to evaluate the role of enterovirus PCR for the diagnosis and management of this clinical entity. The medical records of children who had as discharge diagnosis aseptic or viral meningitis were reviewed. A total of 506 children, median age 5 years, were identified. The annual incidence rate was estimated to be 17/100,000 children less than 14 years of age. Most of the cases occurred during summer (38%) and autumn (24%). The dominant clinical symptoms were fever (98%), headache (94%) and vomiting (67%). Neck stiffness was noted in 60%, and irritation in 46% of the patients. The median number of CSF cell count was 201/mm3 with polymorphonuclear predominance (>50%) in 58.3% of the cases. Enterovirus RNA was detected in CSF in 47 of 96 (48.9%) children tested. Children with positive enterovirus PCR had shorter hospitalization stay as compared to children who had negative PCR or to children who were not tested (P = 0.01). There were no serious complications or deaths.ConclusionsEnteroviruses accounted for approximately one half of cases of aseptic meningitis. PCR may reduce the length of hospitalization and plays important role in the diagnosis and management of children with aseptic meningitis.

The impact of viral co-infections and recently discovered viruses on the epidemiology of respiratory infections in children is still unclear. To simultaneously detect viruses that are involved in the aetiology of respiratory infections, we used a DNA/RNA microarray assay that identifies 17 different viruses or viral subtypes. Rhinopharyngeal washes were taken from 611 children (aged 1 month to 14 years) who presented in the emergency department with respiratory infections from June 2010 to June 2011 and were treated as outpatients (299, 48.9%) or hospitalized (312, 51.1%). Lower respiratory tract infection was diagnosed more often in hospitalized children (68% versus 36%, p 0.001). Of 397 children in which microarrays detected viral infection (70.1%), a single virus was found in 228 (57.4%) and two or more viruses in 169 (42.5%). The most prevalent viruses among children with positive samples were respiratory syncytial virus (RSV) in 225 (56.6%), parainfluenza virus (PIV) in 118 (29.7%), rhinovirus (RV) in 73 (18.4%), followed by influenza in 56 (14.1%), adenoviruses in 31 (7.8%), bocavirus in 25 (6.3%), human metapneumovirus in 15 (3.7%) and enteroviruses in 12 (3%). Most common viral co-infections were RSVA-RSVB in 46 children (27.2%), RSV-Influenza in 20 (11.8%), RSV-RV in 18 (10.6%) and PIV-RV in 13 (7.7%). Multiple logistic regression analysis revealed that viral co-infections were associated with increased probability for hospitalization (OR 1.52, 95% CI 1.01-2.29, p 0.04), and previous pneumococcal vaccination was associated with decreased probability for hospitalization (OR 0.52, 95% CI 0.33-0.81, p 0.004). We conclude that viral co-infections are involved in a significant proportion of children with an acute respiratory infection and may increase the severity of clinical presentation and the risk for hospitalization.

Streptolysin O (SLO) is a cholesterol-dependent cytolysin produced by the important human pathogen, group A Streptococcus (Streptococcus pyogenes or GAS). In addition to its cytolytic activity, SLO mediates the translocation of GAS NAD-glycohydrolase (NADase) into human epithelial cells in vitro. Production of both NADase and SLO is associated with augmented host cell injury beyond that produced by SLO alone, but the mechanism of enhanced cytotoxicity is not known. We have now shown that expression of NADase together with SLO dramatically enhanced the lytic activity of GAS culture supernatants for erythrocytes but had no effect on SLO-mediated poration of synthetic cholesterolrich liposomes. This result revealed a previously unknown contribution of NADase to the cytolytic activity associated with GAS production of SLO. Purified recombinant SLO bound NADase in vitro, supporting a specific, physical interaction of the two proteins. Exposure of human keratinocytes to wild-type GAS, but not to a NADase-deficient mutant strain, resulted in profound depletion of cellular NAD ؉ and ATP. Furthermore, expression of recombinant Group A Streptococcus (Streptococcus pyogenes or GAS), 2 the causative agent of streptococcal pharyngitis, skin and soft tissue infections, and severe invasive infections, produces more than 40 secreted proteins, many of which have been implicated in disease pathogenesis (1-3). Among the secreted products of GAS are streptolysin O (SLO) and NAD-glycohydrolase (NADase). SLO is a member of the pore-forming cytolysin family known as cholesterol-dependent cytolysins due to their ability to bind cholesterol in eukaryotic cell membranes (4, 5). Individual toxin molecules oligomerize and insert into the cell membrane to form transmembrane pores up to 30 nm in diameter. Sufficient doses of SLO result in cell lysis (4, 6, 7). Purified SLO is a potent toxin that is lethal within minutes of intravenous injection in animal models, primarily because of cardiotoxicity (8, 9). GASNAD-glycohydrolases are enzymes that catalyze the hydrolysis of the nicotinamide-ribose bond of NAD ϩ to yield nicotinamide and adenosine diphosphoribose. These enzymes have been described in several bacterial species and in a variety of eukaryotes (10, 11). The GAS NADase differs from most other known bacterial NAD-glycohydrolases in that it is secreted instead of being cell-associated and it possesses ADP ribosyl cyclase activity (12-14). Thus, in addition to producing adenosine diphosphoribose, the GAS NADase can convert NAD ϩ to cyclic adenosine diphosphoribose as a side reaction. Cyclic adenosine diphosphoribose is an important second messenger and signaling molecule for the regulation and mobilization of intracellular calcium stores (14 -16). The GAS genes encoding SLO (slo) and NADase (nga) are expressed from an operon that is transcribed under the control of a promoter upstream of nga. Some studies have identified downstream of nga a second weak promoter for slo (17, 18). The two proteins are functionally linked in that SLO is re...