Research progress on vaccines against human respiratory syncytial virus / 中国生物制品学杂志

@#Human respiratory syncytial virus(hRSV)is one of the main pathogens that cause lower respiratory tract infection in infants and the elderly.hRSV genome contains 10 genes with a full length of 15 222 bp,encoding 11 proteins(9structural proteins and 2 non-structural proteins).Different proteins play different roles in the pathogenesis of hRSV.With the in-depth research on the biological and structural characteristics of hRSV,various types of hRSV vaccines have been developed,making rapid progress.For example,hRSV attenuated live vaccine hRSV ?NS2/?1313/I1314L has entered Phase II clinical trial,and hRSV subunit protein vaccine Pre-F-GCN4t has entered Phase III clinical trial.In this paper,the biological characteristics of hRSV and the types of hRSV vaccines with rapid progress are reviewed so as to provide a reference for the development of hRSV vaccines in China.

In silico analysis of amino acid variation in human respiratory syncytial virus: insights into immunodiagnostics

BACKGROUND The highly contagious nature of human respiratory syncytial virus (HRSV) and the gravity of its infection in newborns and vulnerable adults pose a serious public health problem. Thus, a rapid and sensitive diagnostic test for viral detection that can be implemented upon the first appearance of symptoms is needed. The genetic variation of the virus must be considered for immunodiagnostic purposes. OBJECTIVES To analyse HRSV genetic variation and discuss the possible consequences for capture immunoassay development. METHODS We performed a wide analysis of N, F and G protein variation based on the HRSV sequences currently available in the GenBank database. We also evaluated their similarity with homologous proteins from other viruses. FINDINGS The mean amino acid divergences for the N, F, and G proteins between HRSV-A and HRSV-B were determined to be approximately 4%, 10% and 47%, respectively. Due to their high conservation, assays based on the full-length N and F proteins may not distinguish HRSV from human metapneumovirus and other Mononegavirales viruses, and the full-length G protein would most likely produce false negative results due to its high divergence. MAIN CONCLUSIONS We have identified specific regions in each of these three proteins that have higher potential to produce specific results, and their combined utilisation should be considered for immunoassay development.

Characteristics of Respiratory Syncytial Virus isolated from Acute Respiratory Infectious Disease in Busan

Human respiratory syncytial virus (HRSV) is known as the leading cause of respiratory tract illness in infancy and elderly children worldwide. We investigate the prevalence pattern and genetic characteristics in the second variable region G protein gene of HRSV during 5 consecutive seasons from 2010 to 2015. A total of 4,793 specimens (throat swabs) were collected from patients with acute respiratory tract. HRSV were evaluated and classified as HRSV A (n=111) or HRSV B (n=64) by real-time RT-PCR or RT-PCR. In general HRSV were detected in winter season. Coughing, fever, rhinorrhea and sputum were confirmed main symptoms in patients with HRSV. There were no significant differences in clinical characteristics or severity according to the HRSV subgroup infections. Out of 175 HRSV positive samples, 94 samples were successfully sequenced using G gene. Phylogenetic analysis revealed that 62 HRSV-A strains clustered into genotypes ON1 (n=54, 87.1%), NA1 (n=7), NA2 (n=1) and 32 HRSV-B strains clustered into three genotypes: BA4 (n=28, 87.5%), BA5 (n=2), BA6 (n=2). These results provide a better understanding of HRSV prevalence pattern and genetic characteristics.

Genetic variability in G2 and F2 region between biological clones of human respiratory syncytial virus with or without host immune selection pressure

Human respiratory syncytial virus (HRSV) is an important respiratory pathogens among children between zero-five years old. Host immunity and viral genetic variability are important factors that can make vaccine production difficult. In this work, differences between biological clones of HRSV were detected in clinical samples in the absence and presence of serum collected from children in the convalescent phase of the illness and from their biological mothers. Viral clones were selected by plaque assay in the absence and presence of serum and nucleotide sequences of the G2 and F2 genes of HRSV biological clones were compared. One non-synonymous mutation was found in the F gene (Ile5Asn) in one clone of an HRSV-B sample and one non-synonymous mutation was found in the G gene (Ser291Pro) in four clones of the same HRSV-B sample. Only one of these clones was obtained after treatment with the child's serum. In addition, some synonymous mutations were determined in two clones of the HRSV-A samples. In conclusion, it is possible that minor sequences could be selected by host antibodies contributing to the HRSV evolutionary process, hampering the development of an effective vaccine, since we verify the same codon alteration in absence and presence of human sera in individual clones of BR-85 sample.

Seasonality of viral respiratory infections in Southeast of Brazil: the influence of temperature and air humidity

Viruses are the major cause of lower respiratory tract infections in childhood and the main viruses involved are Human Respiratory Syncytial Virus (HRSV), Human Metapneumovirus (HMPV), Influenzavirus A and B (FLUA and FLUB), Human Parainfluenza Virus 1, 2 and 3 (HPIV1, 2 and 3) and Human Rhinovirus (HRV). The purposes of this study were to detect respiratory viruses in hospitalized children younger than six years and identify the influence of temperature and relative air humidity on the detected viruses. Samples of nasopharyngeal washes were collected from hospitalized children between May/2004 and September/2005. Methods of viral detection were RT-PCR, PCR and HRV amplicons were confirmed by hybridization. Results showed 54% (148/272) of viral positivity. HRSV was detected in 29% (79/272) of the samples; HRV in 23.1% (63/272); HPIV3 in 5.1% (14/272); HMPV in 3.3% (9/272); HPIV1 in 2.9% (8/272); FLUB in 1.4% (4/272), FLUA in 1.1% (3/272), and HPIV2 in 0.3% (1/272). The highest detection rates occurred mainly in the spring 2004 and in the autumn 2005. It was observed that viral respiratory infections tend to increase as the relative air humidity decreases, showing significant association with monthly averages of minimal temperature and minimal relative air humidity. In conclusion, viral respiratory infections vary according to temperature and relative air humidity and viral respiratory infections present major incidences it coldest and driest periods.

Seasonality of viral respiratory infections in Southeast of Brazil: the influence of temperature and air humidity

Viruses are the major cause of lower respiratory tract infections in childhood and the main viruses involved are Human Respiratory Syncytial Virus (HRSV), Human Metapneumovirus (HMPV), Influenzavirus A and B (FLUA and FLUB), Human Parainfluenza Virus 1, 2 and 3 (HPIV1, 2 and 3) and Human Rhinovirus (HRV). The purposes of this study were to detect respiratory viruses in hospitalized children younger than six years and identify the influence of temperature and relative air humidity on the detected viruses. Samples of nasopharyngeal washes were collected from hospitalized children between May/2004 and September/2005. Methods of viral detection were RT-PCR, PCR and HRV amplicons were confirmed by hybridization. Results showed 54% (148/272) of viral positivity. HRSV was detected in 29% (79/272) of the samples; HRV in 23.1% (63/272); HPIV3 in 5.1% (14/272); HMPV in 3.3% (9/272); HPIV1 in 2.9% (8/272); FLUB in 1.4% (4/272), FLUA in 1.1% (3/272), and HPIV2 in 0.3% (1/272). The highest detection rates occurred mainly in the spring 2004 and in the autumn 2005. It was observed that viral respiratory infections tend to increase as the relative air humidity decreases, showing significant association with monthly averages of minimal temperature and minimal relative air humidity. In conclusion, viral respiratory infections vary according to temperature and relative air humidity and viral respiratory infections present major incidences it coldest and driest periods.

Performance of indirect immunofluorescence assay, immunochromatography assay and reverse transcription-polymerase chain reaction for detecting human respiratory syncytial virus in nasopharyngeal aspirate samples

Comparison of the use of indirect immunofluorescence assay (IFA), immunochromatography assay (ICA-BD) and reverse transcription-polymerase chain reaction (RT-PCR) for detecting human respiratory syncytial virus (HRSV) in 306 nasopharyngeal aspirates samples (NPA) was performed in order to assess their analytical performance. By comparing the results obtained using ICA-BD with those using IFA, we found relative indices of 85.0 percent for sensitivity and 91.2 percent for specificity, and the positive (PPV) and negative (NPV) predictive values were 85.0 percent and 91.2 percent, respectively. The relative indices for sensitivity and specificity as well as the PPV and NPV for RT-PCR were 98.0 percent, 89.0 percent, 84.0 percent and 99.0 percent, respectively, when compared to the results of IFA. In addition, comparison of the results of ICA-BD and those of RT-PCR yielded relative indices of 79.5 percent for sensitivity and 95.4 percent for specificity, as well as PPV and NPV of 92.9 percent and 86.0 percent, respectively. Although RT-PCR has shown the best performance, the substantial agreement between the ICA-BD and IFA results suggests that ICA-BD, also in addition to being a rapid and facile assay, could be suitable as an alternative diagnostic screening for HRSV infection in children.

Vigilância de oito vírus respiratórios em amostras clínicas de pacientes pediátricos no sudeste do Brasil / Surveillance of eight respiratory viruses in clinical samples of pediatric patients in Southeast Brazil

OBJETIVO: Detecção de oito vírus respiratórios mais comuns: vírus respiratório sincicial humano (VRSH), vírus influenza tipo A e B (IA e IB), vírus da parainfluenza 1, 2 e 3 (VPIH1, 2 e 3), adenovírus (Ad) e metapneumovírus humano (MPVH), a fim de estabelecer a etiologia das infecções respiratórias agudas (IRA) e a epidemiologia desses vírus em crianças pequenas atendidas no Hospital Universitário da Universidade de São Paulo, em São Paulo, Brasil, durante o ano de 2003. MÉTODOS: A vigilância epidemiológica foi realizada em todas as crianças menores de 5 anos hospitalizadas por causa de doenças do trato respiratório inferior (DTRI) entre 1º de janeiro de 2003 e 20 de dezembro de 2003, no hospital universitário. Amostras coletadas de nasofaringe foram analisadas quanto à presença de vírus respiratórios através da reação em cadeia da polimerase e detectadas pelo programa GeneScan. RESULTADOS: Das 336 amostras coletadas, 187 (55,6 por cento) foram positivas para pelo menos um dos vírus respiratórios estudados. De todas as crianças, o VRSH foi identificado em 24,1 por cento, o MPVH em 17,8 por cento, o VPIH3 em 8,3 por cento, o Ad em 6,8 por cento, o IA em 5 por cento, o VPIH1 em 0,6 por cento, sendo que nenhum vírus foi detectado em 44,1 por cento. Infecções virais duplas foram detectadas em 7,1 por cento de todas as amostras (12,8 por cento das amostras positivas). O VPIH2 e o IB não foram detectados no presente estudo. CONCLUSÕES: Este estudo confirma que as crianças menores de 5 anos, e especialmente aquelas menores de 1 ano, apresentam uma alta taxa de hospitalização devido aos seguintes vírus: VRSH, MPVH, VPIH, influenza e adenovírus. Foi possível determinar a etiologia e epidemiologia da maioria das IRAs e traçar o perfil de sazonalidade dos vírus respiratórios mais comuns entre as crianças pequenas.

OBJECTIVE: Detection of the eight most common respiratory viruses: Human respiratory syncytial virus (HRSV), influenza virus A and B (IA and IB), parainfluenza viruses 1, 2 and 3 (HPIV1, 2 and 3), adenovirus (Ad) and human metapneumovirus (HMPV), in order to establish the etiology of acute respiratory infections (ARIs) and the epidemiology of these viruses in young children seen at Hospital Universitário, Universidade de São Paulo, in São Paulo, Brazil, during 2003. METHODS: The epidemiological surveillance was conducted in all children younger than 5 years hospitalized at the Hospital for lower respiratory tract infections (LRTI) from January 1, 2003 to December 30, 2003. Nasal and throat samples were scanned for respiratory viruses by polymerase chain reaction and detected by the GeneScan assay. RESULTS: Of 336 samples collected from 336 patients, 187 (55.6 percent) were positive for at least one of the respiratory viruses studied. Of all the children, HRSV was identified in 24.1 percent, HMPV in 17.8 percent, HPIV3 in 8.3 percent, Ad in 6.8 percent, IA in 5 percent, HPIV1 in 0.6 percent, but no virus could be detected in 44.1 percent. Dual virus infections were detected in 7.1 percent of all samples (12.8 percent of positive samples). HPIV2 and IB were not detected in the present study. CONCLUSIONS: This study confirms that children younger than 5 years and particularly younger than 1 year have a high hospitalization rate due to HRSV, HMPV, HPIV, influenza and adenovirus. We were able to determine the etiology and epidemiology of most ARIs and trace the seasonal profile of the commonest respiratory viruses among young children.

Construction of adenoviral vectors expressing F and G glycoproteins of human respiratory syncytial virus (HRSV)

Human Respiratory Syncytial Virus (HRSV) was first characterized in 1957 and has since been recognized as the most common viral cause of severe respiratory tract infection in young infants worldwide. Despite many years of research there is still no effective treatment or any immediate prospect of a vaccine. The HRSV genome is composed of single stranded negative sense RNA and the virion consists of a nucleocapsid packaged within a lipid envelope. The envelope contains spike-like projections, each being a homo-oligomer of one of three transmembrane viral envelope proteins: the attachment protein G, the fusion protein F involved in viral penetration and the small hydrofobic protein SH. The aim of this work was to construct two recombinant replication-defective adenoviruses carrying separately F and G genes from HRSV. This system was chosen because adenovirus delivers genes into target cells with high efficiency in a variety of cell lines and can be used in vitro and in vivo. In order to obtain the recombinant viruses, we did RT-PCR of RNA extracted from the HRSV A2 strain, the genes F and G were cloned in to pAdeno-X vectors. pAdeno-F and pAdeno-G were transfected in HEK-293 cells for the production of recombinant viruses, that expressed efficiently these two proteins and provide us the means for doing functional assays and immunization tests.

O Vírus Sincicial Respiratório Humano (HRSV) foi isolado e caracterizado pela primeira vez em 1957 e é considerado como o patógeno viral mais freqüente do trato respiratório de bebês e crianças. Apesar de muitos anos de pesquisa, não há ainda um tratamento específico ou uma vacina licenciada. Seu genoma é composto por uma fita simples de RNA polaridade negativa e o vírion consiste em um nucleocapsídeo empacotado por um envelope lipídico. O envelope contém projeções, chamadas espículas, constituídas de homoligômeros de uma das 3 glicoproteínas de membrana: a proteína de ligação G ("attachment"), a proteína de fusão F ("fusion") e a proteína SH ("small hydrofobic"). O objetivo deste trabalho foi construir dois adenovirus recombinantes defectivos em replicação expressando separadamente os genes F e G do HRSV. Este sistema foi escolhido porque os vetores adenovirais possuem a capacidade de inserir genes em uma grande variedade de linhagens celulares in vitro e in vivo. Para obtenção destes vetores adenovirais, um RT-PCR de RNA extraído do protótipo A2 de HRSV foi feito e os genes F e G clonados em vetores pAdeno-X. pAdeno-F e pAdeno-G foram transfectados em células HEK-293 para a produção do vírus recombinante, que expressaram corretamente essas duas proteínas constituem-se ferramentas para imunização e estudos funcionais.