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.     

The role of non-viral antigens in the cotton rat model of respiratory syncytial virus vaccine-enhanced disease

In the 1960s, infant immunization with a formalin-inactivated respiratory syncytial virus (FI-RSV) vaccine candidate caused enhanced respiratory disease (ERD) following natural RSV infection. Because of this tragedy, intensive effort has been made to understand the root causes of how the FI-RSV vaccine induced a pathogenic response to subsequent RSV infection in vaccinees. A well-established cotton rat model of FI-RSV vaccine-enhanced disease has been used by numerous researchers to study the mechanisms of ERD. Here, we have dissected the model and found it to have significant limitations for understanding FI-RSV ERD. This view is shaped by our finding that a major driver of lung pathology is cell-culture contaminants, although FI-RSV immunization and RSV challenge serve as co-factors to exacerbate disease. Specifically, non-viral products from the vaccine and challenge preparations that are devoid of RSV give rise to alveolitis, which is considered a hallmark of FI-RSV ERD in the cotton rat model. Although FI-RSV immunization and RSV challenge promote more severe alveolitis, they also drive stronger cellular immune responses to non-viral antigens. The severity of alveolitis is associated with T cells specific for non-viral antigens more than with T cells specific for RSV. These results highlight the limitations of the cotton rat ERD model and the need for an improved animal model to evaluate the safety of RSV vaccine candidates.     

AIK-C measles vaccine expressing fusion protein of respiratory syncytial virus induces protective antibodies in cotton rats

Respiratory syncytial virus (RSV) is the most common cause of respiratory infection in infants, and no vaccine is available. In this report, recombinant AIK-C measles vaccines, expressing the RSV G or F protein of subgroup A (MVAIK/RSV/G or F), were investigated as a RSV vaccine candidate. MVAIK/RSV/G or F had the original ts phenotype and expressed RSV/G or F protein. Cross-reactive neutralizing antibodies against RSV subgroups A and B were detected in cotton rats immunized intramuscularly with MVAIK/RSV/F but not MVAIK/RSV/G. In cotton rats infected with RSV, RSV was recovered and lung histopathological finding was compatible with interstitial pneumonia, demonstrating thickening of alveolar walls and infiltration of mononuclear cells. When cotton rats immunized with MVAIK/RSV/F were challenged with homologous RSV subgroup A, no infectious RSV was recovered and very mild inflammation was noted without RSV antigen expression. When they were challenged with subgroup B, protective efficacy decreased. When cotton rats immunized with MVAIK/RSV/G were challenged with RSV subgroup A, low levels of infectious virus were recovered from lung. When challenged with subgroup B, no protective effects was demonstrated, demonstrating large amounts of RSV antigen in bronchial-epithelial cells. MVAIK/RSV/F is promising candidate and protective effects should be confirmed in monkey model.     

Immunization of cotton rats with the fusion (F) and large (G) glycoproteins of respiratory syncytial virus (RSV) protects against RSV challenge without potentiating RSV disease

A formalin-inactivated respiratory syncytial virus (RSV) vaccine tested 22 years ago failed to protect infant vaccinees against RSV infection or disease. Instead, lower respiratory tract disease was enhanced during subsequent infection by RSV. Enhancement of pulmonary pathology is also observed when cotton rats are immunized with formalin-inactivated RSV and subsequently infected with this virus. A major question that must be addressed for each new paramyxovirus vaccine is whether the immunogen possesses the capacity to potentiate disease. In the present study, we evaluated a newly developed purified F and G glycoprotein vaccine over a wide dosage range for immunogenicity, efficacy and capacity to potentiate pulmonary pathology in cotton rats. In addition, a formalin-inactivated RSV vaccine, which served as a positive control for enhancement of pulmonary pathology, was evaluated simultaneously. The results of these comparisons indicate that the purified F and G glycoprotein vaccine was highly immunogenic and was efficacious even in animals that developed low levels of serum-neutralizing antibodies. Furthermore, the F and G vaccine did not induce potentiation of pulmonary pathology. In contrast, formalin-inactivated RSV potentiated RSV pulmonary histopathology, but there was a sparing of potentiation at high and low doses. Both the formalin-inactivated RSV and purified F and G preparations induced a high level of serum antibodies capable of binding to purified F and G glycoproteins but both sets of antibodies had significantly reduced neutralizing activity. These results are encouraging because they suggest that purified paramyxovirus glycoproteins might be used safely as a vaccine. However, it would be desirable to use a glycoprotein preparation that stimulates antibodies that have a high level of neutralizing activity similar to those characteristic of adult human convalescent sera.     

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.     

Respiratory syncytial virus (RSV) fusion protein expressed by recombinant Sendai virus elicits B-cell and T-cell responses in cotton rats and confers protection against RSV subtypes A and B

The respiratory syncytial virus (RSV) is a serious pediatric pathogen for which there is currently no clinically approved vaccine. This report describes the design and testing of a new RSV vaccine construct (rSV-RSV-F), created by the recombination of an RSV F sequence with the murine parainfluenza virus-type 1 (Sendai virus, SV) genome. SV was selected as the vaccine backbone for this study, because it has previously been shown to elicit high-magnitude, durable immune activities in animal studies and has advanced to human safety trials as a xenogenic vaccine for human parainfluenza virus-type 1 (hPIV-1). Cells infected with the recombinant SV expressed RSV F protein, but F was not incorporated into progeny SV virions. When cotton rats were inoculated with the vaccine, high-titer RSV-binding and neutralizing antibodies as well as interferon-γ-producing T-cells were induced. Most striking was the protection against intra-nasal RSV challenge conferred by the vaccine. The rSV-RSV-F construct was also tested as a mixture with a second SV construct expressing the RSV G protein, but no clear advantage was demonstrated by combining the two vaccines. As a final analysis, the efficacy of the rSV-RSV-F vaccine was tested against an array of RSV isolates. Results showed that neutralizing and protective responses were effective against RSV isolates of both A and B subtypes. Together, experimental results encourage promotion of this recombinant SV construct as a vaccine candidate for the prevention of RSV in humans.     

Modelling the impact of respiratory syncytial virus (RSV) vaccine and immunoprophylaxis strategies in New Zealand

BackgroundMathematical models of respiratory syncytial virus (RSV) transmission can help describe seasonal epidemics and assess the impact of potential vaccines and immunoprophylaxis with monoclonal antibodies (mAb).MethodsWe developed a deterministic, compartmental model for RSV transmission, which was fitted to population-based RSV hospital surveillance data from Auckland, New Zealand. The model simulated the introduction of either a maternal vaccine or a seasonal mAb among infants aged less than 6 months and estimated the reduction in RSV hospitalizations for a range of effectiveness and coverage values.ResultsThe model accurately reproduced the annual seasonality of RSV epidemics in Auckland. We found that a maternal vaccine with effectiveness of 30–40% in the first 90 days and 15–20% for the next 90 days could reduce RSV hospitalizations by 18–24% in children younger than 3 months, by 11–14% in children aged 3–5 months, and by 2–3% in children aged 6–23 months. A seasonal infant mAb with 40–60% effectiveness for 150 days could reduce RSV hospitalizations by 30–43%, 34–48% and by 14–21% in children aged 0–2 months, 3–5 months and 6–23 months, respectively.ConclusionsOur results suggest that either a maternal RSV vaccine or mAb would effectively reduce RSV hospitalization disease burden in New Zealand. Overall, a seasonal mAb resulted in a larger disease prevention impact than a maternal vaccine.     

新生儿呼吸合胞病毒下呼吸道感染临床分析

目的探讨新生儿呼吸合胞病毒下呼吸道感染的临床特点。方法选取2008年5月-2011年4月,在医院进行治疗的呼吸合胞病毒下呼吸道感染260例新生儿为研究对象,对其采用氧疗、雾化吸入及抗病毒、抗菌药物治疗,后将其治疗有效率、住院时间及治疗前后的血清IL-4、IL-6、IL-12、CRP、IgE及外周血T淋巴细胞亚群水平进行统计比较。结果 260例患儿痊愈239例,好转20例,无效1例,总有效率为99.62%,住院时间为(12.8±2.8)d,治疗后IL-4、IL-6、IL-12分别为(7.55±2.69)、(16.69±7.14)、(56.48±13.39)ng/L,CRP为(6.88±2.39)mg/L,CD8+为(26.52±5.69)%均高于治疗前;治疗后IgE为(808.34±205.30)μg/L,CD3+、CD4+分别为(63.12±5.20)%、(45.58±5.20)%,CD4+/CD8+为(1.68±0.43)%,均高于治疗前,治疗前后差异有统计学意义(P<0.05)。结论经合理治疗,新生儿呼吸合胞病毒下呼吸道感染治疗效果较佳,可显著改善炎性因子及免疫状态。     

2008-2012年武汉市儿童流行性感冒病毒及呼吸道合胞病毒流行特征

目的 了解2008-2012年武汉市14岁及以下流行性感冒(简称流感)样症状患儿流感病毒和呼吸道合胞病毒(RSV)的感染情况及病毒流行特征.方法 于2008年7月至2012年6月在武汉市某医院门诊和急诊患者中,选择14岁以下的流感样病例作为研究对象,共2854例.研究对象各采集1份咽拭子标本,共采集2854份.从流感样病例咽拭子标本中提取病毒核酸,应用实时荧光定量逆转录PCR (RT-PCR)对流感病毒和RSV进行核酸检测和亚型鉴定,分析两种病毒感染的时间和年龄特征.结果 2854例患儿检测流感病毒阳性758例,阳性率为26.6％.其中,甲型流感病毒阳性547例(19.2％)、乙型流感病毒阳性211例(7.4％).流感病毒每年流行呈双峰特征,即夏季峰和冬春季峰.6～ 14岁组患儿流感病毒检测阳性率为48.0％(275/573),明显高于3～5岁组[26.6％ (213/801)]和3岁以下组[18.3％ (270/1480)],差异有统计学意义(x2=187.432,P＜0.01).患儿中RSV检测阳性219例,阳性率为7.7％,其中,RSV-A阳性108例,RSV-B阳性112例,混合感染1例.RSV流行呈明显的季节性,发病主要集中在秋、冬、春季,占全部发病病例的96.8％(212/219),每年流行情况波动较大.RSV-A、RSV-B交替流行,交替周期为2年,2008年9月至2009年5月及2009年12月至2010年3月以RSV-B感染为主,分别占76.6％(36/47)和96.9％(62/64)；2010年11月至2011年3月及2011年9月至2012年4月以RSV-A感染为主,分别占92.5％(37/40)和100.0％(48/48).但随着年龄的增长,RSV-A和RSV-B阳性率不断下降(RSV-A:x2=36.223,P ＜0.01；RSV-B:x2=36.281,P＜0.01),其中,RSV-A在＜1、1、2、3、4、5 ～9、10～ 14岁年龄组阳性率分别为7.0％ (26/373) 、5.9％ (39/662) 、4.0％ (18/445) 、3.2％ (13/406)、1.3％(3/236)、1.4％(7/517)、0.9％ (2/215)；RSV-B在各年龄组的阳性率分别为6.4％(24/373) 、6.0％(40/662)、4.5％(20/445)、4.4％(18/406)、1.3％(3/236)、1.0％(5/517) 、0.9％(2/215).0～3岁组RSV阳性197例,占RSV阳性患者总数的90.0％(197/219).2009年11月甲型H1N1流感暴发流行时,RSV阳性率为3.0％ (3/100),低于2008、2010、2011年同月份RSV检测阳性率[分别为18.2％ (6/33)、10.8％ (10/93) 、10.0％ (4/40)],差异有统计学意义(x2=8.450,P＜0.05)；2011年1月甲型H1N1流感暴发流行时,RSV阳性率为5.7％(3/53),低于2009、2010、2012年同月份RSV检测阳性率[分别为21.7％(5/23) 、28.6％(22/77)、16.0％(8/50)],差异有统计学意义(x2=11.233,P＜0.05)；2011年9月,流感夏季峰缺失时,RSV检测阳性率为25.0％(10/40),高于2008、2009、2010年同月份RSV检测阳性率[分别为11.4％ (4/35) 、1.7％ (2/118)、0.0％ (0/109)],差异有统计学意义(x2=32.521,P ＜0.01).结论 流感病毒和RSV是武汉地区流感样患儿重要的病原,两种病毒流行的季节性、年龄分布等特征各不相同,流感病毒对RSV的流行存在着一定的抑制作用.     

A chimeric glycoprotein of human respiratory syncytial virus termed FG induces T-cell mediated immunity in mice

Popliteal lymph node cells taken from mice vaccinated with the FG glycoprotein were exposed in vitro to respiratory syncytial virus (RSV) antigens. Proliferation to FG or RSV antigens was blocked with anti-CD4 monoclonal antibody treatment. FG-vaccinated mice developed classical late delayed type hypersensitivity (DTH) reactions when exposed to FG antigen in vivo.     

Antibody response to the central unglycosylated region of the respiratory syncytial virus attachment protein in mice

We examined the humoral immune response to the unglycosylated central region of the respiratory syncytial virus (RSV) attachment (G) protein in mice following intranasal challenge at day 0 (primary) and day 21 (secondary) with subtype A (A2 strain) or B (B1 strain) RSV preparations. Our serological screening reagents included bacterially derived glutathione S-transferase (GST) fusion proteins, each bearing a portion of the RSV G central core (CC; residues 151-190), proximal central core (PCC; residues 151-172), and the distal central core (DCC; residues 173-190) and purified RSV G proteins from subtype A and B viruses. Convalescent sera collected on day 21 following primary RSV infection bore robust IgG response primarily against the homosubtypic RSV G DCC with relatively modest antigen affinity/avidity as demonstrated by brief incubation with 6M urea. In contrast, sera collected on day 42 following secondary homosubtypic RSV infection bore IgG titers of higher magnitudes and antigen affinity/avidity against the homosubtypic RSV G CC, PCC, and/or the DCC regions and full-length RSV G protein but not against the heterosubtypic RSV G protein or recombinant CC subdomains. In contrast, heterosubtypic secondary RSV infection elicits a broad array of IgG responses with titers of varying magnitudes to homo- and heterosubtypic RSV G CC regions as well as to purified F, Ga, and Gb proteins with the notable exception of minimal response to the RSV G DCC domain associated with the secondary RSV challenge. Our results have implications for RSV G-based serological assays as well as prophylactic immunotherapy and RSV vaccine development.     

杭州地区小儿呼吸道合胞病毒感染流行特点与气象学因素

目的阐明杭州地区小儿呼吸道合胞病毒(RSV)感染流行特点及影响RSV感染流行的气象学因素。方法连续3年对住院肺炎患儿中RSV的检出率进行动态观察,将月平均气温、相对湿度及雨天分别与月RSV检出率进行相关性分析。结果3年共检测患儿13642例,RSV阳性率为25.8%,其中≤1岁组检出率33.1%,1～3岁组19.7%,>3岁组5.1%,各年龄组间检出率差异有统计学意义(χ2=763.7,P=0.000)。RSV感染率总体上11月份开始明显增高,流行持续到次年的3-4月份,但每年的流行仍存在差别。雨天与RSV检出率之间r=0.32(P=0.066);相对湿度与RSV检出率之间r=-0.27(P=0.117);平均气温与RSV检出率之间r=-0.83(P=0.000),RSV检出率与气温之间的回归方程式:检出率(%)=52.933-1.914×气温(℃)。结论RSV是目前引起小儿肺炎的主要病毒。RSV在≤1岁组感染率最高,年龄越大感染率越低。RSV在杭州地区的流行见于冬春季,但存在变化,低气温是导致RSV感染流行的主要因素。     

呼吸道合胞病毒感染导致大鼠肾病病理模型持续感染证据研究

目的探讨呼吸道合胞病毒(RSV)感染SD大鼠的肾病病理模型中,RSV持续感染证据。 方法选取2个月龄、体重为180～200 g的清洁级雄性SD大鼠62只为研究对象。采用随机数字表法将其随机分为RSV组(n＝35)与对照组(n＝27)。①对RSV组SD大鼠,采用毒力疱斑形成单位(PFU)为6×10⁶ RSV液(0.4 mL/d腹腔注射＋0.2 mL/d鼻腔滴注)×3 d处理,建立RSV感染SD大鼠肾病病理模型。对照组采用磷酸盐缓冲液(PBS)(0.4 mL/d腹腔注射＋ 0.2 mL/d鼻腔滴注)×3 d处理,建立对照组PBS模型。两组均以造模前1 d计算为第0天,造模结束时计算为造模成功第1天。通过电子显微镜观察RSV感染SD大鼠肾病病理模型造模结束后第7与60天的肾小球超微结构改变,判断SD大鼠肾病病理模型造模是否成功。②RSV组SD大鼠肾病病理模型造模成功后,将35只SD大鼠平均分为7个亚组(RSV 0、7、15、30、60、90及120 d亚组),每个亚组均为5只SD大鼠;将对照组27只PBS模型SD大鼠,按照上述时间点分为7个亚组(PBS 0、7、15、30、60、90及120 d亚组),每个亚组分别为5、5、5、3、3、3、3只SD大鼠。分别测定各个RSV亚组与PBS亚组SD大鼠的24 h尿蛋白定量、血清白蛋白及血清肌酐水平等指标,并进行统计学分析。③于上述指标测定完毕后,分别于上述时间点处死RSV亚组与PBS亚组SD大鼠。同时,切除SD大鼠左肾下2/3、左肺下2/3和脾下2/3,以4%多聚甲醛固定24 h后,石蜡包埋后切片,采用苏木精-伊红(HE)染色,光镜下观察各个亚组SD大鼠肾、肺、脾组织是否发生病理学形态结构改变。④以RSV G蛋白单克隆抗体为一抗,制作RSV组SD大鼠肾、肺、脾组织免疫组化检测标本,于光镜下寻找各个RSV亚组RSV G蛋白存在证据。⑤以RSV G蛋白单克隆抗体为一抗,采用与绿色荧光试剂结合,同时采用4',6-二脒基-2-苯基吲哚(DAPI)染料蓝染细胞核,制作各个RSV亚组SD大鼠肾组织荧光免疫检测标本,于荧光显微镜下寻找RSV G蛋白存在证据及其与细胞核关系。⑥切除剩余RSV亚组左肾上1/3,以3%戊二醛固定后,送至四川大学华西医院病理科制作电子显微镜标本,于电子显微镜下观察各个RSV亚组SD大鼠肾小球超微结构是否发生病理改变,同时寻找RSV颗粒。 结果①RSV组SD大鼠肾病病理模型造模成功,对照组PBS模型亦造模成功。②各个RSV亚组与PBS亚组SD大鼠的24 h尿蛋白定量水平检测结果显示,RSV 0与90 d亚组分别与相同时间点PBS亚组比较,差异均无统计学意义(P>0.05);而RSV 7、15、30、60及120 d亚组,却分别较相同时间点PBS亚组均显著升高,并且差异均有统计学意义(t＝6.9、3.1、3.9、2.2、5.6,P<0.05)。血清白蛋白及血清肌酐水平检测结果显示,7个RSV亚组分别与相同时间点的7个PBS亚组比较,差异均无统计学意义(P>0.05)。③于光镜下观察各个RSV亚组SD大鼠的肾、肺、脾组织形态结构(HE染色)结果显示,除RSV 0 d亚组外,其余各个亚组肾、肺、脾组织形态结构均出现不同程度病理改变。④于光镜下观察各个RSV亚组SD大鼠的肾、肺、脾组织免疫组化技术检测结果显示,除RSV 0 d亚组外,其余各个RSV亚组肾、肺、脾组织中,均有特异性RSV G蛋白呈持续阳性表达。各个RSV亚组肾小球RSV G蛋白定量分析结果显示,RSV 0、7、15及30 d亚组RSV G蛋白表达水平呈逐渐增强趋势,并维持在高水平,但在RSV 60、90及120 d亚组又呈逐渐减弱趋势。⑤于荧光显微镜下观察各个RSV亚组SD大鼠肾组织RSV G蛋白荧光免疫检测结果显示,RSV G蛋白与DAPI蓝染的细胞核重叠。⑥透射电子显微镜下观察发现,RSV组肾组织中RSV颗粒持续存在,RSV 120 d亚组肾小球超微结构仍存在病理改变。 结论RSV G蛋白在RSV感染SD大鼠肾病病理模型造模成功后,大鼠肾、肺、脾组织中RSV G蛋白呈持续阳性表达,证明RSV呈持续感染状态。这一结果或提示,包括RSV在内的呼吸道病毒持续感染,是导致微小病变型肾病综合征(MCNS)患儿免疫功能紊乱的主要原因。     

Association of chronic alcohol consumption and increased susceptibility to and pathogenic effects of pulmonary infection with respiratory syncytial virus in mice.

Chronic alcohol abuse by human beings has been shown to be associated with increased susceptibility to pulmonary infections and severity of inflammatory responses associated with pulmonary infection. On the basis of the higher likelihood of exposure to respiratory viruses, people who abuse alcohol would logically be susceptible to respiratory viral infections. To test this hypothesis, mice were provided alcohol in drinking water for 13-16 weeks with the Meadows-Cook protocol and infected intranasally with respiratory syncytial virus. At various times after infection, severity of infection was determined by evaluation of cellular and cytokine composition of bronchoalveolar lavage fluid (BALF) and histologic evaluation of inflammation. Infection was associated with neutrophil infiltration in both groups, but the proportion and number of neutrophils in BALF were significantly greater in the alcohol consumption group than in the control group. Concentrations of tumor necrosis factor-alpha and monocyte chemoattractant protein-1 in BALF in the alcohol consumption group were increased. Interferon (IFN)-gamma concentrations were lower in the alcohol consumption group at later times of infection. Pulmonary inflammation was cleared by 3-5 days after infection in the control group. In contrast, pulmonary inflammation was evident in the alcohol consumption group after 7 days of infection, and some mice showed severe inflammation with hemorrhage and edema. IFN-alpha/beta was evident in BALF at low concentrations in the alcohol consumption group for several days after infection, and increased mRNA for IFN-alpha/beta was also evident in the alcohol consumption group. This was accompanied by the presence of virus in this group at these times of infection. Chronic alcohol consumption increased severity of pulmonary infection with a virus that naturally infects hosts by an aerosol route. Infection of mice that had consumed alcohol chronically was more severe in terms of increased proinflammatory cytokine production, inflammation, and a failure to clear the virus from the lungs.     

The genetic variability of glycoproteins among respiratory syncytial virus subtype A in China between 2009 and 2013

Human respiratory syncytial virus (RSV) is the leading cause of acute lower respiratory tract infections in infants and children under 5 years of age. The novel genotype ON1 has a 72-nucleotide duplication, which is the largest duplicated genome portion of RSV. Whether the ON1 genotype will follow the pattern of the BA genotype, which has a 60-nucleotide duplication, and become the predominant RSV-A strain is a global concern. To obtain information regarding the prevalence of the ON1 genotype in Chongqing in Southwestern China, we examined the circulation pattern of RSV-A identified over four consecutive years (June 2009 to August 2013). In this study, 312 (12%) RSV-A strains were isolated from 2601 nasopharyngeal aspirates, and partial G gene was sequenced successfully in 250 isolates. Of the sequenced Chongqing RSV-A isolates, 237 (94.8%) strains were the NA1 genotype, 4 (1.6%) strains were the NA3 genotype, 4 (1.6%) strains were the NA4 genotype, 1 (0.4%) strain was the GA1 genotype, and 4 (1.6%) strains were identified as the ON1 genotype. Analysis of the distribution, phylogeny, and evolution of the ON1 strains that were collected globally until December 2013 revealed that the ON1 genotype has rapidly disseminated across the world under positive selection pressures. Future studies will determine whether this new genotype will continue to spread and become the dominant strain of RSV-A worldwide. These findings may contribute to the understanding of RSV evolution and to the potential development of a vaccine against RSV.     

The association between age and the development of respiratory syncytial virus neutralising antibody responses following natural infection in infants

To determine the age at which infants mount significant neutralising antibody responses to both natural RSV infection and live vaccines that mimic natural infection, RSV-specific neutralising antibodies in the acute and convalescent phase sera of infants with RSV infection were assayed. Age-specific incidence estimates for hospitalisation with severe RSV disease were determined and compared to age-specific neutralising antibody response patterns. Disease incidence peaked at between 2 and 3.9 months of life. Following natural infection, relative to the mean acute phase antibody titre, the mean convalescent titre was lower in the 0–1.9 month age class, no different in the 2–3.9 month age class and greater in all age classes greater than 4 months. These data suggest effective vaccination with live vaccines that mimic natural infection may not be achieved before the age of 4 months. Maternal vaccination may be an alternative to direct infant vaccination in order to protect very young babies.     

An insect cell derived respiratory syncytial virus (RSV) F nanoparticle vaccine induces antigenic site II antibodies and protects against RSV challenge in cotton rats by active and passive immunization

Post-infectious immunity to respiratory syncytial virus (RSV) infection results in limited protection as evidenced by the high rate of infant hospitalization in the face of high titer, maternally derived RSV-specific antibodies. By contrast, RSV fusion (F) glycoprotein antigenic site II humanized monoclonal antibodies, palivizumab and motavizumab, have been shown to reduce RSV-related hospitalization in infants. Immunogenicity and efficacy studies were conducted in cotton rats comparing a recombinant RSV F nanoparticle vaccine with palivizumab and controlled with live RSV virus intranasal immunization and, formalin inactivated RSV vaccine. Active immunization with RSV F nanoparticle vaccine containing an alum adjuvant induced serum levels of palivizumab competing antibody (PCA) greater than passive administration of 15 mg/kg palivizumab (human prophylactic dose) in cotton rats and neutralized RSV-A and RSV-B viruses. Immunization prevented detectable RSV replication in the lungs and, unlike passive administration of palivizumab, in the nasal passage of challenged cotton rats. Histology of lung tissues following RSV challenge showed no enhanced disease in the vaccinated groups in contrast to formalin inactivated ‘Lot 100’ vaccine. Passive intramuscular administration of RSV F vaccine-induced immune sera one day prior to challenge of cotton rats reduced viral titers by 2 or more log₁₀ virus per gram of lung and nasal tissue and at doses less than palivizumab. A recombinant RSV F nanoparticle vaccine protected lower and upper respiratory tract against both RSV A and B strain infection and induced polyclonal palivizumab competing antibodies similar to but potentially more broadly protective against RSV than palivizumab.     

Enhanced pulmonary pathology associated with the use of formalin-inactivated respiratory syncytial virus vaccine in cotton rats is not a unique viral phenomenon

The specificity of viral antigens in the formalin-inactivated, alum-precipitated respiratory syncytial virus (FI-RSV) vaccine in augmenting the pulmonary inflammatory response was evaluated. Cotton rats were immunized with a FI-RSV vaccine derived from Vero cells, a monkey cell line, or HEp-2 cells, a human cell line. The FI-RSV/Vero and the FI-RSV/HEp-2 vaccines were prepared similarly to the original Lot-100 FI-RSV vaccine that was associated with enhanced disease in the mid-1960s field trials. Each vaccine was administered intramuscularly at various doses and intervals. At I, 4 or 7 weeks after the last vaccine dose, cotton rats were challenged with 10⁶ plaque-forming units of live RSV grown in HEp-2 cells. For controls, FI-parainfluenza, FI-HEp-2 and alum vaccines, and live RSV primary infection were used. For measuring virus replication and histopathology, lungs were harvested at 4 and 8 days postchallenge. A dose-response relationship to vaccine dose was observed for ELISA, neutralizing and antifusion antibodies. All animals given three doses or two of the higher doses of FI-RSV/Vero vaccine developed significant neutralizing antibody, were protected against pulmonary virus replication and had similar low levels of histopathology compared with live RSV and controls. Two immunizations of the lowest dose of FI-RSV/Vero vaccine did not induce neutralizing antibody, did not provide protection of the lung against RSV and did not enhance the pulmonary cellular response. However, FI-RSV/HEp-2 vaccine was associated with significant enhanced pulmonary histopathology despite inducing high titres of neutralizing antibody and protecting the lungs against RSV infection. FI-HEp-2 vaccine was also associated with significantly enhanced pulmonary histopathology. Under the conditions tested, non-viral antigens played a major role in the augmentation of the pulmonary cellular response in cotton rats that were given FI-RSV/HEp-2 vaccine.