Identification of dual receptor-binding specific strains of human H5N1 viruses in China.

Objective: Both the 2, 6 linkage and its topology on target cells are critical for the recognition by human influenza virus. The binding preference of avian flu virus H5N1 HA to the 2, 3-linked sialylated glycans is considered the major factor limiting its efficient infection and transmission in humans. To monitor potential adaptation of H5N1 virus in human population, the surveillance of receptor-binding specificity was undertaken in China.

[The proliferation of H1N1 subtype influenza viruses in A549 and BEAS-2B cells].

Objective: Analyze the proliferation of different host H1N1 subtype influenza viruses in A549 and BEAS-2B cells.

Methods: Human, avain and swine three hosts of the H1N1 influenza viruses infected A549 and BEAS-2B cells and analyze the characteristics of different periods after inocubation. Determine the receptor binding specificity of influenza virus by hemagglutination (HA) test with RBCs with two types of receptor.

[The study of multiple RT-PCR-based reverse dot blot hybridization technique for detecting influenza viruses].

Objective: To establish a multiplex RT-PCR-based reverse dot blot hybridization technique to detect influenza viruses.

Methods: Obtain the HA nucleotide sequences of seasonal influenza H1N1, seasonal influenza H3N2, influenza H1N1 and human avian influenza H5N1 from GenBank. Design primers in conservative district and probes t in high variable region respectively, after analyzing the HA nucleotide sequences of influenza virus through the Vector NTI 9.

[Expression and purification of avian influenza virus H5N1 NP protein, and screening interaction proteins of host cells in vitro].

Objective: To express and purify H5N1 influenza virus (A/Anhui/1/2005) NP in prokaryotic system and to explore the NP-interacting proteins of human bronchial epithelial cells BEAS-2B in vitro.

Methods: The full length H5N1 NP gene fragment was amplified by PCR, inserted into prokaryotic expression vector (pET30a) to generate NP expression plasmid pET30a-NP. After transforming pET30a-NP into E.

[The virus isolation analysis of the H5N1 subtype human avain influenza cases in mainland China from 2005 to 2009].

Objective: To analyse the correlation between the virus isolation and the specimen collection of the H5N1 human high pathogenic avain influenza cases in Mainland China.

Methods: The specimens were collected in Mainland China from 2005.10 to 2009.

[Characterization of pseudotyped viruses coated with hemagglutinin of H5N1 avian influenza].

Objective: To construct pseudovirus bearing H5N1 HA based on a lentivirus vector system. Then we study the biological feature of the pseudovirus. With the newly established viral particles, we performed the serological tests.

Visual detection of pandemic influenza A H1N1 Virus 2009 by reverse-transcription loop-mediated isothermal amplification with hydroxynaphthol blue dye.

A sensitive reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for rapid visual detection of pandemic influenza A H1N1 virus infection. The reaction was performed in one step in a single tube at 65 degrees C for 60 min with the addition of hydroxynaphthol blue (HNB) dye prior to amplification. The detection limit of the RT-LAMP assay was approximately 60 copies, and no cross-detection was observed.

[Laboratory confirmation of the first influenza A (H1N1) imported case in Mainland China].

The clinical throat swab specimen of an imported suspected case of influenza A (H1N1) was detec ted with real-time PCR, RT-PCR and subsequently confirmed by gene sequencing. The presence of influ enza A (H1N1) virus confirmed the first case with A (H1N1) infection in Mainland China.

[Establishment of a method for rapid detection of the nucleic acid of the novel A (H1N1) influenza virus].

A new flu caused by a novel influenza A(H1N1) virus has spread over the United States, Mexico and more than 40 other countries. And because of the immediate global concern, WHO has announced that the current level of influenza pandemic alert is raised to phase 5, indicating approaching of an influenza pandemic. As patients suffering from the influenza A (H1N1) have the similar symptoms as patients with seasonal influenza, differential detection and identification of the influenza virus have to depend on specific laboratory tests.

[Construction and characterization of reverse genetic system for H9N2 avian influenza].

Objective: To provide a technology platform for vaccine development as well as the research on transmission and pathogenesis, the reverse genetic system for H9N2 avian influenza virus was established.

Methods: Eight full-length cDNAs of avian influenza virus A/Guangzhou/333/99 (H9N2) were amplified by RT-PCR and separately cloned into the transcription/expression vector, pCI-polI. The 8 plasmids DNA was cotransfected into 293T cell, the cell supernatant was collected and inoculated into embryonated eggs, the rescued virus from the allantoic fluid was identified by hemagglutinination assay.

[Detection of influenza viruses/avian influenza viruses and identification of virulence using a microarray].

Objective: To establish the DNA microarray to detect influenza viruses and avian influenza viruses, and identify their virulence.

Methods: Hemagglutinin (HA), neuramidinase (NA) and nucleoprotein(NP) genes were chosen simultaneously as targets for designing a microarray used for detection of viruses and identification virulence. The nucleic acid were amplified by single primer amplication (SPA).

[Laboratory diagnosis and molecular characterization of highly pathogenic avian influenza virus (H5N1) in human in Hunan Province in 2005-2006].

To determine the etiologic agents of two atypical pneumonia human cases in Hunan Province in 2005-2006 and to study their pathogenic potential, the patients' respiratory tract samples and sera were collected. The respiratory tract samples were tested by real-time RT-PCR and RT-PCR methods, and the sera by hemagglutination-inhibition assay. Virus was isolated from case 2 and its genome was sequenced and analyzed.

[The firstly confirmed pregnant woman case of avian influenza A (H5N1) by etiological research in China].

To investigate the cause of death of a pregnant woman with undefined pneumonia reported from the People's Hospital of Tongling City in Anhui Province on November 8, 2005, the patient's tracheal aspirates and serum samples were collected and tested by RT-PCR and Real-time PCR to detect viral nucleic acids of HA of A/H5N1, A/H7N7, A/H9N1 and A/M. Tracheal aspirates were inoculated into special pathogen free (SPF) embryonated eggs for cultivation and identification of virus. The HA gene of the virus was sequenced and analyzed.

[Characteristic analysis of NA gene of human influenza viruses (H3N2) isolated from 1996 to 2005 in China].

The NA genes of 395 strains of human H3N2 influenza virus isolated from 1996 to 2005 in China were sequenced, analyzed with bioinformatics tools. The NA nucleotide sequence of phylogenetic tree showed a main evolution branch with multiple short side branches. The strains in the same year may be divided into several branches.

[Study on the correlation of human influenza A/H3N2 hemagglutinin gene variation and the epidemic from 1995 to 2005 in China].

To study the correlation of human influenza A/H3N2 hemagglutinin gene variation and the epidemic from 1995 to 2005 in China, 550 HA1 sequences of H3N2 viruses isolated in China were analyzed with phylogenetic tree. The results showed that the evolution of HA1 represented a long trunk with short side branches. The animo acid changes of HA1 mostly located at the antigenic sites or aside of them, but also may occur at the other sites simultaneously.

[Antigenic and genetic study of influenza virus (H1N1) circulated in China in 2004-2005].

Background: To analyze the genetic and antigenic characteristics of hemagglutinin (HA) gene of human influenza (H1N1) virus isolated from the mainland of China since 2004 to 2005.

Methods: The single-way hemagglutination inhibition (HI) tests were used to test the antigenic characteristics, and the HA1 gene was sequenced based on the antigenic results.

Results: The single-way HI results showed that no virus isolates had 4-folds greater HI titer compared with A/Shanghai/1/1999 (H1N1) in 2004, but there was 6.

[Development of methods for detection of H5N1 from human clinical specimens].

Background: To establish a method for H5N1 RNA detection and laboratory diagnosis of suspected human avian influenza (H5N1) virus infected cases.

Methods: The typing and sub-typing primers were designed according to M and H5 and N1 gene respectively, and the RT-PCR and real-time PCR were developed using these primers.

Results: The RT-PCR and real-time PCR could be used for H5N1 detection specifically, and there was no cross reaction with other influenza subtypes such as H1 and H3.

[Adamantane resistance among influenza A (H3N2) viruses isolated from the mainland of China].

Background: To study the incidence of adamantane resistance among influenza A (H3N2) viruses isolated from the mainland of China since 1989 through our influenza surveillance system, and to provide more information for the clinical usage of adamantane drugs.

Methods: Totally 584 influenza A (H3N2) virus strains were randomly selected from our surveillance network since 1989, the adamantane drug resistance related gene M2 of all 584 strains was sequenced, and the drug sensitivity of viruses was also assayed by using biological methods in cells.

Results: No adamantane resistant strains were detected among the strains isolated from 1989 to 1999, but there was a surprisingly increased resistance rate of 56% in 2003 compared with 3.

[Establishment of molecular differential diagnostic (MDD) technology for detection of common respiratory viruses].

Background: To provide rapid laboratory evidence for diagnosis of respiratory infection and help diagnose accurately and reduce the spread of disease, so that the patients can be diagnosed and treated early.

Methods: Thirteen kinds of respiratory viruses were detected by using Genaco's MDD technology.

Results: All the specimens were detected, the total positive rate was 100%; the sensitivity of the method was 10e2 (pfu/ml).

[Antigenic and genetic study of influenza virus B circulated in China in 2004-2005].

Background: To analyze the genetic and antigenic characteristics of hemagglutinin (HA) gene of human influenza B virus isolated from the mainland of China since 2004-2005.

Methods: The single-way hemagglutinin inhibition (HI) tests were used to test the antigenic characteristics, and the HA1 gene was sequenced based on the antigenic results.

Results: The Yamagata-like and Victoria-like viruses co-circulated in 2004-2005.

[Analysis of human H5N1 virus hemagglutinin gene isolated from the mainland of China].

Background: To analyze the genetic and antigenic characteristics of human H5N1 virus isolated from the mainland of China.

Methods: The hemagglutinin (HA) gene of human H5N1 virus were sequenced and analyzed.

Results: The results of HA gene sequencing showed that all the virus isolates belong to the same group because of the high similarity, but they were different from the virus isolated from Thailand and Vietnam.

[Clinical characteristic analysis of the first human case infected by influenza A (H5N1) in Jiangxi Province].

Objective: To describe the clinical, laboratory and radiological presentation of a human case infected by influenza A (H5N1), and to understand its management and prognosis.

Methods: The clinical and autopsy data of the first human case infected by influenza A (H5N1) in Jiangxi Province were collected and analyzed.

Results: The first case infected by influenza A (H5N1) in Jiangxi Province was confirmed by laboratory findings with reverse transcription-polymerase chain reaction (RT-PCR) and influenza A (H5N1) isolation.