To construct a recombinant replication-defective human adenovirus type 5 expressing Cap protein of PCV2 and test the immunological efficacy in mice. In this study, the recombinant replication-defective human adenovirus type 5, named as rAd5-Cap (wt-rAd5), was constructed through homologous recombination internally in the HEK293AD cells after co-transfection of the Pac I-linearized backbone plasmid and the shuttle plasmid pacAd5CMV-Cap containing the open reading frame (ORF2) of the porcine circovirus type 2 (PCV2) cap protein or pacAd5CMV without inserted fragment. Furthermore, the rAd5-Cap could induce the expression of PCV2 cap protein in the HEK293AD cells with high efficacy evaluated by the RT-PCR and indirect immunofluorescence assay (IFA). The virus titer of rAd5-Cap could reach up to 10(8.5) TCID50/mL similarly to that of wt-rAd5, indicating that there was little affect on the virus proliferation after the insertion of PCV2 cap protein gene. The humeral immune responses could be activated and detected 14 days after the inoculation of the mice with 10(7) TCID50 rAd5-Cap intramuscularly, and constantly in crease in another 14 days. These molecular biological and animal experiments results demonstrated that the PCV2 cap protein could be efficiently expressed by the recombinant adenovirus rAd5-Cap in eukaryotic cells and induce robust immune responses in mice, which laid a good foundation for the development of new type vaccine against porcine circovirus.
Adenoviruses, Human ; genetics ; Animals ; Antibodies, Viral ; blood ; Capsid Proteins ; genetics ; immunology ; Circovirus ; immunology ; Defective Viruses ; genetics ; HEK293 Cells ; Humans ; Mice ; Recombinant Proteins ; biosynthesis ; immunology ; Virus Replication

PURPOSE: The Purposes of this study are to identify the circulating etiologic viruses of acute lower respiratory tract infection in children and to understand the relation with clinical diagnosis. METHODS: We obtained a total of 418 nasopharyngeal aspirates from children admitted for their acute lower respiratory tract infections at three tertiary hospitals in Seoul from September 2008 to March 2009. We performed multiplex RT-PCR to identify 14 etiologic viruses and analyzed their emerging patterns and clinical features. RESULTS: Average age of patients was 16.4 months old and the ratio of male to female was 1.36. Viruses were detected in 56.2% of a total of 418 samples. Respiratory syncytial virus (35%) was the most frequently detected and followed by human rhinovirus (22%), human bocavirus, adenovirus, human metapneumovirus, parainfluenza virus, influenza virus and human coronavirus. Co-infection reached 21.9% of positive patients. CONCLUSION: When we manage the patients with acute lower respiratory infectious diseases, we should remind the role of various viral pathogens, which might be circulating by seasons and by local areas.
Adenoviruses, Human ; Child ; Coinfection ; Communicable Diseases ; Coronavirus ; Female ; Human bocavirus ; Humans ; Male ; Metapneumovirus ; Orthomyxoviridae ; Paramyxoviridae Infections ; Respiratory Syncytial Viruses ; Respiratory System ; Respiratory Tract Infections ; Rhinovirus ; Seasons ; Tertiary Care Centers ; Viruses

Bovine coronavirus (BCoV) causes severe diarrhea in newborn calves, and is associated with winter dysentery in adult cattle and respiratory infections in calves and feedlot cattle. Although the Korean BCoV vaccine strain, BC94, was isolated in 1995, there has still been no report of a molecular characterization of the vaccine strain. To characterize the vaccine strain, relationships between BC94 and field strains were investigated, based on sequence analysis and cross-immunity. We determined the complete sequences of the HE, N, and S genes from BC94 and four NVRQS isolates (SUN5, A3, 0501, 0502). Due to its major role in antigenicity, the spike proteins of the BCoVs were analyzed. BC94 showed distinctive genetic divergence from field isolates collected from 2002 to 2005. BC94, SUN5, and A3 had no virulence-specific sequence and there was a single amino acid change, from asparagine to lysine at residue 175, in the polymorphic region. Strains 0501 and 0502 had virulence-specific sequences at all seven sites. Although the recently isolated Korean BCoVs and BC94 were genetically different, the cleavage site of spike genes at 763~768 (KRRSRR) and the antigenic domain II of the spike protein, amino acid position 528, were conserved in all NVRQS isolates. The antigenic relatedness of KCD9, representative of recent Korean BCoVs, was compared with the Korean vaccine strain BC94. KCD9 showed cross-reactivity against BC94 by virus neutralization (VN) test. These results suggest that BC94 is antigenically closely related to field isolates and is still effective as a vaccine strain.
Adult ; Animals ; Asparagine ; Cattle ; Coronavirus, Bovine ; Diarrhea ; Dysentery ; Humans ; Infant, Newborn ; Korea ; Lysine ; Proteins ; Respiratory Tract Infections ; Sequence Analysis ; Sprains and Strains ; Viruses

Bovine coronavirus (BCoV) causes severe diarrhea in newborn calves, and is associated with winter dysentery in adult cattle and respiratory infections in calves and feedlot cattle. Although the Korean BCoV vaccine strain, BC94, was isolated in 1995, there has still been no report of a molecular characterization of the vaccine strain. To characterize the vaccine strain, relationships between BC94 and field strains were investigated, based on sequence analysis and cross-immunity. We determined the complete sequences of the HE, N, and S genes from BC94 and four NVRQS isolates (SUN5, A3, 0501, 0502). Due to its major role in antigenicity, the spike proteins of the BCoVs were analyzed. BC94 showed distinctive genetic divergence from field isolates collected from 2002 to 2005. BC94, SUN5, and A3 had no virulence-specific sequence and there was a single amino acid change, from asparagine to lysine at residue 175, in the polymorphic region. Strains 0501 and 0502 had virulence-specific sequences at all seven sites. Although the recently isolated Korean BCoVs and BC94 were genetically different, the cleavage site of spike genes at 763~768 (KRRSRR) and the antigenic domain II of the spike protein, amino acid position 528, were conserved in all NVRQS isolates. The antigenic relatedness of KCD9, representative of recent Korean BCoVs, was compared with the Korean vaccine strain BC94. KCD9 showed cross-reactivity against BC94 by virus neutralization (VN) test. These results suggest that BC94 is antigenically closely related to field isolates and is still effective as a vaccine strain.
Adult ; Animals ; Asparagine ; Cattle ; Coronavirus, Bovine ; Diarrhea ; Dysentery ; Humans ; Infant, Newborn ; Korea ; Lysine ; Proteins ; Respiratory Tract Infections ; Sequence Analysis ; Sprains and Strains ; Viruses

PURPOSE: The purpose of this study is to identify the viral etiology of acute respiratory illnesses and to determine epidemiology in outpatients in Busan, Korea. METHODS: We collected nasal wash samples from 990 patients who visited the hospital for acute respiratory illnesses between January 2007 and December 2008. Extracted DNA or RNA from specimens was used for viral detection by an RT-PCR method. RESULTS: Of a total of 990 samples, viruses were detected in 351 cases (35.5%). The ratio of male to female was 1.6:1 and 93.7% were less than 5 years old. Rhinovirus was detected year-round in 202 cases (57.5%), respiratory syncytial virus from October to March in 57 cases (16.2%), adenovirus year-round in 37 cases (10.5%), influenza virus from December to April in 21 cases (6%), bocavirus from January to August in 15 cases (4.3%), parainfluenza virus from April to July in 9 cases (2.6%), coronavirus from January to July in 7 cases (2%), and enterovirus from June to September in 3 cases (0.9%). CONCLUSION: We identified the etiology and epidemiology of viruses that caused the acute respiratory diseases that were prevalent in Busan, 2007-2008. Further surveillance will be necessary.
Adenoviridae ; Bocavirus ; Coronavirus ; DNA ; Enterovirus ; Female ; Humans ; Male ; Orthomyxoviridae ; Outpatients ; Paramyxoviridae Infections ; Respiratory Syncytial Viruses ; Rhinovirus ; RNA ; Viruses

PURPOSE: This study aimed to investigate the association between respiratory virus infection and pneumococcal colonization in children. METHODS: From May 2009 to June 2010, nasopharyngeal (NP) aspirates were obtained from patients under 18 years old who visited Seoul National University Children's Hospital for respiratory symptoms. NP samples were used to detect respiratory viruses (influenza virus A and B, parainfluenza virus 1, 2 and 3, respiratory syncytial virus A and B, adenovirus, rhinovirus A/B, human metapneumovirus, human coronavirus 229E/NL63 and OC43/HKU1) by RT-PCR and pneumococcus by culture. RESULTS: Median age of the patients was 27 months old. A total of 1,367 NP aspirates were tested for respiratory viruses and pneumococcus. Pneumococcus was isolated from 228 (16.7%) of samples and respiratory viruses were detected from 731 (53.5%). Common viruses were rhinovirus (18.4%), respiratory syncytial virus (RSV) A (10.6%), adenovirus (6.9%), influenza virus A (6.8%). Pneumococcal isolation rate was significantly higher in the cases of positive virus detection than negative detection [21.3% (156/731) vs. 11.3% (72/636), P<0.001]. For individual viruses, pneumococcal isolation rate was positively associated with detection of influenza virus A [24.7% (23/93) vs 16.1% (205/1274), P=0.001], RSV A [28.3% (41/145) vs 15.3% (187/1222), P=0.001], RSV B [31.3% (10/32) vs 16.3% (218/1335), P=0.042], rhinovirus A/B [22.6% (57/252) vs 15.3% (171/1115), P=0.010]. CONCLUSION: The study revealed that pneumococcal isolation from NP aspirates is related with respiratory virus detection. The result of this study could be used to investigate how respiratory viruses and pneumococcus cause clinical diseases.
Adenoviridae ; Child* ; Colon* ; Coronavirus ; Humans ; Metapneumovirus ; Orthomyxoviridae ; Paramyxoviridae Infections ; Respiratory Syncytial Viruses ; Rhinovirus ; Seoul ; Streptococcus pneumoniae

Obesity is connected with numerous diseases, such as type 2 diabetes, atherosclerosis, cancer, and nervous system dysfunctions. Obesity is affected by genetic, environmental, and cultural factors. However, numerous studies indicate that several pathogens might cause obesity. This review discusses recent data and the characteristics of pathogens that are implicated in obesity. In particular, human adenovirus 36 (Ad36) is the most clearly implicated virus in human obesity. It was recently shown that obese groups from the USA, Korea, and Italy have a higher prevalence of serum antibodies against Ad36. The mechanisms of Ad36-induced obesity remain unclear. However, glucose uptake and inflammation are possible mechanisms of Ad36-induced obesity. Overall, this new understanding of causes of obesity has developed into the concept of 'infectobesity' and the possibility of developing a 'vaccine' or 'therapeutic agents' for obesity.
Adenoviruses, Human ; Antibodies ; Atherosclerosis ; Glucose ; Humans ; Inflammation ; Italy ; Korea ; Nervous System ; Obesity ; Prevalence ; Viruses

Adenoviruses, Human ; Child ; Humans ; Orthomyxoviridae ; Respiratory Syncytial Viruses ; Respiratory Tract Infections ; virology

PURPOSE: Gene therapy (ex vivo) has recently been used as a means of delivering bone morphogenetic proteins (BMPs) to sites of tissue regeneration. In the present study, we investigated the effect of co-transduction of adenoviruses expressing BMP-2 and BMP-7 on osteogenesisof C2C12 cells in vitro. METHODS: A replication-defective human adenovirus 5 (Ad5) containing a cDNA for BMPs in the E1 region of the virus (Ad5BMP-2 and Ad5BMP-7) was constructed by in vivo homologous recombination. Functional activity of Ad5BMP-2 and Ad5BMP-7 were evaluated in mouse stromal cells (W20-17cells). C2C12 cells are transduced with various MOI (multiplicity of infection) of Ad5BMP-2 and Ad5BMP-7 to assess most effective and stable titer. Based on this result, C2C12 cells were transduced with Ad5BMP-2 and Ad5BMP-7 alone or by combination. BMPs expression, alkaline phosphatase (ALPase) activity, cell proliferation, and mineralization were assessed. RESULTS: Ad5BMP-2 and Ad5BMP-7 are successfully transduced to W20-17 cells, and secreted BMPs stimulated cell differentiation. Also, C2C12 cells transduced with Ad5BMPs showed expression of BMPs and increased ALPaseactivity. In all groups, cell proliferation was observed over times. At 7days, cells co-transduced with Ad5BMP-2 and Ad5BMP-7 showed lower proliferation than the others. C2C12 cells co-transduced with Ad5BMP-2 and Ad5BMP-7 had greater ALPaseactivity than that would be predicted if effect of individual Ad5BMPs were additive. Little mineralized nodule formation was detected in cells transduced with individual Ad5BMPs. In contrast, Ad5BMP-2 and Ad5BMP-7 combination stimulated mineralization after culturing for 10 days in mineralizing medium. CONCLUSIONS: Present study demonstrated that adenoviruses expressing BMPs gene successfully produced BMPs protein and these BMPs stimulated cells to be differentiated into osteoblastic cells. In addition, the osteogenic activity of Ad5BMPs can be synergistically increased by co-transduction of cells with Ad5BMP-2 and Ad5BMP-7.
Adenoviridae ; Adenoviruses, Human ; Alkaline Phosphatase ; Animals ; Bone Morphogenetic Protein 2 ; Bone Morphogenetic Protein 7 ; Bone Morphogenetic Proteins ; Cell Differentiation ; Cell Proliferation ; DNA, Complementary ; Durapatite ; Genetic Therapy ; Homologous Recombination ; Mice ; Osteoblasts ; Osteogenesis ; Regeneration ; Stromal Cells ; Viruses

PURPOSE: This study was performed to investigate the epidemiologic characteristics of human bocavirus (HBoV)-associated lower respiratory tract infections (LRTIs) in children. METHODS: Nasopharyngeal aspirate samples were obtained from 658 children who had been hospitalized for LRTIs in Seoul National University (SNU) Children's Hospital and SNU Bundang Hospital from March 2000 to September 2005. Multiplex RT-PCR was performed to detect 11 respiratory viruses including respiratory syncytial virus, adenovirus, rhinovirus, parainfluenza viruses 1 and 3, influenza viruses A and B, human metapneumovirus, HBoV, human coronavirus (HCoV) OC43/229E, and HCoV-NL63. Clinical data were reviewed retrospectively. RESULTS: Overall, respiratory viruses were detected in 325 (49.4%) among 658 patients. HBoV was detected in 62 cases (9.4%) and was responsible for 19.1% of virus-positive cases. HBoV was prevalent among infants and young children aged from 3 months to 5 years with the mean age of 25.3 months. Co-detection of HBoV and other respiratory viruses was observed in 37.1% which is significantly higher than average co-detection rate (12.3%) among overall virus-positive cases (P=0.000). HBoV was identified mainly in late spring and early summer from May to July. CONCLUSION: This study describes epidemiologic features of HBoV in Korean children compared with those associated with other respiratory viruses. HBoV was prevalent among LRTIs in childhood, especially in late spring and early summer season in Korea.
Adenoviridae ; Aged ; Bronchiolitis ; Child ; Coronavirus ; Coronavirus NL63, Human ; Human bocavirus ; Humans ; Infant ; Korea ; Metapneumovirus ; Orthomyxoviridae ; Paramyxoviridae Infections ; Pneumonia ; Respiratory Syncytial Viruses ; Respiratory Tract Infections ; Rhinovirus ; Seasons