Human adenovirus (HAdV) infection in children with acute respiratory tract infections in Guangzhou, China, 2010-2021: a molecular epidemiology study.

BACKGROUND: Human adenovirus (HAdV) infection can cause a variety of diseases. It is a major pathogen of pediatric acute respiratory tract infections (ARIs) and can be life-threatening in younger children. We described the epidemiology and subtypes shifting of HAdV among children with ARI in Guangzhou, China. METHODS: We conducted a retrospective study of 161,079 children diagnosed with acute respiratory illness at the Guangzhou Women and Children's Medical Center between 2010 and 2021. HAdV specimens were detected by real-time PCR and the hexon gene was used for phylogenetic analysis. RESULTS: Before the COVID-19 outbreak in Guangzhou, the annual frequency of adenovirus infection detected during this period ranged from 3.92% to 13.58%, with an epidemic peak every four to five years. HAdV demonstrated a clear seasonal distribution, with the lowest positivity in March and peaking during summer (July or August) every year. A significant increase in HAdV cases was recorded for 2018 and 2019, which coincided with a shift in the dominant HAdV subtype from HAdV-3 to HAdV-7. The latter was associated with a more severe disease compared to HAdV-3. The average mortality proportion for children infected with HAdV from 2016 to 2019 was 0.38% but increased to 20% in severe cases. After COVID-19 emerged, HAdV cases dropped to 2.68%, suggesting that non-pharmaceutical interventions probably reduced the transmission of HAdV in the community. CONCLUSION: Our study provides the foundation for the understanding of the epidemiology of HAdV and its associated risks in children in Southern China.

The Seroprevalence of Some Pathogen Specific IgM in Children with Acute Respiratory Tract Infections in Guangzhou Region, 2011 - 2012.

BACKGROUND: Acute respiratory tract infections (ARTIs) are the leading cause of morbidity and death in children < 5 years worldwide. The aim of this study is to analyze the seroprevalence of nine pathogen specific IgMs in children with ARTIs with respect to gender, age, and seasonality in the Guangzhou region. METHODS: Serum samples were collected from 20160 children with ARTIs admitted to the Guangzhou Women and Children's Medical Center between 2011 and 2012. Serum-specific IgM antibodies to nine respiratory pathogens, Mycoplasma pneumonia (MP), Legionella pneumophila (LP), Coxiella burnetii (C. burnetii), Chlamydophila pneumonia (CP), adenovirus (ADV), respiratory syncytial virus (RSV), type A and type B influenza virus (IVA and IVB), and parainfluenza virus (PIV), were detected using immunofluorescence assay. RESULTS: The male-to-female ratio of all patients was 1.9:1. The median age was 3 years and 8 months with a significant difference in seropositivity to respiratory tract pathogens between children from different age groups. Seropositivity was detected in 43.53% of the children with the top three pathogens being MP (33.15%), RSV (10.27%), and ADV (6.63%), followed by IVB (2.63%), LP (2.25%), IVA (1.59%), PIV (1.57%), CP (0.27%), and C. burnetii (0.13%). The prevalence of single, double, and triple seropositivity was 70.20% (6160/8775), 25.22% (2213/8775), and 4.57% (401/8775), respectively. The total IgM seropositivity for any kind of pathogen in the nine kinds of pathogens peaked in winter (46.53%), while the nadir was observed in summer (41.97%). CONCLUSIONS: The top three seroprevalence of nine kinds of pathogen specific IgM was MP, followed by RSV and ADV. The epidemic pathogen specific IgM had a season-specific seropositivity distribution. Seroprevalence of the pathogen should be a focus of attention.

[Complete genomic sequence analysis on human enterovirus 71 strain in Guangzhou, in 2008 and 2010].

OBJECTIVE: To study the genomic genotypes and variation of human enterovirus 71 (EV71) infected infants in Guangzhou city, in 2008 and 2010. METHODS: Primers were designed on the basis of the genomic sequence of EV71 SHZH03 strain (AY465356) in the GenBank, and EV71 genome amplified by RT-PCR. PCR-products were directly sequenced and the genomic nucleotide sequences were analyzed with the programs of Clustal W/X, DNASTAR and MEGA 4.1. RESULTS: 9 strains of EV71 genome appeared to be 7405 bp in length. The genomic sequences of EV71 Guangzhou strains were compared with those of EV71 in GenBank, which revealed that the homology with EV71 genotype C4a Fuyang strains ranged between 98% - 99%. Homology with genotype C4b were 92% - 94%, with genotypes C1, C2, C3 as 82% - 83%, with genotypes B3, B4, B5 as 81% - 83% and the homology with genotype A was 80%. When compared the VP1 genes of EV71 Guangzhou strains with genotypes A, B, C virus, we revealed that the highest homology was also with genotype C4a. When compared the VP1 amino acid sequences of EV71 Guangzhou strains with genotype A, B, C virus by Clustal W program, the results revealed that the amino acid residue Q at position 22 in VP1 gene was transformed to H, while 213 (S→T) and 1764 (V→I) mutations in polyprotein were discovered. CONCLUSION: Data from the sequences and phylogenetics analysis on those Guangzhou strains in 2008 and 2010 revealed that those isolates belong to genotype C4a, with the homology with Fuyang strains as 98%-99%. Mutation of amino acid residue H at position 22 in VP1 gene was discovered and the neutralizing antibody of EV71 might have been conversed by this residue. 213 (S→T) and 1764 (V→I) mutations in polyprotein were also discovered.

[Preparation, characterization and preliminary application of monoclonal antibodies against the expressed Norovirus capsid protein].

AIM: To prepare the monoclonal antibodies (mAbs) against Norovirus capsid protein for the development of a rapid assay of Norovirus and to investigate the pathogenesis of this virus. METHODS: Sp2/0-Ag-14 myeloma cells were fused with spleen cells of BALB/c mice immunized with the recombinant protein of Norovirus NVgz01 (DQ369797), which was overexpressed in E.coli. The monoclonal antibodies against Norovirus capsid protein were screened by selective culture medium. The obtained Ig subtypes, titer and specificity of the mAbs were examined by ELISA and Western blot respectively. RESULTS: After cell fusion and subcloning, four hybridoma cell lines which secreted monoclonal antibodies specifically against Norovirus capsid protein were obtained and named as N2C3, N7C2, N4B1 and N8A9. Indirect ELISA and Western blot assay showed that the four mAbs specifically recognized the Norovirus capsid protein expressed in E.coli. The native Norovirus capsid protein in the stool samples were also recognized by them. The Sandwich ELISA, a rapid detection assay of the expressed and native Norovirus capsid protein, demonstrated that N2C3 and N7C2 were matched successfully. CONCLUSION: The mAbs against GII Norovirus capsid protein have been developed, which can be used for the development of detection assay and the basic research of Norovirus.

[Preparation and characterization of specific monoclonal antibodies against hexon of HAdV 3].

OBJECTIVE: To obtain the monoclonal antibody against hexon protein of human adenovirus. METHODS: BALB/c mice were immunized with purified recombinant hexon protein, and the spleen cells of the mice were isolated and fused with myloma cells. Four hybridoma cell strains were screened by indirect ELISA and cultured, and the sensitivity, specificity and virus neutralizing activity were analyzed with ELISA, Western blotting and neutralizing test. RESULTS: The mouse ascites produced by these hybridoma cells contained specific monoclonal antibodies against hexon protein of human adenovirus as identified by ELISA and Western blot, and the antibody generated by 4C6 strain showed human adenovirus type 3-neutralizing activity. CONCLUSION: The monoclonal antibodies against hexon protein with high specificity have been successfully obtained, and these antibodies can be useful in developing assays for early diagnosis of HAdV3 infection and also in study of therapeutic drugs of the infection.

[Cloning, expression and immunocharacterization of the capsid protein of human Norwalk virus Guangzhou strain NVgz01].

OBJECTIVE: To clone, express and characterize the capsid protein of human Norwalk virus Guangzhou strain NVgz01. METHODS: On the basis of successful construction of full-genome clones and sequence analysis of human norovirus Guangzhou strain NVgz01, the full capsid gene was ligated into pET28a (+) for expression. After IPTG induction, the recombinant protein was purified through metal (Ni(2+)) chelating affinity chromatography. Western blotting and enzyme-linked immunosorbent assay (ELISA) were used to determine the antigenicity of the recombinant protein. RESULTS: The recombinant capsid gene was overexpressed in E.coli, yielding the recombinant protein with relative molecular mass of 62x10(3) that was highly purified through metal (Ni(2+)) chelating affinity chromatography. IDEIA Norovirus Kit and immunoassay showed that the recombinant protein had good antigenicity. CONCLUSION: The capsid gene of norovirus Guangzhou strain has been cloned and expressed, which can be useful for developing diagnostic reagents or vaccine of norovirus.

[Role of bFGF and TGF-beta1 in primary cultured prostatic stromal cells].

OBJECTIVE: To study the role of the basic fibroblast growth factor (bFGF) and transforming growth factor beta1 (TGF-beta1) in benign prostatic hyperplasia (BPH). METHODS: The human stromal cells of BPH were isolated and cultured. The proliferation of the stromal cells cultured in serum-free medium was detected by MTT method, the phenotype changes of smooth muscle cells detected by immunohistochemical method, and the effect of different concentrations of bFGF and TGF-beta1 on the cultured stromal cells of BPH observed. RESULTS: bFGF stimulated the cultured BPH stromal cell proliferation (P < 0.05, P < 0.01) and decreased the expression of smooth muscle cell (SMC) phenotype in higher concentration (10 microg/L). TGF-beta1 (> 1 microg/L) inhibited stromal cell proliferation and increased the expression of SMC phenotype (P < 0.05, P < 0.01). 5 microg/ml bFGF and TGF-beta1 (0.001 microg/L, 0.01 microg/L) promoted stromal cell proliferation (P < 0.01), while 5 microg/L bFGF and TGF-beta1 (0.1 microg/L, 1 microg/L, 10 microg/L) inhibited it, slightly in 0.1 microg/L (P > 0.05) and significantly in 1 microg/L and 10 microg/L (P < 0.01), and increased the expression of SMC phenotype in higher concentration (> 1 microg/L, P < 0.01). CONCLUSION: bFGF stimulates the proliferation of the prostatic stromal cells of BPH in a time- and dose-dependent fashion and decreases the expression of SMC phenotype, TGF-beta1 inhibits the growth of stromal cells and induces the differentiation of stromal cells to SMC, both playing an important role in the mechanism of BPH.