In July 2009, some farms of breeding Muscovy ducks on the peak of egg laying suffered the decrease of hatching rate and the quality of the eggs showing low mortality and no evident respiratory symptoms. The swelling and congestive ovary was visible after autopsy. This study was brought out for the diagnosis of these cases. The virus was isolated and identified by the methods of virus culture in chicken embryo, physical and chemical properties test, hemagglutinin test, NDV (Newcastle diseases Virus) interference test, electron microscope observation, pathogenicity test and the gene sequence analysis. The results indicated the virus showed the characters of inducing dwarf embryo after inocubation, the sensibility to lipid solvent and the hemagglutination capacity after pancreatic enzyme treatment, the typical morphology of coronavirus, the interference to NDV replication and the homology among 84.7% - 99% of the particial N gene sequences to the reference IBV (Avian infectious bronchitis virus) strains. The strain was identified as IBV isolate and this study confirmed the pathogenicity of IBV to Muscovy ducks.
Amino Acid Sequence ; Animals ; Chick Embryo ; Coronavirus Infections ; veterinary ; virology ; Ducks ; virology ; Female ; Infectious bronchitis virus ; classification ; genetics ; isolation & purification ; Molecular Sequence Data ; Phylogeny ; Poultry Diseases ; virology ; Sequence Alignment

We wished to ascertain the prevalence as well as the genetic and antigenic variation of infectious bronchitis viruses (IBVs) circulating in the Guangxi Province of China in recent years. The S1 gene of 15 IBV field isolates during 2012-2013 underwent analyses in terms of the similarity of amino-acid sequences, creation of phylogenetic trees, recombination, and serologic identification. Similarities in amino-acid sequences among the 15 isolates of the S1 gene were 54.3%-99.6%, and 43.3%-99.3% among 15 isolates and reference strains. Compared with the vaccine strain H120, except for GX-YL130025, the other 14 isolates showed a lower similarity of amino-acid sequences of the S1 gene (65.1-81.4%). Phylogenetic analyses of the S1 gene suggested that 15 IBV isolates were classified into eight genotypes, with the predominant genotype being new-type II. Recombination analyses demonstrated that the S1 gene of the GX-NN130048 isolate originated from recombination events between vaccine strain 4/91 and a LX4-like isolate. Serotyping results suggested that seven serotypes prevailed during 2012-2013 in Guangxi Province, and that only one isolate was consistent with the vaccine strain H120 in serotype (which has been used widely in recent years). The serotype of recombinant isolate GX-NN130048 was different from those of its parent strains. These results suggested that not only the genotype, but also the serotype of IBV field isolates in Guangxi Province had distinct variations, and that increasing numbers of genotypes and serotypes are in circulation. We showed that recombination events can lead to the emergence of new serotypes. Our study provides new evidence for understanding of the molecular mechanisms of IBV variations, and the development of new vaccines against IBVs.
Animals ; Antibodies, Viral ; blood ; Chickens ; China ; Coronavirus Infections ; blood ; veterinary ; virology ; Genetic Variation ; Genotype ; Infectious bronchitis virus ; classification ; genetics ; immunology ; isolation & purification ; Molecular Sequence Data ; Phylogeny ; Poultry Diseases ; blood ; virology ; Sequence Homology, Amino Acid ; Spike Glycoprotein, Coronavirus ; chemistry ; genetics ; immunology

In order to investigate the prevalence and track genetic and antigenic evolutions of infectious bronchitis virus (IBV) and their prevalence in Guangxi, China since 1985, gene amplification and sequencing and virus neutralization (VN) test on chicken embryo tracheal organ cultures were used in genotyping and serotyping of 28 IBV isolates during 2009-2011 in Guangxi. The results of N gene sequencing and comparison showed that the 28 isolates and reference strains were classified into three groups, and most isolates belonged to group Ill, while the isolates in 1985-2008 belonged to groups IV and II. The data of VN test indicated that the 28 isolates belonged to 6 serotypes; among them, 71. 4% belonged to serotypes 1, 2, and 3, and 11 (39.3%) shared the same serotype with the current vaccine strains. Given the data of our previous study, it is found that prevalent serotypes and their proportions varied in different areas of Guangxi and during different periods. These data lay a good foundation for developing an oil-emulsified inactivated polyvalent vaccine containing local dominant serotypes for the effective prevention and control of infectious bronchitis.
Animals ; Antibodies, Viral ; immunology ; Chick Embryo ; Chickens ; China ; epidemiology ; Coronavirus Infections ; epidemiology ; immunology ; veterinary ; virology ; Infectious bronchitis virus ; classification ; genetics ; immunology ; isolation & purification ; Molecular Sequence Data ; Phylogeny ; Poultry Diseases ; epidemiology ; immunology ; virology

OBJECTIVETo analyze the diagnosis and treatment characteristics of patients with severe Influenza A.

METHODA retrospective investigation on the clinical manifestation, chest radiography, electronic fiber bronchoscopy and the histology of the cast, rescue course and outcome was conducted in 15 children with severe influenza A during January to May of 2013.

RESULTEleven cases were male, the range of age was 2 to 6 years; 5 cases were female, the range of age was 1 month to 6 years, accouting for 4.2% of hospitalized children with influenza. Three patients had an underlying chronic disease, two had nephrotic syndrome, and one had congenital heart disease. All the 15 cases were diagnosed as severe influenza A virus infection complicated with pneumonia and respiratory failure, of whom 10 cases were infected with H1N1 virus , the other 5 cases could not be identified as H1N1 virus by using H1N1 kit, but none of the 15 cases were infected with H7N9 virus. Of 15 cases, 8 had atelectasis, 4 had pneumothorax, 3 had pneumomediastinum, 4 had pleural effusion, 1 had pneumorrhagia; 12 patients required mechanical ventilation. 1 only required noninvasive mask CPAP, 2 did not require assisted ventilation, they were just given mask oxygen. Seven cases' sputum culture showed combined infection with bacteria and fungi, sputum smear examination detected: G(+) cocci in 2 cases, and G(-) bacilli in the other 2. By using electronic fiber bronchoscopy, bronchial cast was detected in 5 patiens. Histological examination of the bronchial cast revealed a fibrinous exudation containing large quantity of eosinophils, neutrophils in 1 patients, fibrinous exudation and necrotic material containing large quantity of neutrophils in 4 patients. After the bronchial casts were removed, 4 patients were improved greatly. All patients were treated with postural drainage of left and right side position, massage of electric oscillation, strengthening the sputum suction aiming to improve pulmonary ventilation function. Three patients died: 1 case was compliicated with nephrotic syndrome, another case had congenital heart disease, and 1 case hads pneumorrhagia, renal failure and multiple organ dysfunction syndrome (MODS).

CONCLUSIONThe mortality of severe Influenza A is higher if it is complicated with underlying chronic diseases. In children undergoing rapid and progressive respiratory distress with lung atelectasis, consolidation or emphysema on chest X-ray, plastic bronchitis should be considered. Electronic fiber bronchoscopy should be performed early Lung physicotherapeutics still are important assistant measures for improving the pulmonary ventilation function.

Antiviral Agents ; therapeutic use ; Bronchitis ; diagnosis ; therapy ; virology ; Bronchoscopy ; methods ; Child ; Child, Preschool ; Female ; Humans ; Infant ; Influenza A Virus, H1N1 Subtype ; Influenza, Human ; diagnosis ; mortality ; therapy ; Intensive Care Units ; Intubation, Intratracheal ; Male ; Oxygen Inhalation Therapy ; Pneumonia, Viral ; diagnosis ; therapy ; Pulmonary Atelectasis ; diagnosis ; therapy ; virology ; Rare Diseases ; Respiration, Artificial ; Retrospective Studies ; Sputum ; microbiology ; Treatment Outcome

To explore the expression potential of heterogeneous genes using the backbone of infectious bronchitis virus (IBV) Beaudette strain, the ectodomain region of the Spike gene (1,302 bp) of IBV H120 strain was amplified by RT-PCR and replaced into the corresponding location of the IBV Beaudette strain full-length cDNA. This recombinant was designated as BeauR-H120(S1). BeauR-H120(S1) was directly used as the DNA template for the transcription of viral genomic RNA in vitro. Then, the transcription product was transfected into Vero cells by electroporation. At 48 h post-transfection, the transfected Vero cells were harvested, and passaging continued. A syncytium was not observed until the recombinant virus had passed through four passages. The presence of rBeau-H120(S1) was verified by the detection of the replaced ectodomain region of the H120 Spike gene using RT-PCR. Western blot analysis of rBeau-H120 (S1)-infected Vero cell lysates demonstrated that the nucleocapsid (N) protein was expressed, which implied that rBeau-H120(S1) could propagate in Vero cells. The TCIDs0 and EIDs0 data demonstrated that the titer levels of rBeau-H120(S1) reached 10(590+/-0.22)TCID50/mL and 10(6.13+/-0.23)EID50/mL in Vero cells and 9-day-old SPF chicken embryos, respectively. Protection studies showed that the percentage of antibody-positive chickens, which were vaccinated with rBeau-H120(S1) at 7 days after hatching, rose to 90% at 21 days post-inoculation. Inoculation provided an 85% rate of immune protection against a challenge of the virulent IBV M41 strain (103EID50/chicken). This recombinant virus constructed using reverse genetic techniques could be further developed as a novel genetic engineering vaccine against infectious bronchitis.
Animals ; Cercopithecus aethiops ; Chick Embryo ; Chickens ; Coronavirus Infections ; veterinary ; virology ; Infectious bronchitis virus ; chemistry ; genetics ; growth & development ; metabolism ; Poultry Diseases ; virology ; Protein Structure, Tertiary ; Spike Glycoprotein, Coronavirus ; chemistry ; genetics ; metabolism ; Transfection ; Vero Cells

Infectious bronchitis virus (IBV) poses a severe threat to the poultry industry and causes heavy economic losses worldwide. Vaccination is the most effective method of preventing infection and controlling the spread of IBV, but currently available inactivated and attenuated virus vaccines have some disadvantages. We developed a chimeric virus-like particle (VLP)-based candidate vaccine for IBV protection. The chimeric VLP was composed of matrix 1 protein from avian influenza H5N1 virus and a fusion protein neuraminidase (NA)/spike 1 (S1) that was generated by fusing IBV S1 protein to the cytoplasmic and transmembrane domains of NA protein of avian influenza H5N1 virus. The chimeric VLPs elicited significantly higher S1-specific antibody responses in intramuscularly immunized mice and chickens than inactivated IBV viruses. Furthermore, the chimeric VLPs induced significantly higher neutralization antibody levels than inactivated H120 virus in SPF chickens. Finally, the chimeric VLPs induced significantly higher IL-4 production in mice. These results demonstrate that chimeric VLPs have the potential for use in vaccines against IBV infection.
Animals ; Antibodies, Viral/blood ; *Chickens ; Chimera/genetics/immunology ; Coronavirus Infections/prevention & control/*veterinary/virology ; Female ; *Immunity, Innate ; Infectious bronchitis virus/genetics/*immunology ; Influenza A Virus, H5N1 Subtype/genetics/immunology ; Injections, Intramuscular/veterinary ; Mice ; Mice, Inbred BALB C ; Neuraminidase/genetics ; Poultry Diseases/*prevention & control/virology ; Recombinant Fusion Proteins/genetics/immunology ; Spike Glycoprotein, Coronavirus/genetics/*immunology ; Vaccines, Synthetic/administration & dosage/genetics/immunology ; Vaccines, Virus-Like Particle/administration & dosage/genetics/*immunology ; Viral Proteins/genetics

This study aimed to understand the dynamic distribution of infectious bronchitis virus (IBV) Jin-13 strain in SPF chickens. Ninety-day-old SPF chickens were inoculated with Jin-13, a virulent strain, and dissected at day 1, 4, 7, 10, 14, 21, 28 or 35 post-inoculation (dpi). Samples of heart, liver, spleen, lung, trachea, kidney and duodenum were collected and the N gene was detected by Sybr Green I real-time quantitative RT-PCR assays. The established method had a good linear correlation from 7.77 x 10(8) to 10(0) copies/microL. SPF chickens developed typical clinical signs of IBV at the 4th dpi, and the IBV viral concentration of tissues and organs gradually increased with a peak of up to 7.13 x 10(4) copies/microL. The viral concentration of most organs decreased by the 10th dpi, but those of the kidney, trachea and lung remained positive for IBV at 28 dpi and the heart was still positive for IBV at > 35 dpi. The results of this study, showed that the Jin-13 strain can cause prolonged virus excertion in chickens with severe renal damage.
Animals ; Chickens ; Coronavirus Infections ; veterinary ; virology ; Infectious bronchitis virus ; isolation & purification ; pathogenicity ; physiology ; Lung ; virology ; Poultry Diseases ; virology ; Reverse Transcriptase Polymerase Chain Reaction ; Specific Pathogen-Free Organisms ; Trachea ; virology ; Virulence

The genome of CK/CH/SD09/005, an isolate of infectious bronchitis virus (IBV), was characterized to enable the further understanding of the epidemiology and evolution of IBV in China. Twenty-five pairs of primers were designed to amplify the full-length genome of CK/CH/SD09/005. The nucleotide sequence of CK/CH/SD09/005 was compared with reference IBV strains retrieved from GenBank. The phylogenic relationship between CK/CH/SD09/005 and the reference strains was analyzed based on S1 gene sequences. The complete genome of CK/CH/SD09/005 consisted of 27691 nucleotides (nt), excluding the 5' cap and 3' poly A tail. The whole-genome of CK/CH/SD09/005 shared 97 - 99% nucleotide sequence homology with the GX-NN09032 strain, which was the only complete genome that was closely related to CK/CH/SD09/005. When compared with all reference strains except GX-NN09032, CK/CH/SD09/005 showed the highest similarity to ck/CH/LDL/091022 and SDIB821/2012 (QX-like) in the replicase gene (Gene 1) and 3'UTR, with a sequence identity rate of 97% and 98%, respectively. However, CK/CH/SD09/005 exhibited lower levels of similarity with ck/CH/LDL/091022 and SDIB821/2012 in S-3a-3b-3c/ E-M-5a-5b-N with a sequence identity of 72% - 90%. CK/CH/SD09/005 showed the highest level of nucleotide identity with Korean strain 1011, and Chinese strains CK/CH/LXJ/02I, DK/CH/HN/ZZ2004 and YX10, in ORF 3c/E (97%), 5a (96%), 5b (99%) and N (96%), respectively. ORFs 3a, 3b and M of CK/CH/SD09/005 exhibited no more than 90% homology with the reference strains, excluding GX-NN09032. The phylogenic analysis based on the S1 gene revealed that CK/CH/SD09/005 and 39 published strains were classified into seven clades (genotypes). CK/CH/SD09/005 was distributed in clade IV with several isolates collected between 2007 and 2012. CK/CH/SD09/005 showed 66% - 69% and 72% - 81% nucleotide identities with the IBV strains of other six clades in the S1 and S2 subunits, respectively. More over, multiple substitutions were found throughout the entire S gene of CK/CH/SD09/005, while insertions and deletions were located within the S1 gene. These results indicated that CK/CH/SD09/005 is a novel variant that may be derived from the QX-like strains that are prevalent in China. Multiple genetic mechanisms, including recombinations, mutations, insertions and deletions, are likely to have contributed to the emergence of this IBV strain.
Animals ; Chickens ; China ; Coronavirus Infections ; veterinary ; virology ; Genome, Viral ; Genomics ; Infectious bronchitis virus ; classification ; genetics ; isolation & purification ; Molecular Sequence Data ; Phylogeny ; Poultry Diseases ; virology ; Sequence Homology, Amino Acid ; Viral Proteins ; chemistry ; genetics

The protective efficacy of DNA plasmids encoding avian infectious bronchitis virus (IBV) S1, N, or M protein was investigated in chickens. Chickens were inoculated monovalently (with plasmid pVAX1-16S1, pVAX1-16M, or pVAX1-16N alone) or multivalently (combination of the three different plasmids, pVAX1-16S1/M/N). A prime-boost immunization protocol against IBV was developed. Chickens were immunized with the multivalent DNA vaccine twice and then boosted with an inactivated vaccine once. Antibody titers of the chickens immunized with pVAX1-16S1/M/N were much higher than those of the monovalent groups (p < 0.01). A protective rate up to 90% was observed in the pVAX1-16S1/M/N group. The serum antibody titers in the prime-boost birds were significantly higher than those of the multivalent DNA vaccine group (p < 0.01) but not significantly different compared to the inactivated vaccine group at 49 days of age. Additionally, the prime-boost group also showed the highest level of IBV-specific cellular proliferation compared to the monovalent groups (p < 0.01) but no significant difference was found compared to the multivalent DNA vaccine group, and the prime-boost group completely protected from followed viral challenge.
Aging ; Animals ; Antibodies, Viral/blood ; Cell Proliferation ; Chickens ; Coronavirus Infections/prevention & control/*veterinary/virology ; Immunization, Secondary/veterinary ; Infectious bronchitis virus/*immunology ; Poultry Diseases/*prevention & control/virology ; T-Lymphocyte Subsets/cytology/physiology ; Vaccines, DNA/immunology ; Vaccines, Inactivated/immunology ; Viral Vaccines/*immunology

INTRODUCTIONSingapore had its first case of pandemic influenza A (H1N1) 2009 on 26 May 2009. As of 3 August 2009, 440 children with confirmed H1N1were admitted to KK Women's and Children's Hospital (KKH).

MATERIALS AND METHODSThis is a retrospective case control study of children admitted from 26 May 2009 to 19 July 2009 with H1N1infection. Cases and controls were first differentiated by whether they were complicated or non-complicated in nature, and subsequently analysed with regards to possible independent risk factors.

RESULTSWe analysed 143 admitted children; 48 cases and 95 controls (1: 2 ratio). Significant comorbidity was found in 20.3% (n = 29) of patients with the majority having asthma (n = 18, 12.6 %) followed by obesity (n = 7, 4.9%). Binary logistic regression analysis showed risk factors for complicated disease were comorbidity (adjusted OR 6.0, 95% CI, 2.5 to 14.6, P < 0.0001) and age <2 years (adjusted OR 9.8, 95% CI, 2.4 to 40, P = 0.001). Age less than 5 years was not found to be a risk factor.

CONCLUSIONIn the early stages of an evolving influenza epidemic when oseltamivir stocks are low, oseltamivir treatment for influenza can be streamlined and offered to those at highest risk who are under 2 years old or have significant comorbidity to prevent complicated disease.

Adolescent ; Age Factors ; Antiviral Agents ; therapeutic use ; Asthma ; complications ; Bronchitis ; complications ; Case-Control Studies ; Child ; Child, Preschool ; Cohort Studies ; Female ; Fluid Therapy ; Gastritis ; virology ; Gastroenteritis ; virology ; Humans ; Infant ; Influenza A Virus, H1N1 Subtype ; isolation & purification ; Influenza, Human ; complications ; Male ; Obesity ; complications ; Oseltamivir ; therapeutic use ; Pneumonia, Bacterial ; complications ; Retrospective Studies ; Risk Factors ; Seizures ; complications ; Singapore