Parylene Double-Layer Coated Screen-Printed Carbon Electrode for Label-Free and Reagentless Capacitive Aptasensing of Gliadin.

Celiac patients are required to strictly adhere to a gluten-free diet because even trace amounts of gluten can damage their small intestine and leading to serious complications. Despite increased awareness, gluten can still be present in products due to cross-contamination or hidden ingredients, making regular monitoring essential. With the goal of guaranteeing food safety for consuming labeled gluten-free products, a capacitive aptasensor was constructed to target gliadin, the main allergic gluten protein for celiac disease. The success of capacitive aptasensing was primarily realized by coating a Parylene double-layer (1000 nm Parylene C at the bottom with 400 nm Parylene AM on top) on the electrode surface to ensure both high insulation quality and abundant reactive amino functionalities. Under the optimal concentration of aptamer (5 µM) used for immobilization, a strong linear relationship exists between the amount of gliadin (0.01-1.0 mg/mL) and the corresponding ΔC response (total capacitance decrease during a 20 min monitoring period after sample introduction), with an R2 of 0.9843. The detection limit is 0.007 mg/mL (S/N > 5), equivalent to 0.014 mg/mL (14 ppm) of gluten content. Spike recovery tests identified this system is free from interferences in corn and cassava flour matrices. The analytical results of 24 commercial wheat flour samples correlated well with a gliadin ELISA assay (R2 = 0.9754). The proposed label-free and reagentless capacitive aptasensor offers advantages of simplicity, cost-effectiveness, ease of production, and speediness, making it a promising tool for verifying products labeled as gluten-free (gluten content <20 ppm).

Vapor Sublimation and Deposition to Fabricate a Porous Methyl Propiolate-Functionalized Poly-p-xylylene Material for Copper-Free Click Chemistry.

Conventional porous materials are mostly synthesized in solution-based methods involving solvents and initiators, and the functionalization of these porous materials usually requires additional and complex steps. In the current study, a methyl propiolate-functionalized porous poly-p-xylylene material was fabricated based on a unique vapor sublimation and deposition process. The process used a water solution and ice as the template with a customizable shape and dimensions, and the conventional chemical vapor deposition (CVD) polymerization of poly-p-xylylene on such an ice template formed a three-dimensional, porous poly-p-xylylene material with interconnected porous structures. More importantly, the functionality of methyl propiolate was well preserved by using methyl propiolate-substituted [2,2]-paracyclophane during the vapor deposition polymerization process and was installed in one step on the final porous poly-p-xylylene products. This functionality exhibited an intact structure and reactivity during the proposed vapor sublimation and deposition process and was proven to have no decomposition or side products after further characterization and conjugation experiments. The electron-withdrawing methyl propiolate group readily provided efficient alkynes as click azide-terminated molecules under copper-free and mild conditions at room temperature and in environmentally friendly solvents, such as water. The resulting methyl propiolate-functionalized porous poly-p-xylylene exhibited interface properties with clickable specific covalent attachment toward azide-terminated target molecules, which are widely available for drugs and biomolecules. The fabricated functional porous materials represent an advanced material featuring porous structures, a straightforward synthetic approach, and precise and controlled interface click chemistry, rendering long-term stability and efficacy to conjugate target functionalities that are expected to attract a variety of new applications.

Epidemiological features of Kawasaki disease in Taiwan, 1976-2007: results of five nationwide questionnaire hospital surveys.

BACKGROUND: Kawasaki disease (KD) affects mainly children younger than 5 years of age, leading to coronary artery lesions, and even to life-threatening myocardial infarction. In Taiwan, KD was encountered for the first time in 1976; then, it continued to occur in increasing numbers. METHODS: For the survey of epidemiological features of KD in Taiwan, we conducted five nationwide questionnaire hospital surveys in 1987, 1992, 1994, 2001, and 2008, respectively. In each survey, a special questionnaire form, together with a request letter and diagnostic guidelines for KD, was sent to the chairman of the Department of Pediatrics of all hospitals with 100 or more beds in Taiwan. RESULTS: KD patients increased to a total of 14,399 patients by 2007, with the highest number of 1018 in 2001, and the highest incidence of 66.24 per 100,000 children < 5 years of age in 2006. Of the 14,399 patients, the male-to-female ratio ranged from 1.5 to 1.7. Of these patients, 57.6-65.2% were < 2 years of age, 23.3-26.6% were 2-4 years of age, and 11.4-15.8% were ≥ 5 years of age. Coronary artery lesions were noted in 20.2-31.5% of patients. Fourteen cases expired, documenting that the fatality rate decreased from 0.4% to 0.03% during the 31 years from 1976 to 2007. CONCLUSION: In Taiwan, KD was encountered for the first time in 1976, and it continued to occur, reaching the highest annual incidence of 66.24 per 100,000 children < 5 years of age in 2006. In Taiwan, the first emergence of KD came in 1976; the annual increment of the incidence rate was lower (2.41 in Taiwan vs. 4.17 Japan), and no significant KD outbreak was observed in Taiwan.

Comparison of real-time polymerase chain reaction and serological tests for the confirmation of Mycoplasma pneumoniae infection in children with clinical diagnosis of atypical pneumonia.

BACKGROUND: Mycoplasma pneumoniae is a common pathogen of respiratory tract infection in children, and its correct and rapid diagnosis is a clinical challenge. Real-time polymerase chain reaction (RT-PCR) has been used frequently for the detection of this pathogen. MATERIALS AND METHODS: Medical records from all children with a clinical diagnosis of mycoplasma pneumonia and whose respiratory samples were tested for M. pneumoniae (using RT-PCR) during 2011 were reviewed retrospectively. We compared the sensitivity and specificity of serological assays versus those of RT-PCR for diagnosis of M. pneumoniae infections. We also reviewed retrospectively clinical characteristics, and laboratory and imaging findings of children with laboratory evidence of M. pneumoniae infection. RESULTS: In 2011, 290 children were diagnosed to have mycoplasma pneumonia clinically and had their respiratory samples tested for M. pneumoniae by RT-PCR. Fifty-four children (19%) had a positive result. Meanwhile, 63% (182/290) of these children also underwent serological tests, out of whom 44 (24%) were found to be positive for immunoglobulin M (IgM). Using PCR as a gold standard, M. pneumoniae IgM assay was found to show a sensitivity of 62.2% and a specificity of 85.5%. Positive and negative predictive values of IgM were 52.3% and 89.9%, respectively. In M. pneumoniae IgM-positive children, a negative PCR result was associated with more coinfection by other pathogens and longer duration of prehospitalization fever. Bacterial loads of M. pneumoniae were not correlated with clinical outcomes. CONCLUSION: The majority of clinically diagnosed mycoplasma pneumonia was unconfirmed. Mycoplasma pneumoniae IgM has poor sensitivity and a positive predictive value. Interpretation of Mycoplasma pneumoniae IgM should be done with caution.

Estimation of the incidence of Kawasaki disease in Taiwan. A comparison of two data sources: nationwide hospital survey and national health insurance claims.

BACKGROUND: Kawasaki disease (KD), first described by Dr. Tomisaku Kawasaki in 1967, was found for the first time in Taiwan in 1976. It continued to occur in increased numbers. For the study of incidence rates and epidemiological features of KD, we conducted five nationwide hospital surveys (NHS) in 1987, 1992, 1994, 2001 and 2008, respectively. We estimated also the annual incidence rates of KD during 1996-2007, based on the National Health Insurance (NHI) database, which had been implemented since 1995, covering 98% of the population in Taiwan. METHODS: The annual incidence rates of KD during the twelve years, from1996 to 2007, estimated by the NHS and the NHI claims were compared, analyzed and discussed. RESULTS: During 1996-2007, a total of 9,938 cases of KD were reported by the Departments of Pediatrics of all hospitals surveyed, and a total of 11,849 cases of KD were claimed in the NHI database. The annual number of cases and incidence rates of KD based on NHI claims constantly surpassed those by the NHS. The ratio of the two incidence rates varied from 1.10 to 1.33. They were well correlated (r = 0.902, p < 0.001) with a linear equation, NHI = 16.07 + 0.93*NHS. The changes in annual incidence rate by the NHI were mean 1.149, p = 0.07, 95% CI -0.082 - 2.382, and those by the NHS were mean 1.562, p <0.001, CI 0.656 - 2.468. CONCLUSION: The annual incidence rates of KD can be estimated by the NHI claims and by the classic NHS. The values estimated by the NHI claims constantly outnumbered those by the NHS. Some pitfalls involved in the NHI claims are discussed.

Recommendations for rotavirus vaccine.

Rotavirus infection has been the leading cause of gastroenteritis among children in Taiwan. Studies have shown that 40% of hospitalization for acute gastroenteritis can be prevented through the use of vaccines, including a live, attenuated monovalent rotavirus vaccine and a pentavalent, human-bovine reassortant rotavirus vaccine. In 2009, the World Health Organization suggested that rotavirus vaccine should be included in all national immunization programs. This review summarizes issues and recommendations discussed during an expert meeting in Taiwan. The recommendations included: (1) rotavirus vaccine should be offered to all healthy infants (including those without contraindications, such as immunodeficiency) at an appropriate age; (2) either monovalent or pentavalent vaccine can be administered concurrently with routine injected vaccines; (3) the administration of rotavirus vaccine must be administered at least 2 weeks prior to oral polio vaccination; (4) the first vaccine dose for infants should be administered between age 6 weeks and age 14 weeks 6 days and the course should be completed by age 8 months 0 day; (5) pentavalent vaccines can be administered at 2 months, 4 months, and 6 months while monovalent vaccines can be taken at 2 months and 4 months; (6) a combined use of monovalent and pentavalent vaccine is justified only when the previous dose is unavailable or unknown; and (7) rotavirus vaccines may be given to premature infants, human immunodeficiency virus infected infants and infants who have received or are going to receive blood products.

Waning population immunity to measles in Taiwan.

To evaluate the population immunity to measles in Taiwan where the coverage rate of the measles vaccine was >95% for more than a decade, anti-measles IgG was determined in 3552 Taiwanese volunteers in 2007. The overall seroprevalence was 74.7% (95% confidence interval [CI]: 73.3-76.1%). In subgroups aged 2-25 years, to whom at least 2 doses of measles-containing vaccine were given, there was a declining trend of seropositivity with age from 94.5% at 2 years to 50.6% at 21-25 years (p<0.0001). Age (odds ratio [OR]: 1.0464, 95% CI: 1.043-1.085) and male gender (OR: 1.466, 95% CI: 1.131-1.901) were independent factors predicting seronegative sera in this population. Seroprevalence was uniformly >95% in the older population (≥ 35 years) who had not been immunized against measles. The waning vaccine-induced immunity may have impact on the control of measles in the future, especially when the vaccinated population becomes older.

Viral load and clinical features in children infected with seasonal influenza B in 2006/2007.

BACKGROUND/PURPOSE: In influenza B infection, viral load is believed to be related to the severity of clinical illness. The correlation between viral load and symptoms is not known. We conducted a study to assess the relationship between virus load and clinical features in children infected with influenza B, in the hope that clinical features could be used as surrogate markers of viral load to guide treatment. METHODS: Between December 2006 and February 2007, 228 patients with fever and respiratory symptoms were prospectively enrolled in our tertiary hospital-based study. Real-time reverse transcription polymerase chain reaction (RT-PCR) was performed to determine viral load. RESULTS: Real-time RT-PCR was positive for influenza B in 76 patients. Using virus culture as the gold standard, the sensitivity and specificity were 95% and 87%, respectively. Influenza culture positive rate significantly correlated with viral load (p = 0.03). The median copy number of influenza B virus in the 76 RT-PCR positive patients was 9735 copies/ml (range 4.8×10¹-2.0×106 copies/ml). Samples obtained later in the clinical course tended to have lower viral load (p = 0.7), while patient age (p = 0.72) and fever duration (p = 0.96) positively related to viral load. In patients >3 years of age, myalgia was related to statistically lower viral loads (14300 vs. 1180; p = 0.025). Patients with chills tended to have higher viral loads (72450 vs. 7640; p = 0.1). Patients with abdominal pain tended to have lower viral loads (1998 vs. 12550; p = 0.06). CONCLUSION: Culture rate positively correlated with viral load. Patients with myalgia had a lower viral load.

Are empiric antibiotics for acute exudative tonsillitis needed in children?

BACKGROUND: Empiric antibiotics are frequently given for children with acute exudative tonsillitis. A few studies have investigated the causative agent of acute "exudative" tonsillitis in children to evaluate the necessity of antibiotic therapy. This study tried to explore the common causative agent of acute exudative tonsillitis among children. METHODS: From April 2009 to March 2010, throat swabs were obtained and cultured for viruses and bacteria from children who visited the pediatric emergency rooms of two medical centers in central Taiwan with acute exudative tonsillitis. Demographic data and microbiological results were analyzed. RESULTS: A total of 294 children with acute exudative tonsillitis were enrolled during the 1-year prospective study, and 173 (58.8%) of them were younger than 7 years. Group A streptococci were isolated from only three (1.0%) children, and they were all older than 6 years. A total of 143 viruses were isolated from 140 (47.6%) children. Adenovirus (18.7%) and enterovirus (16.3%) were the most common viral etiologies, followed by influenza virus (5.4%), parainfluenza virus (5.1%), herpes simplex virus Type 1 (2.7%), and respiratory syncytial virus (0.3%). Group A streptococcus only contributed to a minimal portion of acute exudative tonsillitis. CONCLUSION: Routine or immediate antibiotic therapy for acute exudative tonsillitis in children is not necessary.

Serologic status for pandemic (H1N1) 2009 virus, Taiwan.

We studied preexisting immunity to pandemic (H1N1) 2009 virus in persons in Taiwan. A total of 18 (36%) of 50 elderly adults in Taiwan born before 1935 had protective antibodies against currently circulating pandemic (H1N1) 2009 virus. Seasonal influenza vaccines induced antibodies that did not protect against pandemic (H1N1) 2009 virus.

Epidemic pleurodynia caused by coxsackievirus B3 at a medical center in northern Taiwan.

Epidemic pleurodynia is seldom reported in Southeast Asia and there has been no report from Taiwan. We conducted a retrospective chart review of children = 18 years of age in the National Taiwan University Hospital from January 1 to December 31, 2005. Epidemic pleurodynia was defined as an acute illness characterized by sharp localized pain over the chest or upper abdomen. Patients with known heart diseases or pulmonary consolidations were excluded. In total, 28 patients met the case definition of epidemic pleurodynia. Coxsackievirus B3 (CB3) was isolated in 15 (60%) of the 25 throat swab specimens. Four (14%) of the 28 patients presented chest wall tenderness and only one (6%) of the 18 patients tested had an elevated creatinine kinase level. Twenty-one (75%) of the 28 patients described pleuritic chest pains and 10 (45%) of the 22 chest radiographies exhibited pulmonary infiltrates or pleural effusions. Six patients were observed with tonsillar exudates and one was confirmed to have a CB3 urinary tract infection. The clinical features and radiological findings suggest that CB3-associated epidemic pleurodynia might be a disease of the pleura and occasionally spreads to nearby tissues, resulting in chest wall myositis, pulmonary infiltrates and myopericarditis.

Epidemiology and treatment of Bell's palsy in children in northern Taiwan.

BACKGROUND AND PURPOSE: Bell's palsy is not uncommon in children. This study was performed to evaluate the epidemiology of Bell's palsy in the northern Taiwanese pediatric population, and the effectiveness of corticosteroid treatment. METHODS: The medical records of pediatric patients with a primary diagnosis of facial palsy from April 2002 through March 2007 were reviewed. Patients with secondary facial palsy were excluded from the analysis. RESULTS: 289 episodes of facial palsy were identified and the clinical findings of 134 episodes among 132 patients were assessed. The median +/- standard deviation age was 9.9 +/- 4.9 years, and 58.2% of patients were girls. Children were more likely to have episodes of Bell's palsy during the cold season, with a peak in January. The left (67 episodes; 50.0%) and right (64 episodes; 47.8%) facial nerves were involved with similar frequency. Common symptoms were postauricular pain (11.2%) and facial hypoesthesia (9.0%). Of 51 episodes of Bell's palsy with complete follow-up, corticosteroids were given for 44 episodes. Thirty eight patients (86.4%) given corticosteroids had complete recovery and 4 patients (57.1%) recovered without corticosteroids. Rates of complete recovery did not differ significantly between the 2 groups (p = 0.08). There were no significant differences in the recovery rate between early (< or = 3 days) and late (4-7 days) administration. CONCLUSIONS: In northern Taiwan, childhood Bell's palsy peaks from January through March. The majority of children with Bell's palsy recovered completely. There was no significant effect of corticosteroid treatment for children with Bell's palsy.

Safety and immunogenicity of a tetravalent polysaccharide vaccine against meningococcal disease.

BACKGROUND/PURPOSE: Meningococcal disease, including meningitis and sepsis, usually follows an invasive infection by Neisseria meningitidis, and is a major cause of death and morbidity worldwide. Currently available meningococcal vaccines that consist of pure capsular polysaccharides (serogroups A, C, W-135 and Y) are generally safe and efficacious in adults and children aged > 2 years. The purpose of this study was to evaluate the immunogenicity, safety and reactogenicity of a single dose of GlaxoSmithKline Biologicals' Mencevax ACWY vaccine in healthy Taiwanese subjects aged 2-30 years. METHODS: This open, single center, Phase III study was conducted in Taiwan. A single dose of the Mencevax ACWY vaccine was administered to subjects aged 2-30 years. Immunogenicity and safety of the vaccine were evaluated after vaccination. RESULTS: The immunogenicity results obtained 1 month after vaccination with Mencevax ACWY vaccine indicated that the vaccine elicited a good immune response in vaccinees aged 2-30 years. This was both in terms of functional activity directed against meningococcal polysaccharide (A, C, W-135 and Y) as measured by serum bactericidal assay (> 93% activity against all serogroups), and specific IgG concentrations measured by ELISA (> 96% seropositivity to all serogroups). CONCLUSION: The tetravalent polysaccharide Mencevax ACWY meningococcal vaccine was well tolerated and immunogenic in subjects aged 2-30 years in Taiwan.

Coxsackieviruses infection in northern Taiwan--epidemiology and clinical characteristics.

BACKGROUND AND PURPOSE: The epidemiology of coxsackievirus has rarely been investigated in Taiwan. This study was performed to ascertain the epidemiological and clinical characteristics of coxsackievirus infections in Taiwan. METHODS: 457 patients treated at a medical center in northern Taiwan who were positive for coxsackievirus were enrolled in this retrospective study. Patients' medical charts were reviewed for clinical diagnosis, physical examination, laboratory findings, and clinical manifestations. RESULTS: Three serotypes of coxsackievirus A (A9, 5.3%; A10, 7.2%; A16, 87.5%) were identified among 265 patients, 27.4% of whom were admitted to hospital. The mean (+/- standard deviation [SD]) duration of fever and hospital stay was 2.6 +/- 0.5 days and 4.0 +/- 2.1 days, respectively. Complications were noted in 14 patients (5.3%), all of which involved the central nervous system (CNS). All 6 serotypes of coxsackievirus B (B1, 2.6%; B2, 7.8%; B3, 55.7%; B4, 2.1%; B5, 12.5%; B6, 1.0%; non-typable, 18.2%) were identified in 192 patients, 45.3% of whom were admitted to hospital. The mean (+/- SD) duration of fever and hospital stay was 4.1 +/- 1.0 and 3.4 +/- 0.9 days, respectively. Thirty seven patients (19.3%) had complications, including 34 with CNS involvement. Patients with coxsackievirus B infection had higher hospital admission rates (p < 0.001), more CNS involvement (p < 0.001), and longer fever duration (p < 0.001) than those with coxsackievirus A infection. Patients with coxsackievirus A infection tended to have more skin manifestations (p < 0.001) and oral ulcers (p < 0.001). CONCLUSIONS: The most common serotypes were coxsackieviruses A16 and B3. Patients with coxsackievirus B infection were more likely to be admitted to hospital, had longer fever duration, and more CNS involvement than patients with coxsackievirus A infection.

Microbiology and epidemiology of brain abscess and subdural empyema in a medical center: a 10-year experience.

BACKGROUND AND PURPOSE: Brain abscess and subdural empyema are the 2 most common forms of intracranial pyogenic infections. Predisposing factors and etiological agents may change with time. This study examined the epidemiological features of these conditions. METHODS: The medical records of all inpatients with a diagnosis of brain abscess or subdural empyema from 1998 to 2007 were reviewed. The diagnosis was confirmed by imaging study or operative findings. RESULTS: 151 episodes of brain abscess were diagnosed in 150 patients, and 10 patients had subdural empyema. The incidence of brain abscess fluctuated over time, while that of subdural empyema remained stable. The mean +/- standard deviation age of patients with brain abscess was significantly greater than that of patients with subdural empyema (48.5 +/- 19 years vs 25.4 +/- 24 years; p = 0.004). The number of patients with hematogenous brain abscess increased from 7 in 1998 to 2002 to 19 in 2003 to 2007, while that of those with infection related to operation decreased from 10 to 5. Most subdural empyema was related to bacterial meningitis (4 of 10). Etiological agents were identified in 53% of brain abscesses, including Enterobacteriaceae spp. (21.3%), Streptococcus spp. (20%), and mixed pathogens (17.5%). Klebsiella pneumoniae was the most common enteric bacteria isolated (15.3%), especially in patients with diabetes mellitus, but was not observed in children younger than 18 years. CONCLUSIONS: In contrast to western countries, K. pneumoniae plays an important role in intracranial pyogenic infections in Taiwan. The pathogens and routes of infection are different between children and adults.

Clinical characteristics of nosocomial rotavirus infection in children in Taiwan.

BACKGROUND/PURPOSE: The aim of this study was to determine the incidence and clinical characteristics of nosocomial rotavirus infection (NRI) among hospitalized children. METHODS: We collected data of children in the Department of Pediatrics with positive stool rotavirus antigen tests. Cases of an admission diagnosis of acute gastroenteritis or a positive stool rotavirus antigen test within 3 days of admission, representing community-acquired infections, were excluded. Both VP4 and VP7 genotyping of the rotaviruses was done. RESULTS: There were 98 patients who met the inclusion criteria during the 3-year period. The incidence density was 0.58 per 1000 patient-days in our series. Among these patients, 59 (60%) had underlying diseases. The intermediate intensive care unit had the highest incidence density (2.8 per 1000 patient-days). Overcrowding of the care unit, inappropriate hand hygiene, and inadequate isolation and cohorting predisposed to the high rate. Genotypes among 79 (80%) rotaviruses tested showed that 42% belonged to the novel genotype, G9P[8]. CONCLUSION: NRI may cause significant morbidity in hospitalized children, especially young infants and those with underlying diseases. Infection control with hospital surveillance, strict isolation and cohort care should be adopted to prevent the spread of rotavirus among special care units.

Humoral and cellular immune responses to a hepatitis B vaccine booster 15-18 years after neonatal immunization.

BACKGROUND: Whether hepatitis B (HB) vaccine-conferred immunity persists into adulthood is unknown. We aimed to investigate long-term HB immunity in adolescents. METHODS: In 2004-2005, 6156 high school students (15-21 years old) who had been vaccinated with plasma-derived HB vaccine as infants were recruited for HB seromarker screening. The immune response to an HB vaccine booster was evaluated in 872 subjects who were seronegative. HB surface antibody (anti-HBs) titers and levels of HB surface antigen (HBsAg)-specific interferon (IFN)-gamma- or interleukin (IL)-5-secreting peripheral blood mononuclear cells (PBMCs; measured by enzyme-linked immunospot assay) were determined 4 weeks later. RESULTS: Although the vaccine remained highly efficacious in reducing the HBsAg positivity rate, 63.0% of the vaccinees had no protective anti-HBs. After the booster, anti-HBs remained undetectable in 28.7% (158/551) of the subjects who had received complete HB vaccination (4 doses) during infancy. We estimated that 10.1% of the total population had lost their HB vaccine-conferred booster response. HBsAg-specific IFN-gamma- or IL-5-secreting PBMCs remained negative in 27.2% (25/92) of subjects after the booster. CONCLUSIONS: A notable proportion of fully vaccinated adolescents had lost immune memory conferred by a plasma-derived HB vaccine 15-18 years later. This decay of immune memory may raise concerns about the need for a booster vaccine for high-risk groups in the long run.

Astrovirus gastroenteritis in children in Taipei.

BACKGROUND/PURPOSE: A prospective study was initiated to study astroviral infections in Taiwan. METHODS: A total of 415 stool samples were collected and assayed for astrovirus antigen using an enzyme immunoassay. RESULTS: Twelve (2.9%) stool samples from 12 patients were positive for astrovirus antigen. Most patients (8/12) had watery diarrhea which lasted for 2-6 days. The majority of patients recovered without specific treatment, except for two patients who were treated with antibiotics for possible bacterial infections. One patient developed chronic diarrhea and two episodes were nosocomially acquired. A clustering in the autumn and winter, with a peak in December (5/12), was noted. Growth on Caco-2 cells was performed for four specimens with positive astroviral RT-PCR results, and a characteristic cytopathic effect was observed after 4 days. Astroviral RNA was detected in six stool samples using RT-PCR. Five of six strains were serotype 1 and one strain was serotype 3. Sequence homology among the six strains was between 80.5% and 100%. A higher degree of homology (89.9-100%) was noted in the five strains of serotype 1. A phylogenetic study demonstrated two clusters in our strains and Oxford reference strain types 1 and 2. CONCLUSION: Our study results provide further information about the prevalence and span of clinical spectra associated with astroviral infections in Taiwan. The current study showed that infection with astroviruses may be an important cause of gastroenteritis, as well as respiratory symptoms, in infants and children in Taipei.

Establishment of a young mouse model and identification of an allelic variation of zmpB in complicated pneumonia caused by Streptococcus pneumoniae.

OBJECTIVE: Complicated pneumonia, including necrotizing pneumonia, lung abscess, and empyema, caused by Streptococcus pneumoniae in children has been increasing. We thus determined to investigate its virulence in an animal model and to identify virulence factors of S. pneumoniae. DESIGN: Prospective, randomized, controlled animal study. SETTING: University medical laboratory. SUBJECTS: Male Balb/c-strain mice, 3 wks old. INTERVENTIONS: We used a young mouse model to monitor bacterial virulence and a microarray to compare gene expression between S. pneumoniae from children with complicated and noncomplicated pneumonia. Deletion and complementation of a candidate gene were performed to study its role on the virulence of S. pneumoniae. MEASUREMENTS AND MAIN RESULTS: A model of complicated pneumonia in young mice infected with strains of S. pneumoniae from children with complicated pneumonia was established. Using a microarray analysis, differences in zinc metalloprotease B (zmpB) RNA hybridization between two strains from children with complicated pneumonia (NTUH-p28 and NTUH-p15) and a strain (NTUH-p3) from a child with pneumococcal lobar pneumonia were found. Confirmatory assays revealed the signal differences were due to sequence variation in the zmpB gene. Infection with the zmpB deletion mutant of NTUH-p15 showed a significant decrease in the severity of pneumonia and no destructive lung injury. The zmpB complementation strain of NTUH-p15 significantly restored the level of inflammation and caused lung necrosis. For studying the effect of allelic variation of zmpB on the virulence of S. pneumoniae, we added zmpB of NTUH-p15 in the zmpB deletion mutant of NTUH-p3, which resulted in a higher bacterial burden than that in wild-type NTUH-p3. CONCLUSIONS: A young mouse model is established for complicated pneumococcal pneumonia. This model proved that allelic variation of zmpB affects the virulence of S. pneumoniae.