Lipopeptide-adjuvanted respiratory syncytial virus virosomes: A safe and immunogenic non-replicating vaccine formulation

Respiratory syncytial virus (RSV) causes severe respiratory disease in children and the elderly. There is no registered RSV vaccine. Early experimental non-replicating vaccines have been found to exacerbate RSV symptoms upon infection causing enhanced respiratory disease. Here we show that immunization of mice with reconstituted virosomes produced from RSV envelopes and containing the lipopeptide adjuvant (P3CSK4), induces high-titer virus-neutralizing antibodies, and the secretion of IFN-γ through both MHC-I and MHC-II presentation of antigen, with a balanced Th1/Th2 profile. Immunization with RSV virosomes provides sterilizing immunity to virus challenge in mice and cotton rats, while not producing symptoms of enhanced disease. Therefore, these virosomes represent a promising candidate inactivated RSV vaccine formulation.     

A respiratory syncytial virus (RSV) vaccine based on parainfluenza virus 5 (PIV5)

Human respiratory syncytial virus (RSV) is a leading cause of severe respiratory disease and hospitalizations in infants and young children. It also causes significant morbidity and mortality in elderly and immune compromised individuals. No licensed vaccine currently exists. Parainfluenza virus 5 (PIV5) is a paramyxovirus that causes no known human illness and has been used as a platform for vector-based vaccine development. To evaluate the efficacy of PIV5 as a RSV vaccine vector, we generated two recombinant PIV5 viruses – one expressing the fusion (F) protein and the other expressing the attachment glycoprotein (G) of RSV strain A2 (RSV A2). The vaccine strains were used separately for single-dose vaccinations in BALB/c mice. The results showed that both vaccines induced RSV antigen-specific antibody responses, with IgG2a/IgG1 ratios similar to those seen in wild-type RSV A2 infection. After challenging the vaccinated mice with RSV A2, histopathology of lung sections showed that the vaccines did not exacerbate lung lesions relative to RSV A2-immunized mice. Importantly, both F and G vaccines induced protective immunity. Therefore, PIV5 presents an attractive platform for vector-based vaccines against RSV infection.     

Nosocomial respiratory syncytial virus infection: impact of prospective surveillance and targeted infection control

BACKGROUND: Nosocomially acquired respiratory syncytial virus (RSV) infections cause serious problems in hospitalized patients. An increased effort should be made to describe the problems connected with such infections in pediatric hospitals, with the aim of reducing the occurrence of nosocomial RSV infections (NI). METHODS: A specialized database was introduced for surveillance and a multifaceted barrier concept based on the CDC recommendations was developed for the control of NI in a university children's hospital in Germany. RESULTS: Between 1999 and 2002 (November 1-April 30), 283 RSV infections (general population) were prospectively documented. Thirty-nine cases (13.8%) were nosocomial infections (NI) with an incidence density (ID) of 0.99/1000 patient days; 48.7% of all NI were found in prematurely born infants. Following the introduction of a surveillance and prevention policy, a 9-fold decrease of the ID (1.67 vs. 0.18/1000 patient-days) was found when comparing the first and the last season. Intensive care treatment was required in 18% of all documented RSV-infections, in 48.7% of all NI cases and in 43.5% of all RSV-infected prematurely born infants. Overall RSV-related mortality was 0.71%. CONCLUSIONS: Early diagnosis, a strict cohorting and contact isolation policy, and prospective surveillance contribute to the reduction of nosocomial RSV infection.     

郑州市流感和呼吸道合胞病毒感染住院病例的抗生素使用及合理性评估

目的 描述河南省郑州市流感和呼吸道合胞病毒(respiratory syncytial virus, RSV)感染住院病例的抗生素使用情况,评估抗生素使用的合理性。方法 采用前瞻性病例系列研究,2018年11月—2019年11月在河南省儿童医院与郑州市中心医院分别纳入儿童和成人急性呼吸道感染住院病例,收集人口学和临床资料,采集呼吸道标本,使用Real-time PCR法检测流感病毒和RSV,分析流感和RSV感染病例合并细菌感染、抗生素使用及其合理性。结果 共检出669例流感和RSV感染住院病例,38.4%(252/656)合并细菌感染,13例无法判断细菌感染情况。97.9%的病例住院期间使用抗生素,第三代头孢菌素(77.0%)、大环内酯类(20.3%)、青霉素和β-内酰胺酶抑制剂复合制剂(15.5%)、喹诺酮类(9.9%)和碳青霉烯类(7.9%)为常用的种类。从入院到使用抗生素的时间间隔为1.0(1.0,1.0)d,抗生素使用持续时间为6.0(5.0,8.0)d。抗生素用药频度和总费用分别为2.4(1.3,6.0)DDDs和369.7(210.6,1 154.0)元/人。在655例使用...     

Genetic variability of human respiratory syncytial virus in Pune,Western India

Human respiratory syncytial virus (RSV) is one of the most important respiratory viruses causing acute respiratory tract infections amongst children. Based on genotyping of the attachment glycoprotein (G) gene, it is divided into two groups, RSV-A and RSV-B. Infection with one group does not confer immunity against the other and children infected with one antigenic group are more likely to be reinfected with the heterologous group. We tested 854 samples of patients with influenza like illness (ILI)/severe respiratory illness (SARI) during the period 2009–2012 for RSV using a conventional multiplex RT-PCR and found 159 (18.61%) samples to be positive for RSV of which 130 (15.22%) were positive for RSV-B and 29 (3.39%) for RSV-A suggesting that RSV-B was the predominant group circulating in Western India during the study period. Seasonal RSV outbreaks were observed in the monsoon and winter months. RSV was more prevalent amongst children in the 0–24 month age group (21.53%) in comparison to children in the 24–60 month age group (13.01%). Phylogenetic analysis using the G gene of 27 representative RSV-A positive samples revealed that all sequences belonged to the NA1 genotype. Of these, 5 sequences exhibited the novel 72 nucleotide duplication in the C-terminal of the G gene first reported from Ontario, Canada and clustered in the newly designated ON1 genotype. Also, 32 of the 33 RSV-B sequences exhibited the 60 nucleotide duplication associated with genotype BA and phylogenetic analysis showed that these sequences belonged to the genotype BA9 and BA12. We also found one RSV-B sequence belonging to genotype GB2, which has not been previously reported in India.     

2016—2019年杭州市余杭区973例严重急性呼吸道感染病例呼吸道合胞病毒检测结果

收集2016年1月至2019年12月杭州市余杭区第一人民医院的严重急性呼吸道感染病例鼻咽分泌物标本,采用荧光定量PCR方法进行RSV病毒核酸检测;检测后分析不同年龄组、不同月份RSV阳性检出情况。结果显示,共收集并检测973份鼻咽拭子标本中,63份标本RSV核酸检测阳性,阳性检出率为6.47%;核酸检出率男性大于女性,...     

Advances in RSV vaccine research and development – A global agenda

Respiratory syncytial virus (RSV) is an important cause of viral lower respiratory tract illness in infants and children globally, but no vaccine is currently available to protect these vulnerable populations. Live-attenuated vaccine approaches have been in development for decades, but achieving the appropriate balance between immunogenicity and safety has proven difficult. Immunoprophylaxis with the neutralizing monoclonal antibody palivizumab is limited to high-risk infants, but cost requirements for multiple dosing make its use impractical in low- and middle-income countries. A growing number of RSV vaccine candidates using a variety of technologies and targeting diverse populations has emerged in recent years. There are now 60 RSV vaccine candidates in development that target pediatric and elderly populations. While most are at a preclinical stage, 16 candidates are in clinical development. This review summarizes current RSV vaccine research and development, including an overview of the vaccine platforms being used, the development stage of individual vaccine candidates, and gaps to be addressed to facilitate use of these vaccines to meet global health needs.     

A comparison of influenza and respiratory syncytial virus infections among infants admitted to hospital with acute respiratory infections.

Among 741 children under 5 years admitted to hospital with respiratory infections during two winters, infection with influenza A virus was diagnosed in 70 (9%), with influenza B virus in 8 (1%), and with respiratory syncytial virus (RSV) in 259 (35%). Both influenza virus and RSV infections were diagnosed most frequently in children under the age of one year, and diagnosed more frequently in males than females. Influenza illnesses were more severe in boys than girls. Both infections occurred more often, but were not more severe, in children from a conurbation than in those from 'rural' areas. Convulsions were the cause of 36% of admissions with influenza A infections, but were rare in RSV infections. Bronchiolitis was the reason for 39% of admissions with RSV infections, but was rare in influenza infections. It is suggested that infants admitted to hospital are a good source of influenza virus strains for monitoring antigenic variation.     

Respiratory syncytial virus fusion nanoparticle vaccine immune responses target multiple neutralizing epitopes that contribute to protection against wild-type and palivizumab-resistant mutant virus challenge

Human respiratory syncytial virus (RSV) is the leading cause of severe lower respiratory tract infections in newborns, young children, elderly, and immune-compromised. The RSV fusion (F) glycoprotein is a major focus of vaccine development and the target of palivizumab (Synagis®) which is licensed as an immuno-prophylactic for use in newborn children at high risk of infection. However, clinical use of a narrowly targeted monoclonal antibodies leads to the generation of escape mutant strains that are fully resistant to neutralization by the antibody. Herein, we evaluated the RSV F nanoparticle vaccine (RSV F vaccine), produced as near-full-length, pre-fusogenic F trimers that form stable protein-detergent nanoparticles. The RSV F vaccine induces polyclonal antibodies that bind to antigenic site II as well as other epitopes known to be broadly neutralizing. Cotton rats immunized with the RSV F vaccine produced antibodies that were both neutralizing and protected against wild-type RSV infection, as well as against a palivizumab-resistant mutant virus. Use of aluminum phosphate adjuvant with the RSV F vaccine increased site II antibody avidity 100 to 1000-fold, which correlated with enhanced protection against challenge. The breadth of the vaccine-induced antibody response was demonstrated using competitive binding with monoclonal antibodies targeting antigenic sites Ø, II, IV, and VIII found on pre-fusion and post-fusion conformations of RSV F. In summary, we found the RSV F vaccine induced antibodies that bind to conserved epitopes including those defined as pre-fusion F specific; that use of adjuvant increased antibody avidity that correlated with enhanced protection in the cotton rat challenge model; and the polyclonal, high-avidity antibodies neutralized and protected against both wild-type and palivizumab-resistant mutant virus. These data support the ongoing clinical development of the aluminum phosphate adjuvanted RSV F nanoparticle vaccine.     

Core bead chromatography for preparation of highly pure,infectious respiratory syncytial virus in the negative purification mode

Respiratory syncytial virus (RSV) is an important human pathogen, and is the most frequent viral cause of severe respiratory disease in infants. In addition, it is increasingly being recognized as an important cause of respiratory disease in the elderly and immunocompromised. Although a passive prophylactic treatment does exist for high-risk neonates and children, the overall disease burden warrants the development of a safe and effective prophylactic vaccine for use in otherwise healthy newborns and children. RSV is known to be an extremely labile virus, prone to aggregation and loss of infectious titer during virus handling and preparation procedures. To date infective RSV virions have been prepared by methods which are not readily scalable, such as density gradient ultracentrifugation. In this study we describe a scalable, chromatography-based purification procedure for preparation of highly pure, infectious RSV. The purification scheme is based on core bead technology and hollow fiber tangential flow filtration (TFF) and results in a ∼60% recovery of infectious virus titer. This method can be used to prepare highly purified wild type or live-attenuated vaccine strain viruses with titers as high as 1 × 10⁸ plaque forming units per mL. A live-attenuated RSV vaccine prepared by this method was found to be immunogenic and protective in vivo, and its purity was 50–200-fold greater with respect to host cell dsDNA and Vero host cell proteins, than the raw feed stream. The results presented here can be considered a starting point for downstream process development of a live-attenuated vaccine approach for prevention of disease by RSV.     

A dose-ranging study of a subunit Respiratory Syncytial Virus subtype A vaccine with and without aluminum phosphate adjuvantation in adults ≥65 years of age

We studied the safety and immunogenicity of a Respiratory Syncytial Virus (RSV)-A vaccine containing subunit antigens F, G and M in older persons, and its effect on influenza vaccine immunogenicity. In a dose-ranging, placebo-controlled, blinded trial 561 adults ≥65 years of age at five Canadian sites were randomized to one intramuscular injection of either 100, 50 or 25 μg RSV-A-alum vaccine or 100 μg non-adjuvanted RSV-A vaccine, or alum-placebo. All participants were offered inactivated influenza vaccine on day 32. Immunization was well tolerated and reactogenicity was similar between the RSV and influenza vaccines and the alum-placebo. Only the 100 μg non-adjuvanted RSV vaccine achieved the primary immunogenicity outcome of eliciting a ≥4-fold rise in neutralizing antibody (NA) titres against RSV-A in ≥50% of participants at day 32. Geometric mean titres against RSV-A and -B at all points were comparable in 100 μg adjuvanted and non-adjuvanted groups. At day 32, a ≥4-fold haemagluttinin inhibition (HI) antibody response or HI ≥40 to Influenza (A-H3N2) was seen in >74% of participants; no difference was seen between groups. A subunit non-alum-containing RSV-A vaccine was well tolerated in a large population ≥65 years and did not interfere with influenza vaccine immunogenicity. This RSV-A-based vaccine demonstrated NA rise which could provide seasonal protection against severe RSV illnesses from RSV-A or -B and warrants further testing to determine its efficacy in the prevention of clinical illness in elderly persons.     

Chitosan alters inactivated respiratory syncytial virus vaccine elicited immune responses without affecting lung histopathology in mice

Chitosan is a polysaccharide capable of augmenting immune responses with a proven safety record in animals and humans. These properties make it a potentially attractive agent for the prevention and treatment of infectious disease. Infection by respiratory syncytial virus (RSV) is the leading cause of serious lower respiratory disease in young children throughout the world. There is no licensed vaccine available against RSV whereas inactivated vaccine is known to cause enhanced respiratory disease instead of protection. Here, we investigated whether chitosan administered one or three days post-infection could protect animals against RSV infection and whether it could alter immune responses or immunopathology induced by inactivated RSV vaccine when administered twice before RSV infection. We found chitosan could modestly protect animals against RSV infection when given post-infection, while, in conjunction with inactivated RSV vaccine when given pre-infection, it could significantly reduce RSV infection in mice. Further mechanistic investigation revealed that chitosan enhanced antigen-specific immune responses through augmenting the induction of regulatory T cells, lung resident T cells and neutralizing antibodies while reversing Th2-skewed immune responses induced by inactivated RSV vaccine but, surprisingly, failing to reverse lung histopathology. Overall, this study sheds more light on the molecular mechanisms underlying inactivated RSV vaccine-induced disease.     

Immunization with an adjuvanted low-energy electron irradiation inactivated respiratory syncytial virus vaccine shows immunoprotective activity in mice

Respiratory syncytial virus (RSV) is a pathogen causing severe lower respiratory tract disease in infants and the elderly. In spite of the great need for a vaccine against RSV, currently there is no licensed product on the market. A very early vaccine candidate developed in the 1960s based on formaldehyde inactivation (FI) turned out to instead enhance the disease. Our novel inactivation method applied low-energy electron irradiation (LEEI) to produce a killed RSV vaccine. LEEI yielded inactivated virus particles with a reproducible virus antigen conservation above 70%, while FI resulted in highly variable antigen conservation. Immunization of mice with LEEI-RSV elicited a strong immune response, resulting in a drastic reduction in viral load upon challenge in two independent studies. These results have implications for the development of an RSV vaccine and should be validated in further preclinical and clinical studies.     

湖南省2012-2014年发热呼吸道症候群哨点监测结果及分析

目的 通过“发热呼吸道症候群监测”了解湖南省主要呼吸道病毒流行情况及病原谱分布,为发热呼吸道症候群防控及临床诊疗提供科学依据。 方法 2012年1月-2014年12月,从符合发热呼吸道症候群定义的患者身上采集鼻/咽拭子、肺泡灌洗液、痰液样本。采用基于毛细管凝胶电泳的“9+7呼吸道病毒多重PCR检测法”对标本中流感病毒(Flu)、呼吸道腺病毒(AdV)、呼吸道合胞病毒(RSV)、人鼻病毒(hRV)、副流感病毒(PIV)、偏肺病毒(HMPV)、人博卡病毒(hBoV)和人冠状病毒(hCoV)共8种病毒的16个亚型进行核酸检测。使用SPSS17.0统计学软件对个案信息及检测结果进行统计学分析。 结果 2012年1月-2014年12月,从哨点医院采集各类呼吸道标本共961份,其中男性病例标本595份,占61.91%;女性病例标本366份,占38.09%。在961份已检测的标本中,通过PCR检出阳性标本456份,阳性检出率为47.45%。男、女病例标本的阳性检出率差异无统计学意义(P>0.05)。按年龄将病例分为7个年龄组,各年龄组的阳性检出率分别为 1岁以下(52.80%)、1~2岁(66.53%)、2~5岁(49.86%)、6~15岁(25.56%)、16~49岁(11.11%)、50~64岁(17.39%)和65岁以上(19.40%)。在456份病毒阳性的标本中,检出混合感染48例,非混合感染408例。在检出的呼吸道病毒中,AdV、RSV和Flu所占的比例最大(病原谱构成比分别为25.90%、21.31%和17.73%),其次为hRV(14.14%)和PIV(11.75%);HMPV(4.78%)和hBoV(3.98%)检出较少,而hCoV只有两例阳性被检出。男性、女性的病原构成差异无统计学意义(P>0.05)。各年龄段的病原构成差异有统计学意义(P<0.05),其中2岁以下的儿童病例中检出的病毒以RSV(29.36%)和AdV(23.02%)为主,其次为PIV(13.10%)、hRV(11.90%)和Flu(11.11%);2~5岁的儿童病例中,AdV所占比例最大(34.02%),其次为hRV(18.56%)、Flu(18.04%)和RSV(15.46%);6~15岁少儿病例中以Flu(31.43%)和PIV(34.29%)为主,其次为AdV(17.14%)和hRV(14.29%);在16~49岁的病例中,只有Flu检出;在50岁以上老年病例中,病原构成以Flu(68.41%)为主,其余为PIV(10.53%)、HMPV(10.53%)和RSV(10.53%)。监测还发现,Flu、AdV有夏季和冬春季两个流行高峰;RSV有冬春季流行高峰;hRV有秋冬季流行高峰。 结论 所监测的8种呼吸道病毒是引发严重急性呼吸道住院病例的重要病原。5岁以下儿童及65岁以上的老人可能是主要的易感人群。其中,RSV、AdV、 Flu可能是引发儿童病毒性呼吸道感染的主要病原,而Flu则可能是引发成人病毒性呼吸道感染的主要原因。因此,应将这几种病毒作为呼吸道病毒防控的重点,进行更多的监测和更深入的研究。     

Diagnosis and epidemiological surveillance of influenza and respiratory syncytial virus infections: interest of multiplex PCR

OBJECTIVE: Respiratory infections require a rapid etiological diagnosis for efficient management of cases. We evaluated multiplex PCR used for the diagnosis and the epidemiological surveillance of influenza and respiratory syncytial virus (RSV) infections. PATIENTS AND METHODS: Our study included 278 patients (mean age: 37.2+/-22.9 years) with flu or flu-like syndromes, consulting physicians affiliated with the GROG Poitou-Charentes or hospitalized in the Poitiers teaching hospital. A multiplex PCR detecting A(H3), A(H1) and B influenza viruses, and RSV A and B, was performed with both a direct examination by immunofluorescence and cell-culture. RESULTS: We diagnosed a viral infection in 139 (50.0%) patients: 99 cases of influenza A(H3), 2 cases of influenza A(H1), 28 cases of influenza B and 11 cases of RSV infections. The diagnosis yield in GROG patients (52.3%) was significantly higher than that observed in hospitalized patients (34.5%) (P=0.04). All techniques were correlated in 61% of cases. The multiplex PCR yielded 22.3% more positive samples compared to the conventional techniques. All positive samples by conventional techniques were also positive by multiplex PCR. We observed a perfect correlation between viral types and subtypes determined by PCR and cell-culture. CONCLUSION: Multiplex PCR is a sensitive technique allowing an efficient and rapid diagnosis of respiratory infections due to influenza and RSV.