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In addition to the typical respiratory response, new coronavirus disease 2019 (COVID-19) is also associated with very common gastrointestinal symptoms. Cases with gastrointestinal symptoms are more likely to be complicated by liver injury and acute respiratory distress syndrome (ARDS). If not treated in time, coma and circulatory failure may ensue. As severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infects the human body through the combination of angiotensin-converting enzyme 2 (ACE2) in the gastrointestinal tract, the mechanism underlying the gastrointestinal symptoms may involve damage to the intestinal mucosal barrier and promotion of the production of inflammatory factors. Indeed, after cells in the lungs become infected by SARS-CoV-2, effector CD4+ T cells reach the small intestine through the gut-lung axis, causing intestinal immune damage and diarrhea; early extensive use of antibacterial and antiviral drugs can also lead to diarrhea in patients. Thus, treatment options for COVID-19 patients should be promptly adjusted when they have gastrointestinal symptoms. As SARS-CoV-2 has been detected in the feces of COVID-19 patients, future prevention and control efforts must consider the possibility of fecal-oral transmission of the virus.

Background Our previous study found that most Mycoplasma pneumoniae (MP) pneumonia (MPP)patients had elevated serum total immunoglobulin E (IgE) levels. Objective To determine components of MP that can cause an IgE increase in children, and to clarify its specific mechanism. Methods The components of MP cells were isolated by serum IgE from patients with MP pneumonia. These components obtained through the prokaryotic expression were used as allergens to detect the proportion of allergen-specific IgE produced in MPP patients, and the clinical characteristics and related immune parameters of these patients who produced this allergen-specific IgE were also analyzed. In addition, a cell experiment was used to verify the biological effect of these components in vitro. Results P1-specific IgE was detected in serum of MPP children. An approximately 24-kDa polypeptide of P1 protein was obtained through prokaryotic expression purified by nickel agarose affinity chromatography. Approximately 9.2% of MPP patients produced IgE against this polypeptide of P1 protein, which was more likely to be produced in MPP patients with no history of allergies or family history of allergy-related diseases. P1-specific IgE-positive MPP patients had more severe clinical symptoms, with excessive secretion of interleukin (IL)-4 and IL-5 and overdifferentiation of Th0 cells into Th2 cells. Tests also demonstrated that the P1 protein stimulated excessive secretion of IL-4 and IL-5 in peripheral blood mononuclear cells from the peripheral blood of healthy donors. Conclusion Mycoplasma pneumoniae is not only an infectious agent but also an allergen for certain individuals. The P1 protein of MP can induce the production of P1-specific IgE.

Sepsis is an important cause of neonatal morbidity and mortality worldwide. Diagnosis and treatment of neonatal sepsis relies on clinical judgment and interpretation of nonspecific laboratory tests. In a prospective cohort, we measured inflammatory cytokines as a potential biomarker for neonatal sepsis. Serum inflammatory cytokine levels were evaluated in the early stage of neonatal sepsis and after antimicrobial treatment. Receiver operating characteristic curves assessed the diagnostic value of cytokines. We performed multiple logistic regression analysis to characterize the role of each cytokine independently for infants with culture proven sepsis. C-reactive protein, interleukin (IL)-6, IL-10 and IL-6/IL-10 levels were significantly elevated in neonatal sepsis when compared with the control group and there were 1.4 (95% confidence interval (CI): 1.2–1.5), 4.9 (95% CI: 4.6–5.1), 5.1 (95% CI: 4.5–5.6), and 10.2 (95% CI: 9.2–11.1) fold greater odds, respectively, to predict neonatal sepsis when increased. After effective treatment, median IL-6 (pretreatment value: 263.0 pg/ml and post-treatment value: 7.4 pg/ml) and IL-6/IL-10 levels (pretreatment value: 16.6 and post-treatment value: 1.4) significantly decreased. The areas under the curve for IL-6, IL-10, IL-6/IL-10 and C-reactive protein for differential diagnosis were 0.98, 0.82, 0.90, and 0.88, respectively. IL-6 and IL-6/IL-10 outperformed C-reactive protein to diagnose neonatal sepsis. Of the cytokines studied, IL-6 was the most sensitive, whereas IL-6/IL-10 was the most specific predictor of neonatal sepsis.

This study investigated whether the rotavirus infection rate in children is associated with temperature and air pollutants in Hangzhou, China. This study applied a distributed lag non-linear model (DLNM) to assess the effects of daily meteorological data and air pollutants on the rotavirus positive rate among outpatient children. There was a negative correlation between temperature and the rotavirus infection rate. The impact of temperature on the detection rate of rotavirus presented an evident lag effect, the temperature change shows the greatest impact on the detection rate of rotavirus approximate at lag one day, and the maximum relative risk (RR) was approximately 1.3. In 2015, the maximum cumulative RR due to the cumulative effect caused by the temperature drop was 2.5. Particulate matter (PM) 2.5 and PM10 were the primary air pollutants in Hangzhou. The highest RR of rotavirus infection occurred at lag 1–1.5 days after the increase in the concentration of these pollutants, and the RR increased gradually with the increase in concentration. Based on the average concentrations of PM2.5 of 53.9 μg/m 3 and PM10 of 80.6 μg/m 3 in Hangzhou in 2015, the cumulative RR caused by the cumulative effect was 2.5 and 2.2, respectively. The current study suggests that temperature is an important factor impacting the rotavirus infection rate of children in Hangzhou. Air pollutants significantly increased the risk of rotavirus infection, and dosage, lag and cumulative effects were observed.

This study investigated whether respiratory syncytial virus (RSV) infection in children was associated with ambient temperature and air pollutants in Hangzhou, China. A distributed lag non-linear model (DLNM) was used to estimate the effects of daily meteorological data and air pollutants on the incidence of RSV infection among children. A total of 3650 childhood RSV infection cases were included in the study. The highest air pollutant concentrations were in January to May and October to December during the year. The yearly RSV-positive rate was 10.0 % among children with an average age of 4.3 months. The highest RSV-positive rate occurred among patients 0 to 3 months old. Children under 6.5 months old accounted for 80 % of the total patients infected by RSV. A negative correlation was found between ambient temperature and RSV infection, and it was strongest with minimum ambient temperature (r = −0.804, P

Nontypeable Haemophilus influenzae (NTHi) is one of the most common etiologies of acute otitis media, rhinosinusitis, and pneumonia. Outer membrane proteins (OMPs) are the main focus in new vaccine development against NTHi, as the H. influenzae type b (Hib) vaccine does not cover noncapsulated NTHi. The OMPs P6 and protein D are the most promising candidate antigens for an NTHi vaccine, and low antibody levels against them in serum may be correlated with infection caused by NTHi. In the current study, we measured the antibody titers against P6, protein D, and their T- and B-cell combined peptide epitopes in healthy individuals of different ages. We found that children H. influenzae .

Resumen Introduccion El asma es una afeccion inflamatoria de las vias respiratorias. Las infecciones por Mycoplasma pneumoniae pueden exacerbar los sintomas del asma. Se ha demostrado que la interleucina 2 y la interleucina 4 participan en las reacciones inmunitarias e inflamatorias. Hemos estudiado la relacion entre los polimorfismos de la IL2 y la IL4 y su expresion y el riesgo de padecer asma e infeccion por M. pneumoniae en ninos. Metodos Se recluto a 392 ninos asmaticos y 849 controles para el estudio. Se genotiparon 8 polimorfismos en IL2 e IL4 con la plataforma MassARRAY de Sequenom. La infeccion por M. pneumoniae y el numero de copias se establecieron mediante PCR fluorescente. Los niveles sericos de expresion de IL-2 e IL-4 se midieron con ELISA. Resultados Hallamos una relacion significativa entre el polimorfismo rs6534349 de IL2 y el aumento de riesgo de sufrir asma (heterocigoticos, p = 0,029; variantes homocigoticas, p = 0,013), asi como entre el polimorfismo rs2227284 de IL4 y una reduccion del riesgo de padecer asma (heterocigoticos, p = 0,026; variantes homocigoticas, p = 0,001). Ademas, la relacion con otros polimorfismos, excepto el rs2070874, se hizo evidente al agrupar a los ninos asmaticos segun la clasificacion GINA de control y gravedad del asma. Asimismo, los niveles sericos de expresion de IL-2 e IL-4 fueron significativamente mayores en los sujetos no infectados (p = 0,038) e infectados (p = 0,011) por M. pneumoniae, respectivamente. Esta observacion tambien se cumple entre los pacientes asmaticos (p = 0,016 para IL-2 y p = 0,042 para IL-4), pero en los controles no asmaticos solo se cumple en el caso de la IL-4 (p = 0,032). Del mismo modo, observamos que el genotipo GG rs6534349 estaba claramente relacionado con un aumento de las posibilidades de tener una infeccion con alta carga de M. pneumoniae (p = 0,0376). Conclusiones La IL2 y la IL4 podrian ser biomarcadores importantes para calcular el riesgo de padecer asma, asi como infeccion por M. pneumoniae, en ninos.

Objective Kawasaki disease (KD) is the primary cause of heart disease among children, but because its clinical symptoms are nonspecific, it is difficult to diagnose. The purpose of this study was to evaluate laboratory indices for possible use in the early diagnosis of KD and to determine which indices are predictive of a response to intravenous immunoglobulin (IVIG) and can be used to monitor the effects of treatment. Methods Three hundred thirty KD patients, 330 age-matched children with KD-like febrile disease, and 330 age-matched healthy children (controls) were enrolled in this prospective study. Levels of N-terminal pro–brain natriuretic peptide (NT-proBNP), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and cytokines were determined in all study subjects. Results In the derivation cohort, 181 patients in the KD group were compared with 181 patients in the KD-like febrile group. The following indices were found to be useful in the diagnosis of KD: NT-proBNP (area under the curve [AUC] 0.923), ESR (AUC 0.909), CRP (AUC 0.834), and interleukin-6 (IL-6; AUC 0.678). The diagnostic efficiency of each index demonstrated in the derivation cohort was repeated in the 149 KD patients in the validation cohort. There were significant differences in NT-proBNP levels between IVIG-responsive KD patients (n = 270) and IVIG-nonresponsive KD patients (n = 60), with higher NT-proBNP levels in IVIG-nonresponsive KD patients. The NT-proBNP level can effectively distinguish IVIG-responsive KD patients from IVIG-nonresponsive patients, and its AUC was 0.73. There were also significant differences in the NT-proBNP levels before and after treatment, with a significant decline after treatment. Conclusion Serum levels of NT-proBNP can be used in the diagnosis of KD, the prediction of a patient's sensitivity to IVIG treatment, and the monitoring of the effects of IVIG treatment, but more attention must be paid to the scope of its application.

Autophagy is a dynamic process accomplished by the generation and maturation of autophagosomes. Isolation of autophagosomes and subsequent compositional analysis can provide information about their biogenesis mechanism. In this article, HEK293 cells expressing GFP-LC3 were treated by calcium phosphate precipitates (CPP) to induce autophagy. The autophaogomes induced by CPP were tubular and vesicular structures, extensively formed in the cytosol. After all membranes in the cell lysate were fractionated by differential centrifugation, autophagosomes from light and heavy membranes were isolated by immuno-precipitation, using antibodies against GFP-LC3 and Atg5. We found that GFP-LC3 and Atg5 positive autophagosomes represented distinctive subpopulations. Judged from the molecular markers associated, including organelle markers and Atg proteins, GFP-LC3 positive autophagosomes were overall at the later biogenetic stage. Furthermore, both GFP-LC3 and Atg5 positive autophagosomes from light membranes were less mature than those from heavy membranes. We have established a method to isolate subpopulations of autophagsomes for further characterization.

There is growing public concern regarding assay sensitivity to HBsAg mutants in clinical diagnosis and vaccine escape. The aim of this study is to introduce a new HBsAg mutant strain. The serum samples were those of patient X at the age of 3 months and 3 years respectively, and of her mother immediately before parturition, which were used to amplify the HBsAg-coding DNA fragments by PCR. The HBsAg DNA sequences were translated into their corresponding amino acid sequences and then aligned in pubmed with nucleotide blast. The sequencing data of S coding regions shows that patient X has been infected by a new HBV variant with an A to C substitution at nt431, resulting in an Asp(GAC)to Ala(GCC) substitution at aa144 of major protein; CC to AA substitution at nt359 and nt360, resulting in an Pro(CCC) to Gln(CAA) substitution at aa120 of pre “a” epitope; A to G substitution at nt491, resulting in an Glu(GAG) to Gly(GGG) substitution at aa164 of post “a” epitope. Three new mutations (S171F, S174N and Q181R) at the antigenic epitopes of HBV presented by HLA class I molecules are found. The HBV mutant strain causes vaccine escape and occult infection.

To study sialic acid and iron content in breastmilk in Chinese women during different lactation stages. Sialic acid and iron content of colostrum, transitional milk, mature milk, and involutional milk were determined using a neuraminidase assay kit and the ferrozine method, respectively in 88 lactating women (58 Term, 30 Preterm). The mean (SD) sialic acid levels of colostrum, transitional milk, mature milk, and involutional milk were 2201.4 (676.6) mg/L, 1445.9 (423.4) mg/L, 395.3 (96.0) mg/L and 273.0 (76.9) mg/L, respectively. The median iron content were 0.05 mg/L, 0.06 mg/L, 0.25 mg/L and 0.35 mg/L, respectively, in successive stages of lactation. Sialic acid and iron were significantly higher in breast milk of preterm mothers compared to term mothers. Sialic acid and iron content in breast milk vary greatly throughout the lactation stages, which probably reflects the infants’ needs for growth and development at different stages.

Respiratory Syncytial Virus (RSV) infections are the dominant cause of pneumonia in children. In order to determine the epidemiological characteristics and immune status of children with Respiratory Syncytial Virus, a prospective study was performed among patients with RSV infection. Comparisons between RSV pneumonia group and normal control group, RSV pneumonia group had lower IL-2 (median levels, pg/ml: 3.8 vs. 5.1, P

Kawasaki disease (KD) has become the most common cause of acquired heart disease in children and is also a risk factor for ischemic heart disease in adults. However, Kawasaki disease lacks specific laboratory diagnostic indices. Thus, this study analyzed the T cell activation profiles of Kawasaki disease and assessed their value in the diagnosis of Kawasaki disease and the prediction of intravenous immunoglobulin (IVIG) sensitivity. We analyzed human leukocyte antigen-DR (HLA-DR), CD69 and CD25 expression on peripheral blood CD4+ and CD8+ T cells during the acute phase of KD. We compared the percentages of HLA-DR+/CD69+/CD25+ T cells in the CD4+ and CD8+ T cell populations of IVIG-effective and IVIG-resistant groups. Receiver operating characteristic curves were used to assess the diagnostic value of the above parameters. The median percentage of CD8+HLA-DR+ T cells and the median ratio of CD8+HLA-DR+ T cells/CD8+CD25+ T cells were significantly elevated in the patient group compared with those in the control group during the acute phase of KD. Regarding the diagnosis of Kawasaki disease, the area under the ROC curve was 0.939 for the percentage of CD8+HLA-DR+ T cells. There was a significant difference in the ratio of CD8+HLA-DR+ T cells/CD8+CD69+ T cells between IVIG-resistant patients and IVIG-sensitive patients. Regarding IVIG sensitivity, the area under the ROC curve was 0.795 for it. Excessive CD8+ T cell activation, as well as an imbalance between CD8+ T cell activation and inhibition, underlies the pathogenesis of Kawasaki disease. The percentage of CD8+ HLA-DR+ T cells may be used as an index to diagnose Kawasaki disease. IVIG inhibits CD8+ T cell activation, but excessive CD8+ T cell activation may cause IVIG resistance. The ratio of CD8+HLA-DR+ T cells/CD8+CD69+ T cells may be used as a predictor of IVIG sensitivity.

Congenital heart disease (CHD) is extremely rarely reported in 48, XYY, +21 karyotype. Herein, we reported one case of 48,XYY,+21 karyotype with CHD and reviewed the available literature. The phenotypic characteristics of the 4-month-old child showed the presence of features typical of mongoloid slant. X-ray detection showed the form of heart was corpulent and the bilateral mediastinum was broad. Doppler echocardiogram detection showed atrial septal and ventricular septal defects with patent ductus arteriosus, pulmonary hypertension and mild tricuspid regurgitation. Including this case, 63 cases of 48, XYY, +21 chromosome pattern have been reported. However, only 9 cases have CHD.

In order to develop a gene therapy to human cytomegalovirus (HCMV), RNA interference (RNAi) was employed to inhibit the expression of HCMV UL122 gene in vitro. Recombinant vector pUL122-EGFP, which expressed UL122-EGFP fusion protein, and recombinant vectors psi122-1, psi122-2 and psi122-3, which expressed small interfering RNAs (siRNAs) targeted to UL122 were contransfected into AD293 cells. The fluorescence signal of pUL122-EGFP was greatly suppressed by psi122-1 and psi122-2, with an inhibitory rate of 82.0% ± 1.0% and 79.5% ± 2.5%, respectively. The mRNA of pUL122-EGFP of the cells transfected with psi122-1 and psi122-2 was decreased 97.3% ± 0.6% and 98.0% ± 0.1%, respectively. Vector psi122-3 showed a slightly low suppression rate. Therefore, it may be concluded that plasmids encoding siRNAs targeted to UL122 is able to in vitro reduce markedly the expression of UL122-EGFP. And it is very likely that the psi122-1 and psi122-2 are potentially efficacious siRNAs in the gene therapy of HCMV infection in vivo, in which further investigations are required. This study is expected to greatly facilitate the use of the RNAi technology for the anti-HCMV studies. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

A method for the detection of bacterial pathogens in sepsis and bacterial meningitis with 16S rRNA gene— based real-time fluorescent quantitative polymerase chain reaction (FQ-PCR) is developed. A total of 190 blood specimens and 5 cerebrospinal fluid specimens from neonates with suspected sepsis or bacterial meningitis were evaluated with 16S rRNA gene—based real-time FQ-PCR assay. The positive rate of the real-time FQ-PCR assay was significantly higher (25/195, 12.82%) than that of bacterial culture (15/195, 7.69%; P = .002). When bacterial culture was used as a control, the sensitivity of the real-time FQ-PCR was 100%, the specificity was 94.4%, and Youden's index was 0.944. This study suggests that 16S rRNA gene—based real-time FQ-PCR assay is an important and accurate method in the detection of bacterial pathogens of sepsis and bacterial meningitis and should have a promising usage in the diagnosis of sepsis and bacterial meningitis.

Polymerase chain reaction (PCR) techniques have been increasingly used to detect microbial DNA in cerebrospinal fluid (CSF) for the diagnosis of bacterial meningitis. In order to determine the rapidity, sensitivity and specificity of 16S rRNA-based fluorescence quantitative polymerase chain reaction (FQ-PCR), 16S rRNA-based FQ-PCR, CSF bacterial culture and CSF routine analysis were compared in the diagnosis of bacterial meningitis in children. Twenty children who were clinically suspected of bacterial meningitis were included in this study. A total of 2.0 ml of CSF was collected from every child and was subjected to 16S rRNA-based FQ-PCR, CSF culture and CSF routine analysis. Bacterial DNA copies and the cycle threshold (CT) value of the 16S rRNA-based FQ-PCR was recorded, and the results were compared with CSF culture and CSF routine analysis. Seven children were found to be positive with a rate of 35% (7/20) when detected with 16S rRNA-based FQ-PCR and four children displayed a positive rate of 20% (4/20) with the CSF culture method. These two groups displayed a significant difference, with a p-value of 0.002. The method of 16S rRNA-based FQ-PCR demonstrated a high specificity when compared to the standard microbes. A negative correlation was noted between the CT value and the bacteria DNA copies, and the CT value was indicative of the seriousness of bacterial meningitis. 16S rRNA-based FQ-PCR was proved to be a more rapid, sensitive and specific method compared with CSF culture and it should have promising usage in the diagnosis of bacterial meningitis.

The aim of the study was to develop a multiplex PCR-based DNA microarray technology for simultaneous detection and species identification of seven human herpes viruses, namely herpes simplex virus type 1, type 2 (HSV-1, HSV-2), varicella-zoster virus (VZV), Epstein-Barr virus (EBV), cytomegalovirus (CMV), and human herpes virus 6 (HHV-6A, HHV-6B), and to apply this technology to accurate diagnosis of herpesvirus-associated diseases. Primers and oligonucleotide probes were designed and synthesized based on the highly conserved regions of the DNA polymerase gene in human herpes viruses. DNA microarrays were made by printing the oligonucleotide probes onto special glass slides. After amplification and labeling with CY5, the PCR products were hybridized with the DNA microarrays and species identified. Sixty-one cerebrospinal fluid (CSF) and 132 blood specimens were analyzed by this technique, and the results were compared with those of TaqMan PCR. Several specimens were sequenced further after cloning. The PCR products of the seven human herpes viruses ranged from 224 to 252 bp, and could be species identified with DNA microarrays. The detection limits were 10(1) copies/microl for each virus. And the test showed no cross-reaction to DNA extracted from S. aureus, E. coli, hepatitis B virus, Cryptococcus neoformans, Candida albicans and human genome. Among 132 blood and 61 CSF specimens, 55 were tested positive for human herpes virus DNA. Compared with the results of TaqMan PCR, the sensitivity and specificity of the DNA microarray technology was 96.2% and 99.3%, respectively. This multiplex PCR-based DNA microarray technology, which is rapid, specific and sensitive, serves as an effective technique for simultaneous detection and species identification of seven human herpes viruses.

Background: The levels of Th1/Th2 cytokine can alter in pathogenic infection in children with pneumonia. Objectives: To evaluate Th1/Th2 cytokine profile and its diagnostic value in M. pneumoniae pneumonia in children. Patients and Methods: Children with M. pneumoniae mono-infection and 30 healthy children were tested with cytokines assay. We used real time PCR to detect M. pneumoniae in children with pneumonia. Results: M. pneumoniae test was positive in 2188 (16.62%) out of 13161 pneumonia children. Children aged 5 - 9 years had the highest rate and summer was a season with high rate of M. pneumoniae incidence in Zhejiang province. During the course of study, in 526 pneumonia children with M. pneumoniae mono-infection and 30 healthy children cytokines assay was performed. IL-2 level of M. pneumoniae pneumonia children was lower than that of healthy children (median levels, pg/mL: IL-2: 3.2 vs. 5.7, P = 0.00), while IL-4, IL-10 and IFN-γ were higher than in healthy children (median levels, pg/mL: IL-4: 3.2 vs. 1.5, P = 0.00; IL-10: 5.6 vs. 2.5, P = 0.001; IFN-γ: 20.4 vs. 4.8, P = 0.001). Conclusions: IL-2 decreases and IL-4, IL-10 and IFN-γ increase in children with M. pneumoniae pneumonia, which has a promising prospect in diagnosis of this disease in clinical practice.

This study aimed to assess the relevance of laboratory tests in Henoch-Schonlein purpura nephritis (HSPN) classification, and determine accurate classification factors. This prospective study included 694 HSPN patients who underwent ultrasound-guided percutaneous renal biopsy (PRB). Renal specimens were scored according to International Study of Kidney Disease in Children (ISKDC) classification. Meanwhile, blood samples were immediately collected for laboratory examination. The associations between laboratory parameters and HSPN classification were assessed. Significant differences in levels of serum Th1/Th2 cytokines, immunoglobulins, T-lymphocyte subsets, complement, and coagulation markers were obtained between HSPN patients and healthy children. Interestingly, 24h urinary protein (24h-UPRO) levels and urine protein/urine creatinine ratios could determine HPSN grade IIb, IIIa, and IIIb incidences, with areas under ROC curve of 0.767 and 0.731, respectively. At 24h-UPRO >580.35mg/L, prediction sensitivity and specificity were 75.2% and 70.0%, respectively. These values became 53.0% and 82.3%, respectively, with 24h-UPRO exceeding 1006.25mg/L. At urine protein/urine creatinine > 0.97, prediction sensitivity and specificity were 65.5% and 67.2%, respectively, values that became 57.4% and 80.0%, respectively, at ratios exceeding 1.2. Cell and humoral immunity, coagulation and fibrinolytic systems are all involved in the pathogenesis of HSPN, and type I hypersensitivity may be the disease trigger of HSPN. 24h-UPRO levels and urine protein/creatinine ratios could probably forecast the pathological classification of HSPN.

Objective: To purify Mannan-binding lectin (MBL) from human serum and detect its binding ability to several kinds of bacteria common in infectious diseases of children. Methods: MBL was purified from human serum by affinity chromatography on mannan-Sepharose 4B column. Its binding ability to eight species, 97 strains of bacteria was detected by enzyme-linked lectin assay (ELLA). Results: MBL has different binding ability to bacteria and shows strong binding ability to Klebsiella ornithinolytica and Escherichia coli, but shows relatively lower binding ability to Staphylococcus haemolyticus, Enterobacter cloacae and Staphylococcus epidermidis. To different isolates of Klebsiella pneumoniae, Haemophilus influenzae and Staphylococcus aureus, MBL shows quite different binding ability. Conclusions: MBL has different binding ability to different bacteria, and has relatively stronger binding ability to Gram-negative bacteria. Its binding ability to different isolates of certain kinds of bacteria is quite different.

Kawasaki disease (KD) is the primary cause of heart disease among children, but because its clinical symptoms are nonspecific, it is difficult to diagnose. The purpose of this study was to evaluate laboratory indices for possible use in the early diagnosis of KD and to determine which indices are predictive of a response to intravenous immunoglobulin (IVIG) and can be used to monitor the effects of treatment.Three hundred thirty KD patients, 330 age-matched children with KD-like febrile disease, and 330 age-matched healthy children (controls) were enrolled in this prospective study. Levels of N-terminal pro-brain natriuretic peptide (NT-proBNP), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and cytokines were determined in all study subjects.In the derivation cohort, 181 patients in the KD group were compared with 181 patients in the KD-like febrile group. The following indices were found to be useful in the diagnosis of KD: NT-proBNP (area under the curve [AUC] 0.923), ESR (AUC 0.909), CRP (AUC 0.834), and interleukin-6 (IL-6; AUC 0.678). The diagnostic efficiency of each index demonstrated in the derivation cohort was repeated in the 149 KD patients in the validation cohort. There were significant differences in NT-proBNP levels between IVIG-responsive KD patients (n = 270) and IVIG-nonresponsive KD patients (n = 60), with higher NT-proBNP levels in IVIG-nonresponsive KD patients. The NT-proBNP level can effectively distinguish IVIG-responsive KD patients from IVIG-nonresponsive patients, and its AUC was 0.73. There were also significant differences in the NT-proBNP levels before and after treatment, with a significant decline after treatment.Serum levels of NT-proBNP can be used in the diagnosis of KD, the prediction of a patient's sensitivity to IVIG treatment, and the monitoring of the effects of IVIG treatment, but more attention must be paid to the scope of its application.

Respiratory Syncytial Virus (RSV) infections are the dominant cause of pneumonia in children. In order to determine the epidemiological characteristics and immune status of children with Respiratory Syncytial Virus, a prospective study was performed among patients with RSV infection. Comparisons between RSV pneumonia group and normal control group, RSV pneumonia group had lower IL-2 (median levels, pg/ml: 3.8 vs. 5.1, P 0.01), and higher IL-4 (median levels, pg/ml: 3.2 vs. 2.4, P 0.01), IL-10 (median levels, pg/ml: 12.2 vs. 2.3, P 0.01), and IFN-γ (median levels, pg/ml: 13.4 vs. 4.6, P 0.01). The level of IgE among pneumonia patients caused by RSV increased sharply (median levels, mg/L: 48.1 vs. 8.8, P 0.01). Another amazing finding is that after birth, the degree of IgE of the children infected by RSV increases gradually with age. This effect is at its peak in 0.6 years old. The IgE and eosinophil levels were higher when patients suffered from RSV pneumonia with wheeze (IgE median levels, IU/ml: with wheeze: 72.74 vs. without wheeze: 11.5, P 0.05; eosinophil median levels, ×10(9) /l: with wheeze: 0.21 vs. without wheeze: 0.05, P 0.05). The main morbidity crowd is the children under the age of 1 year old. The downregulation of IL2 and the upregulation of IL-4, IL-10, IFN-γ, and IgE happen after RSV infection.

Introduction Asthma is an inflammatory disorder of the airways and the symptoms of asthma could be exacerbated by Mycoplasma pneumoniae infection. Interleukin-2 and interleukin-4 have been implicated in immune and inflammatory reactions. We examined the associations of IL2 and IL4 polymorphisms and expression with the risks of asthma and M. pneumoniae infection in children. Methods A total of 392 asthmatic children and 849 controls were recruited into the study. Eight polymorphisms in IL2 and IL4 were genotyped with Sequenom MassARRAY platform. M. pneumoniae infection and copy number was determined with fluorescence PCR. IL-2 and IL-4 serum expression levels were determined by using ELISA. Results We found a significant association of IL2 rs6534349 polymorphism with increased asthma risk (heterozygotes, P=.029; homozygous variants; P=.013) and of IL4 rs2227284 polymorphism with reduced asthma risk (heterozygotes, P=.026; homozygous variants; P=.001). Besides, the association of other polymorphisms, except rs2070874 polymorphism, became apparent when the asthmatic children were grouped according to GINA classification of asthma control and severity. In addition, IL-2 and IL-4 serum expression levels were significantly higher in M. pneumoniae negative (P=.038) and positive (P=.011) subjects respectively. This observation holds true among asthmatic patients (P=.016 for IL-2 and P=.042 for IL-4), but only the IL-4 observation remained correct among non-asthmatic controls (P=.032). We also observed that the rs6534349 GG genotype was significantly associated with increased odds of getting high load M. pneumoniae infection (P=.0376). Conclusions IL2 and IL4 could be important biomarkers for estimating the risks of asthma and M. pneumoniae infection in children.

Human cytomegalovirus (HCMV) is an ubiquitous pathogen that infects a majority of the world's population. The virus can establish lifelong infection once the human body is infected by HCMV and virus can be reactivated from a latent state in immune suppressed individuals. HCMV has developed several strategies to evade host immune surveillance after millions of years of co-evolution with mankind. One of the classical tricks is encoding homologous to human immune factors or stealing host cellular genes that have significant functions in immune system. Virus encoded immune modulators which participate in regulating the major histocompatibility complex, cellular immunity, humoral immunity, cytokines and chemokines are supposed to play a significant role in the pathogenesis of HCMV. Evaluation of "mutually assured survival" relationship between virus and host provides important insights into viral immunopathogenesis and study of viral immunomodulatory proteins might help us to uncover new human genes that control immunity.

To explore the inhibitory effect of different sources, different concentrations of Mannose-binding lectin (MBL) on human cytomegalovirus infection of human MD-DC cells.The recombinant MBL was acquired by vector construction, and the natural MBL was purified from human plamsa. MD-DC were pre-exposed to several dilutions of the hMBL/rMBL for 30 min, then HCMV suspensions were added to MD-DC for 2 h to compare the inhibitory effect of hMBL/rMBL on the HCMV infection of MD-DC. MD-DC infected by HCMV co-culture with hMBL/rMBL to compare the inhibitory effect of hMBL/rMBL on the HCMV diffusion between MD-DC. HCMV-DNA in MD-DC was detected by fluorescence quantitative PCR. HCMV-PP65 in MD-DC was analyzed with flow cytometry, the ability of MD-DC to capture HCMV was observed with immunofluorescence confocal microscope.In hMBL/rMBL inhibition the ability of MD-DC capture HCMV experiments, the fluorescent quantitative PCR demonstrated that the amount of HCMV-DNA in 1 microg/mL of hMBL/rMBL treated cells was not significantly different from that of control group (P0.05). But the HCMV-DNA in 5 microg/mL and 10 microg/mL hMBL/rMBL treated group were significantly lower than that of control group (P0.05). The significant inhibit effects of 10 microg/mL hMBL/rMBL on the ability of MD-DC capture HCMV were observed by immunofluorescence confocal microscopy and flow cytometry. The inhibit effects of hMBL/rMBL on HCMV diffusion between MD-DC were also observed in 5 microg/mL and 10 microg/mL hMBL/rMBL treated groups at 72 hours.The hMBL/rMBL in physiological concentration range (5-10 microg/mL) can significantly inhibit human cytomegalovirus infection of human MD-DC cells, and the hMBL is more effective than rMBL.

To evaluate the significance of determining ascitic bacterial 16S rRNA by quantitative PCR combined with microarray (PCR-microarray) in the diagnosis of spontaneous bacterial peritonitis (SBP).Ascitic bacterial 16SrRNA was determined by real time fluorescent quantitative PCR-microarray in 76 cases of suspected SBP and 6 cases of non-infectious ascites with chronic liver diseases. The results were compared with ascitic bacterial culture simultaneously.Of 76 ascitic samples, 17 were detected bacteria positive by PCR-microarray, including 8 Grams positive(G+) and 9 Grams negative(G-), which was higher than that by bacterial culture which had only 6 ascitic samples detected positive (all G-); the positive rates were 22.4% vs 7.9%, respectively (P0.01). The bacterial strains detected by both methods in 6 cases had a consistency with each other. No bacteria were detected in another 6 cases of non-infectious ascites with chronic liver diseases.Determination of ascitic bacteria 16S rRNA by PCR-microarray has a higher specificity and sensitivity in the diagnosis of SBP as compared with the bacteria culture. Application of this novel method can not only accelerate SBP diagnosis but also stratify the different pathogens.

To investigate the epidemic characteristics of etiological agents in children with hand, foot and mouth disease (HFMD) and analyze the differences between the severe and mild cases with HFMD seen from 2008 to 2009 in the Children's Hospital.A total of 154 patients with HFMD were enrolled from May 2008 to September 2008 and from May 2009 to September 2009, including 28 severe HFMD patients. Data from 80 cases with suspected herpangina were collected as control. Enterovirus universal type, enterovirus type 71 (EV71) and coxsackie virus group A 16 (CA16) were detected by real-time RT-PCR respectively.The positive rate of enterovirus universal type in the 154 patients with HFMD was 81.82%(126/154). EV71 positive rate in these 126 patients with enterovirus universal type infection was 57.14%(72/126). The positive rate of enterovirus universal type in the 80 cases with suspected herpangina was 68.75%(55/80). There was no EV71 infection in these 80 cases with suspected herpangina. EV71 infection was mainly popular in 2008. Both EV71 and CA16 were prevalent in 2009. The epidemic characteristics of enterovirus infection with HFMD between 2008 and 2009 had significant differences (χ(2) = 23.50, P = 0.000) (P0.01). The epidemic characteristics of enterovirus infection between severe and mild HFMD patients also had significant differences (χ(2) = 29.85, P0.01). There were 28 cases with severe HFMD, in whom the EV71 positive rate was 92.86% (26/28). EV71 positive rate in the mild HFMD was 36.51% (46/126) (χ(2) = 29.22, P0.01). There was no significant difference in the gender (χ(2) = 0.135, P = 0.714) and virus load (t = 0.141, P = 0.889) between the mild and severe HFMD cases. But the age of mild and severe HFMD showed a significant difference (t = 2.926, P = 0.009). Patients who were less than 2 years of age had a proportion of 88.89% (8/9) with severe HFMD. The mean age of mild HFMD patients was 3.19 years.HFMD showed different epidemic characteristics at different times of enterovirus infection. There was no significant difference in the gender and virus load between the mild and severe cases with HFMD. Children under 3 years of age with EV71 infection were at high risk for severe HFMD.

The aim is to investigate the spectrum of disease in 378 infants with human cytomegalovirus infection. In these patients, 27.78% were systemic infection and 72.22% involved single organ infection. Hepatitis, thrombocytopenic purpura, pneumonia were predominant with 33.07%, 13.49%, 6.35% respectively. The rate of HCMV systemic infection in infants younger than 2 weeks was higher than in those older than 2 weeks. The gB genotype analysis in 107 cases showed 53 gBI, 20 gBII, 18 gBIII, 7 gBI+gBII, 5 gBI+gBIII and 4 gBII+gBIII. These results suggest that HCMV can infect multiorgan and has varietal clinic feature. The gBI genotype is most prevalent.

To investigate the frequency of human herpesvirus-6 (HHV-6) encephalitis in pediatric patients under 2 years of age, we developed a method for the simultaneous detection and differentiation of the 2 variants of HHV-6 (HHV-6A and HHV-6B) using subtype-specific, probe-based, real-time PCR (SSPBRT-PCR) and which were further evaluated on 405 cerebrospinal fluid (CSF) specimens from children with suspected encephalitis. A total of 23 (5.70%) out of 405 CSF specimens were positive by SSPBRT-PCR, including 3 cases of HHV-6A and 20 cases of HHV-6B. The positive rate of HHV-6B was significantly higher than that of HHV-6A (P = 0.0004). Compared with the results of the conventional real-time PCR, the sensitivity and specificity of the SSPBRT-PCR assay were 95.24% and 99.22%, respectively. This study suggests a role for both variants of HHV-6 in the pathogenesis of viral encephalitis. SSPBRT-PCR can provide rapid, sensitive, and specific results for identification of HHV-6A and HHV-6B and management of HHV-6 encephalitis.

The aim of this study was to determine differences in the gene expression profile of kidney tissue from type 2 diabetes mellitus (T2D) and control rats using DNA microarray analysis. Total RNA was extracted from the kidney tissue of the T2D and control rats using the original single step method. cDNA retro-transcribed from an equal quantity of mRNA was labeled with Cy5 and Cy3 fluorescence and served as the probes. The mixed probes were hybridized to a DNA microarray. Fluorescence signals were scanned by an ScanArray 4000 laser scanner and further analyzed by QuantArray software. Apoptotic cells were detected in situ using the Roche TUNEL assay. Serum glucose, ApoAI, ApoB, ApoA1/ApoB, cholesterol and triglyceride levels were significantly higher in the T2D rats than in the controls, but there were no significant differences in serum insulin. When the kidney tissue was screened using the DNA microarray, differential expression was found for 41 genes. Five genes in the T2D rats were upregulated by 2-fold compared to the control rats, while 36 genes were down-regulated by 0.5-fold. Moreover, in the renal tubular epithelial cells, there was a significantly greater number of TUNEL-positive cells in the T2D group than in the control group. A total of 41 genes are associated with the occurrence and development of T2D and diabetic nephropathy. The present study suggests that examining differences in gene expression profiles is of benefit to the diagnosis, treatment and prevention of T2D, diabetic nephropathy and other T2D complications.

Human herpesvirus 6 (HHV-6) isolates are classified into two variants, HHV-6A and HHV-6B, based on distinct genetic, antigenic and biological characteristics. HHV-6 has been associated with encephalitis in children recently. This study aimed to establish a real time PCR assay for simultaneous detection of the two subtypes of HHV-6, and apply this new assay to children with suspected encephalitis, then analyze the relationship between the infection with HHV-6 and encephalitis in children.The universal primers and variant-specific TaqMan probes were designed based on the highly conserved sequences of the DNA polymerase gene (U38) of HHV-6. The 5' end of the probes for HHV-6A and HHV-6B was labeled with the fluorescein reporter tetrachloro-6-carboxyfluorescein and 6-carboxyfluorescein (6-FAM), separately, while the 3' end were quenched with 6-carboxy-tetramethylrhodamine. The real time PCR assay for simultaneous detection of HHV-6A and HHV-6B was established. Then, the plasmids of HHV-6A and -6B which were diluted by a 10-fold series from 10(9) to 10(0) copies/microl, together with controls were used for testing both sensitivity and specificity of the real time PCR assay. The cerebrospinal fluid (CSF) specimens from 445 cases of suspected encephalitis were tested with this real time PCR and positive samples were then sequenced.Both HHV-6A (strain ZJ-159) and HHV-6B (strain GS) were positive on the real time PCR assay. There were no cross-reaction with herpes simplex virus type 1, type 2 (HSV-1, HSV-2), varicella-zoster virus (VZV), cytomegalovirus (CMV), Epstein-Barr virus (EBV), hepatitis B virus, Staphylococcus aureus, Mycoplasma pneumoniae and human DNA. A linear regression curve was obtained when plotting Ct values against the log10 of the viral DNA input for both subtypes of HHV-6. The sensitivity threshold was 10 copies/microl for the real time PCR. HHV-6 positive rate by the real time PCR assay was 4.72% (21/445), including 4 cases with HHV-6A infection, 16 cases of HHV-6B infection and 1 case with mixed HHV-6A and HHV-6B infection. The new PCR assay usually took 2 to 3 hours to provide results.This new real time PCR assay can simultaneously detect both subtypes of HHV-6, and have high specificity and sensitivity. It will provide an early and sensitive diagnosis of HHV-6 encephalitis in children.

In order to develop a gene therapy to human cytomegalovirus (HCMV), RNA interference (RNAi) was employed to inhibit the expression of HCMV UL122 gene in vitro. Recombinant vector pUL122-EGFP, which expressed UL122-EGFP fusion protein, and recombinant vectors psi122-1, psi122-2 and psi122-3, which expressed small interfering RNAs (siRNAs) targeted to UL122 were contransfected into AD293 cells. The fluorescence signal of pUL122-EGFP was greatly suppressed by psi122-1 and psi122-2, with an inhibitory rate of 82.0% +/- 1.0% and 79.5% +/- 2.5%, respectively. The mRNA of pUL122-EGFP of the cells transfected with psi122-1 and psi122-2 was decreased 97.3% +/- 0.6% and 98.0% +/- 0.1%, respectively. Vector psi122-3 showed a slightly low suppression rate. Therefore, it may be concluded that plasmids encoding siRNAs targeted to UL122 is able to in vitro reduce markedly the expression of UL122-EGFP. And it is very likely that the psi122-1 and psi122-2 are potentially efficacious siRNAs in the gene therapy of HCMV infection in vivo, in which further investigations are required. This study is expected to greatly facilitate the use of the RNAi technology for the anti-HCMV studies.

To study the relationship of nonsyndromic cleft lip and/or palate (NSCL/P) and poliovirus receptor-related 1 exon3 (PVRL1exon3) polymorphisms in Han People of Jiangzhe area.PVRL1exon3 was examined by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) technique in the 50 patients with NSCL/P and 85 healthy parents.No W185X mutation was found in the PVRL1exon 3.It indicates that there is no relationship between NSCL/P and PVRL1exon3 in Han People in Jiangzhe area.

To explore a new method of rapid and reliable diagnosis of bacterial infectious diseases such as purulent meningitis and septicemia.A pair of universal primers and a set of probes (including universal fluorescence probe, Gram-positive probe and Gram-negative probe) were designed based on the bacterial highly conserved region of 16S rRNA gene. By using the FQ-PCR method, 12 standard strains, 23 clinical cultural isolations and the controls such as HBV, Cryptococcus histolyticus, Blastomyces albicans and human DNA were detected with the three kinds of probes. The correlation among the results of the three kinds of probes detection was analyzed.The determination of 16S rRNA gene with FQ-PCR was a highly specific and sensitive method and not cross-reactive with human DNA, virus or fungi. The least amount of 10 copies of 16S rRNA gene which corresponded to 2 bacteria could be detected with FQ-PCR. Twelve standard strains and 23 clinical cultural isolations were detected by FQ-PCR with the three kinds of probes mentioned above. All samples presented positive results using the universal probe. The results of 16S rRNA gene detected by the Gramjpositive probe were positive to the 18 G+ strains. The results of 16S rRNA gene detected by the Gram-probe were positive to the 17 G- strains.The FQ-PCR technique was established for bacteria quantifying and typing using the universal primer and the double type probes. This method was convenient and rapid in detecting, quantifying and typing bacteria, with a high specificity and sensitivity.

Sepsis is a serious disease with high mortality in newborns. It is very important to have a convenient and accurate method for pathogenic diagnosis of neonatal sepsis. We developed a method of simultaneous detection and Gram classification of clinically relevant bacterial pathogens causing sepsis directly from blood samples with Gram stain-specific-probe-based real-time PCR (GSPBRT-PCR). With GSPBRT-PCR, 53 clinically important strains representing 25 gram-positive and 28 gram-negative bacterial species were identified correctly with the corresponding Gram probe. The limits of the GSPBRT-PCR assay in serial dilutions of the bacteria revealed that Staphylococcus aureus could be detected at concentrations of 3 CFU per PCR and Escherichia coli at concentrations as low as 1 CFU per PCR. The GSPBRT-PCR assay was further evaluated on 600 blood specimens from patients with suspicioon of neonatal sepsis and compared to the results obtained from blood cultures. The positive rate of the GSPBRT-PCR array was 50/600 (8.33%), significantly higher than that of blood culture (34/600; 5.67%) ( P = 0.00003). When blood culture was used as a control, the sensitivity of GSPBRT-PCR was 100%, the specificity was 97.17%, and the index of accurate diagnosis was 0.972. This study suggests that GSPBRT-PCR is very useful for the rapid and accurate diagnosis of bacterial infection and that it can have an important impact on the current inappropriate and unnecessary use of antibiotics in the treatment of newborns.

To evaluate the usefulness of a broad-range real-time PCR assay aimed at the 16S rRNA gene of bacteria in a clinical setting in rapid and reliable diagnosis of neonatal septicemia for improving the speed and accuracy of bacterial detection.The universal primer and TaqMan probe were designed based on the highly conserved sequences of the bacterial 16S rRNA gene. The chosen primers and probe did not show any likely cross hybridization with human, viral or fungal genome sequences. The TaqMan assay used the fluorescent signal on the probe, such as 6-carboxyfluorescin (6-FAM), and quenched by the standard 6-carboxytetramethylrhodamine (TAMRA) probes. The broad-range 16S rRNA gene real-time PCR array was established. Then, three common pathogenic microorganisms including Staphylococcus aureus, Staphylococcus epidermidis and Escherichia coli, which were prepared by a 10-fold dilution series respectively from 10(8) colony forming unit (CFU)/ml to 10(3) CFU/ml, as well as controls, were used for testing of both sensitivity and specificity of the real-time PCR assay. The blood samples from 830 cases of suspected septicemia, who were hospitalized in our neonatal ward and the neonatal intensive care unit (NICU) and developed clinical signs suggestive of infection, were tested with routine culture and bacterial 16S rRNA genes real-time PCR separately. In addition, 30 neonates without infection were enrolled as the negative control group.All the three common pathogenic bacterial species were positive on the 16S rRNA genes real-time PCR assay. There were no cross-reaction with cytomegalovirus (CMV), Epstein-Barr virus (EBV), hepatitis B virus (HBV), fungi, human DNA and blank control, and the technique showed high specificity and sensitivity. The detection limit of the TaqMan assay was tested by amplifying serial dilutions of the three common pathogenic bacterial DNA. The minimal detection limit of the TaqMan system was equivalent to 3 CFU of bacteria, the threshold cycle (CT), which is inversely proportional to the log of the amount of target DNA initially present, was 37.90 by calculation. The real-time PCR assay was evaluated on 830 blood specimens for suspected neonatal septicemia, as compared to the results obtained from the routine bacterial cultures. The positive rate by the real-time PCR assay was 5.18% (43/830) in 830 samples, and was significantly higher than that of blood culture [2.41% (20/830) (P0.01)]. The real-time PCR was positive in all the 20 positive blood culture samples. Thirty non-infectious blood samples were negative by both the PCR assay and blood cultures. When blood culture was used as control, the sensitivity of the real-time PCR assay was 100%, the specificity was 97.16%, and the index of accurate diagnosis was 0.972. Moreover, three of the PCR positive amplicons were confirmed by sequencing to confirm the accuracy of the real-time PCR assay in testing clinical specimens. The sequencing showed that except for one sequence, all the others were demonstrated to be Staphylococcus aureus and Escherichia coli respectively, which was in accord with the results of the blood cultures.The bacterial 16S rRNA genes real-time PCR had been established to diagnose the neonatal septicemia. The sensitivity and specificity the real-time PCR assay were higher than those of blood culture. This technique can provide a rapid way for the etiological diagnosis of neonatal septicemia, and was a convenient and accurate method in etiologic diagnosis of neonatal septicemia.

Recent discoveries indicate that the molecules in glomerular podocytes and slit diaphragms may play an important role in the development of proteinuria and nephrotic syndrome. Mutational analyses of NPHS1 and NPHS2 were performed to verify this hypothesis in sporadic nephrotic syndrome (NS) patients. Clinical characteristics and DNA samples were collected from 38 Chinese children with sporadic steroid-sensitive NS, 22 with steroid-resistant NS and 30 controls. Direct sequencing was performed after PCR amplification of all 29 and 8 exons of the NPHS1 and NPHS2 genes, respectively. In NPHS1, 4 patients had heterozygous missense mutations leading to amino acid substitutions (R800C, Q453R). Furthermore, 3 known single nucleotide polymorphism (SNP) were found (T741T, V763V, S1105S). In NPHS2, 3 patients had novel heterozygous allelic variants leading to amino acid substitutions (S206I, E188D), while 1 patient was found to carry a novel nonsense mutation leading to a truncated protein product (Glu237STOP). Two known polymorphisms were also found (A318A, L346L). The results demonstrate that NPHS1 and NPHS2 mutations are also present in Chinese sporadic NS patients, suggesting that genetic changes of nephrin and podocin may play pathogenetic roles in some patients with sporadic steroid resistant NS.

To study the pathogenic bacteria of lower respiratory tract infection (LRTI), and age and gender distribution and drug resistance of the pathogenic bacteria in children.Sputum specimens for bacterial cultures were collected in sterile tubes from all of the children with LRTI who had been admitted to the Children's Hospital of Zhejiang University between August 2001 and July 2002. Antibiotic susceptibility tests were performed using the Vitek system, the Kirby-Bauer diffuse method and the Etest method after bacteria were identified.Among the 4,238 patients with LRTI during the study period, 1,181 patients were bacteria-positive, with a positive rate of 27.9%. Streptococcus pneumoniae (S. pneumoniae) was the most common (222 strains), followed by Haemophilus influenzae (H. influenzae) (216 strains), Klebsiella pneumoniae (K. pneumoniae) (216 strains), Escherichia coil (E. coli) (169 strains) and Staphylococcus aureus (S. aureus) (89 strains). The isolation rate of S. pneumoniae in females was significantly higher than in males (6.2% vs 4.7%; P0.05). However, the isolation rates of K. pneumoniae and S. aureus in males were higher than in females (5.1% vs 4.1% and 2.5% vs 1.5%, respectively; P0.05). A higher incidence of LRTI due to S. pneumoniae and H. influenzae was found in the 1-3 years group, while the incidence of LRTI due to K. pneumoniae, E. coli, S. aureus and E. cloacae was higher in patients under 1 year of age. Antibiotic susceptibility tests showed that rates of penicillin non-susceptible S. pneumoniae, ampicillin resistant H. influenzae, oxacillin-resistant S. aureus and ESBL-positive K. pneumoniae and E. coli were 55.0%, 16.5%, 41.2%, 42.6% and 4.5%, respectively.S. pneumoniae, H. influenzae, K. pneumoniae, E. coli and S. aureus were common pathogens of LRTI in children. The infection rate varied with age and gender. Antibiotics for treating LRTI should be selected based on the drug susceptibility test.