Human infections with novel reassortant influenza A(H3N2)v viruses, United States, 2011

During July-December 2011, a variant virus, influenza A(H3N2)v, caused 12 human cases of influenza. The virus contained genes originating from swine, avian, and human viruses, including the M gene from influenza A(H1N1)pdm09 virus. Influenza A(H3N2)v viruses were antigenically distinct from seasonal influenza viruses and similar to proposed vaccine virus A/Minnesota/11/2010.     

Host immune response to A(H1N1)pdm09 vaccination and infection: a one-year prospective study on six cohorts of subjects

Background

The long-term immunogenicity after novel vaccine against A(H1N1)pdm09 administration or natural infection has not been well investigated.

Methods

Six cohorts of subjects were followed up for over one year: one-dose A(H1N1)pdm09 vaccine recipient, A(H1N1)pdm09-seasonal trivalent vaccine recipients in different orders, confirmed A(H1N1)pdm09 patients without vaccination, with previous A(H1N1)pdm09 or seasonal influenza vaccination. Peripheral blood mononuclear cells and sera samples were collected at baseline and month 1, 2, 3, 7 and 14 after vaccination (infection). The immunogenicity was determined by hemagglutination-inhibition (HI) and B cell enzyme-linked immunospot (ELISPOT) assays.

Results

Single dose of A(H1N1)pdm09 vaccine elicited antibody titer of greater than 1:40 in 40% adults for 1 year and mean live of this adequate antibody was determined as 8.35 months. In contrast, responses after natural infections had lower peaking level and a relatively longer antibody duration, with estimated mean lives of 11.8 months. Pre-vaccination with the seasonal flu vaccine led to a significant reduction in HI titer to A(H1N1)pdm09 one month after vaccination, while pre-vaccination with A(H1N1)pdm09 had no effect on seasonal influenza vaccination. Seasonal flu vaccination followed by A(H1N1)pdm09 infection elicited boosting effect on antibody response against A(H1N1)pdm09. A similar memory B cell response was elicited from both vaccination and infection by ELISPOT assay.

Conclusions

The long-term decay of immunity for A(H1N1)pdm09 vaccine and natural infection indicates the need of revaccination after the host lose protection acquired from either vaccination or infection. Prior infection, rather than the pre-vaccination with seasonal influenza could act on the host immunity to elicit boosting effect on the A(H1N1)pdm09.     

Novel influenza vaccine M2SR protects against drifted H1N1 and H3N2 influenza virus challenge in ferrets with pre-existing immunity

Current influenza vaccines do not provide effective protection against heterologous influenza viruses. The ability of the novel M2SR influenza vaccine to protect against drifted influenza viruses was evaluated in naïve ferrets and in ferrets with pre-existing immunity to influenza. In naïve ferrets, M2SR provided similar protection against drifted challenge viruses as the comparator vaccine, FluMist®. However, in ferrets with pre-existing immunity, M2SR provided superior protection than FluMist in two model systems.In the first model, ferrets were infected with influenza A H1N1pdm and influenza B viruses to mimic the diverse influenza exposure in humans. The pre-infected ferrets, seropositive to H1N1pdm and influenza B but seronegative to H3N2, were then vaccinated with H3N2 M2SR or monovalent H3N2 FluMist virus (A/Brisbane/10/2007, clade 1) and challenged 6 weeks later with a drifted H3N2 virus (clade 3C.2a). Antibody titers to Brisbane/10/2007 were higher in M2SR vaccinated ferrets than in FluMist vaccinated ferrets in the pre-infected ferrets whereas the opposite was observed in naïve ferrets. After challenge with drifted H3N2 virus, M2SR provided superior protection than FluMist monovalent vaccine.In the second model, the impact of homologous pre-existing immunity upon vaccine-induced protection was evaluated. Ferrets, pre-infected with H1N1pdm virus, were vaccinated 90 days later with H1N1pdm M2SR or FluMist monovalent vaccine and challenged 6 weeks later with a pre-pandemic seasonal H1N1 virus, A/Brisbane/59/2007 (Bris59). While cross-reactive serum IgG antibodies against the Bris59 HA were detected after vaccination, anti-Bris59 hemagglutination inhibition antibodies were only detected post-challenge. M2SR provided better protection against Bris59 challenge than FluMist suggesting that homologous pre-existing immunity affected FluMist virus to a greater degree than M2SR.These results suggest that the single replication intranasal M2SR vaccine provides effective protection against drifted influenza A viruses not only in naïve ferrets but also in those with pre-existing immunity in contrast to FluMist viruses.     

Impact of pre-existing immunity on the induction of functional cross-reactive anti-hemagglutinin stalk antibodies following vaccination with an AS03 adjuvanted pandemic H1N1 vaccine

The 2009 pandemic H1N1 (A(H1N1)pdm09) virus had a highly divergent hemagglutinin (HA) compared to pre-2009 seasonal H1N1 strains. Most peoples were immunologically naïve to the A(H1N1)pdm09, although hospital workers were exposed early in the pandemic before pandemic vaccines became available. Here, we evaluated how pre-existing antibodies influence the induction of cross-functional HA stalk antibodies following A(H1N1)pdm09 vaccination.Fifty-six healthcare workers vaccinated with AS03 adjuvanted A(H1N1)pdm09 vaccine were chosen by their pre-vaccination priming status (primed HI titers ≥ 40 or unprimed < 40). We analyzed the HA head- and stalk-specific serum IgG subclasses at pre- and 21 days post-vaccination. We also assessed the functionality of the HA stalk-specific antibodies to neutralize virus and mediate antibody dependent cellular cytotoxicity (ADCC).Primed individuals had higher pre-existing HA head- and stalk-specific IgG1, as well as higher ADCC functionality of stalk antibodies. However, following vaccination with the adjuvanted pandemic vaccine, only the quantity of HA head specific IgG1 antibodies were significantly higher than in unprimed individuals. The priming status did not impact upon the cross-reactive HA stalk specific IgG antibodies or their ability to neutralize virus or induce ADCC post-vaccination. In conclusion, a single dose of AS03 adjuvanted pandemic vaccine elicited similar levels of functional antibodies in naïve and primed individuals. These findings are important for understanding the immunological factors that impact or modulate pandemic vaccine responses in humans.     

Coupling sensitive in vitro and in silico techniques to assess cross-reactive CD4(+) T cells against the swine-origin H1N1 influenza virus

The outbreak of the novel swine-origin H1N1 influenza in the spring of 2009 took epidemiologists, immunologists, and vaccinologists by surprise and galvanized a massive worldwide effort to produce millions of vaccine doses to protect against this single virus strain. Of particular concern was the apparent lack of pre-existing antibody capable of eliciting cross-protective immunity against this novel virus, which fueled fears this strain would trigger a particularly far-reaching and lethal pandemic. Given that disease caused by the swine-origin virus was far less severe than expected, we hypothesized cellular immunity to cross-conserved T cell epitopes might have played a significant role in protecting against the pandemic H1N1 in the absence of cross-reactive humoral immunity. In a published study, we used an immunoinformatics approach to predict a number of CD4⁺ T cell epitopes are conserved between the 2008-2009 seasonal H1N1 vaccine strain and pandemic H1N1 (A/California/04/2009) hemagglutinin proteins. Here, we provide results from biological studies using PBMCs from human donors not exposed to the pandemic virus to demonstrate that pre-existing CD4⁺ T cells can elicit cross-reactive effector responses against the pandemic H1N1 virus. As well, we show our computational tools were 80-90% accurate in predicting CD4⁺ T cell epitopes and their HLA-DRB1-dependent response profiles in donors that were chosen at random for HLA haplotype. Combined, these results confirm the power of coupling immunoinformatics to define broadly reactive CD4⁺ T cell epitopes with highly sensitive in vitro biological assays to verify these in silico predictions as a means to understand human cellular immunity, including cross-protective responses, and to define CD4⁺ T cell epitopes for potential vaccination efforts against future influenza viruses and other pathogens.     

Impact of prior infection and repeated vaccination on post-vaccination antibody titers of the influenza A(H1N1)pdm09 strain in Taiwan schoolchildren: Implications for public health

BackgroundThe objective of this study was to evaluate the effects of prior-infection and repeated vaccination on post-vaccination antibody titers.MethodsA(H1N1)pdm09 strain was included in 2009 pandemic monovalent, 2010–2011, and 2011–2012 trivalent influenza vaccines (MIVpdm09, TIV10/11, TIV11/12) in Taiwan. During the 2011–2012 influenza season, we conducted a prospective sero-epidemiological cohort study among schoolchildren from grades 1 – 6 in the two elementary schools in Taipei with documented A(H1N1)pdm09 vaccination records since 2009. Serum samples were collected at pre-vaccination, 1-month, and 4-months post-vaccination (T1, T2, T3). Anti-A(H1N1)pdm09 hemagglutination inhibition titers (HI-Ab-titers) were examined. We also investigated the impact of four vaccination histories [(1) no previous vaccination (None), (2) vaccinated in 2009–2010 season (09v), (3) vaccinated in 2010–2011 season (10v), and (4) vaccinated consecutively in 2009–2010 and 2010–2011 seasons (09v + 10v)] and pre-vaccination HI-Ab levels on post-vaccination HI-Ab responses as well as adjusted vaccine effectiveness (aVE) against serologically-defined infection from T2 to T3.ResultsTIV11/12 had zero serious adverse events reported. A(H1N1)pdm09 strain in TIV11/12 elicited seroprotective Ab-titers in 98% of children and showed promising protection (aVE: 70.3% [95% confidence interval (CI): 51.0–82.1%]). Previously unvaccinated but infected children had a 3.96 times higher T2 geometric mean titer (T2-GMT) of HI-Ab than those naïve to A(H1N1)pdm09 (GMT [95% CI]: 1039.7[585.3–1845.9] vs. 262.5[65.9–1045], p = 0.046). Previously vaccinated children with seroprotective T1-Ab-titers had a higher T2-GMT and a greater aVE than those with non-seroprotective T1-Ab-titers. Repeatedly vaccinated children had lower T2-GMT than those receiving primary doses of TIV11/12. However, after controlling prior infection and T1-Ab-titers, differences in T2-GMT among the four vaccination histories became insignificant (p = 0.16).ConclusionThis study supports the implementation of annual mass-vaccination with A(H1N1)pdm09 in schoolchildren for three consecutive influenza seasons when vaccine and circulating strains were well matched, and found that prior infection and pre-vaccination HI-Ab levels positively impacted post-vaccination HI-Ab responses.     

Identification of HLA class II H5N1 hemagglutinin epitopes following subvirion influenza A (H5N1) vaccination

Prophylactic immunization against influenza infection requires CD4+ T-helper cell activity for optimal humoral and cellular immunity. Currently there is one FDA approved H5N1 subvirion vaccine available, although stockpiles of this vaccine are insufficient for broad population coverage and the vaccine has only demonstrated modest immunogenicity. Specific activation of CD4+ T-helper cells using class II H5N1 HA peptide vaccines may be a useful component in immunization strategy and design. Identification of HLA class II HA epitopes was undertaken in this report by obtaining PBMCs from volunteers previously immunized with an H5N1 inactivated subvirion vaccine, followed by direct ex vivo stimulation of CD4+ T cells against different sources of potential HA class II epitopes. In the 1st round of analysis, 35 donors were tested via IFN-γ ELISPOT using pools of overlapping HA peptides derived from the H5N1 A/Thailand/4(SP-528)/2004 virus, recombinant H5N1 (rHA) and inactivated H5N1 subvirion vaccine. In addition, a series of algorithm-predicted epitopes coupled with the Ii-Key moiety of the MHC class II-associated invariant chain for enhanced MHC class II charging were also included. Specific responses were observed for all 20 peptide pools, with 6–26% of vaccinated individuals responding to any given pool (donor response frequency) and a magnitude of response ranging from 3- to >10-fold above background levels. Responses were similarly observed with the majority of algorithm-predicted epitopes, with a donor response frequency of up to 29% and a magnitude of response ranging from 3–10-fold (11/24 peptides) to >10-fold above background (7/24 peptides). PBMCs from vaccine recipients that had detectable responses to H5N1 rHA following 1st round analysis were used in a 2nd round of testing to confirm the identity of specific peptides based on the results of the 1st screening. Sixteen individual HA peptides identified from the library elicited CD4+ T cell responses between 3- and >10-fold above background, with two peptides being recognized in 21% of recipients tested. Eight of the putative MHC class II epitopes recognized were found in regions showing partial to significant sequence homology with New Caledonia H1N1 influenza HA, while eight were unique to H5N1 HA. This is the first study to identify H5N1 HA epitope-specific T cells in vaccine recipients and offers hope for the design of a synthetic peptide vaccine to prime CD4+ T-helper cells. Such a vaccine could be used to provide at least some minimal level of H5N1 protection on its own and/or prime for a subsequent dose of a more traditional but supply-limited vaccine.     

2012-2018年青岛市A型(H1N1)pdm09流感病毒奥司他韦耐药株基因特征

目的 了解2012-2018年青岛市人群A型(H1N1)pdm09流感病毒奥司他韦耐药株基因特征。方法 收集2012年4月-2018年3月间青岛市A(H1N1)pdm09毒株397份,逆转录聚合酶链反应(RT-PCR)扩增神经氨酸酶(Neuraminidase,NA)和血凝素(Hemagglutinin,HA)基因全长,序列测定后进行耐药位点和氨基酸变异及进化分析。结果 5株发生了H275Y突变,为奥司他韦耐药株;另有4株S247N突变,可能为奥司他韦耐药株。2012-2018年H275Y突变株检出率依次为2.8 %、2.0 %、0.0 %、1.1 %、0.0 %和0.7 %。NA和HA进化树显示,2012-2013年青岛H275Y突变株与美国耐药株A/Tennessee/03/2013更接近,2013-2014年青岛H275Y突变株与国内株和日本札幌耐药株更接近,这两个年度耐药株的毒株起源可能有所不同。突变株在酶活性位点、抗原决定簇、受体结合位点及其他功能位点(如HA位点D222、Q223和NA位点V241I、N369K和N386K)的转变与野生敏感株一致。结论 青岛市A型(H1N1)pdm09流感病毒奥司他韦耐药株明显增加且流行起源不同,但并未取得比野生株更强的流行能力。奥司他韦仍可作为流感预防和治疗的有效手段。     

A polyvalent influenza A DNA vaccine induces heterologous immunity and protects pigs against pandemic A(H1N1)pdm09 virus infection

The composition of current influenza protein vaccines has to be reconsidered every season to match the circulating influenza viruses, continuously changing antigenicity. Thus, influenza vaccines inducing a broad cross-reactive immune response would be a great advantage for protection against both seasonal and emerging influenza viruses. We have developed an alternative influenza vaccine based on DNA expressing selected influenza proteins of pandemic and seasonal origin. In the current study, we investigated the protection of a polyvalent influenza DNA vaccine approach in pigs. We immunised pigs intradermally with a combination of influenza DNA vaccine components based on the pandemic 1918 H1N1 (M and NP genes), pandemic 2009 H1N1pdm09 (HA and NA genes) and seasonal 2005 H3N2 genes (HA and NA genes) and investigated the protection against infection with virus both homologous and heterologous to the DNA vaccine components.     

Antigenic and genetic characterization of influenza viruses circulating in Bulgaria during the 2015/2016 season

Influenza virological surveillance is an essential tool for early detection of novel genetic variants of epidemiologic and clinical significance. The aim of this study was to determine the antigenic and molecular characteristics of influenza viruses circulating in Bulgaria during the 2015/2016 season. The season was characterized by dominant circulation of A(H1N1)pdm09 viruses, accounting for 66% of detected influenza viruses, followed by B/Victoria-lineage viruses (24%) and A(H3N2) viruses (10%). All sequenced influenza A(H1N1)pdm09, A(H3N2) and B/Victoria-lineage viruses belonged to the 6B.1, 3C.2a and 1A genetic groups, respectively. Amino acid analysis of 57 A(H1N1)pdm09 isolates revealed the presence of 16 changes in hemagglutinin (HA) compared to the vaccine virus, five of which occurred in four antigenic sites, together with 16 changes in neuraminidase (NA) and a number of substitutions in proteins MP, NP, NS and PB2. Despite the many amino acid substitutions, A(H1N1)pdm09 viruses remained antigenically closely related to A/California/7/2009 vaccine virus. Bulgarian A(H3N2) strains (subclade 3C.2a) showed changes at 11 HA positions four of which were located in antigenic sites A and B, together with 6 positions in NA, compared to the subclade 3C.3a vaccine virus. They contained unique HA1 substitutions N171K, S312R and HA2 substitutions I77V and G155E compared to Bulgarian 3C.2a viruses of the previous season. All 20 B/Victoria-lineage viruses sequenced harboured two substitutions in the antigenic 120-loop region of HA, and 5 changes in NA, compared to the B/Brisbane/60/2008 vaccine virus. The results of this study reaffirm the continuous genetic variability of circulating seasonal influenza viruses and the need for continued systematic antigenic and molecular surveillance.     

2014-2019年青岛市A型流感病毒进化分析

  目的   了解2014-2019年青岛市人群A型流感病毒(influenza A virus, IAV)流行病学和遗传特征。   方法   提取9 807份流感样病例咽拭子标本的病毒RNA, 采用多重实时反转录PCR方法鉴定分型; 采用一步法反转录PCR扩增IAV血凝素(hemagglutinin, HA)和神经氨酸酶(neuraminidase, NA)基因, 并进行序列测定和基因分析。   结果   2014-2019年青岛市流感病毒流行具有北半球典型季节性流感特征。夏季小量流行主要是H3N2。青岛市IAV中H1N1pdm09和H3N2分别占57.6%和42.4%, 并呈现交替流行。基因群分属于6B和3C。两者HA和NA基因进化都处在净化选择下, 但抗原进化可能具有不同的进化模式。与2009年毒株相比, 2019年青岛市H1N1pdm09和H3N2的HA分别已发生21个和30个氨基酸替换, 均主要发生在头部, 分别包含5个和18个抗原位点。H1N1pdm09检测到两株神经氨酸酶抑制剂抗性突变, 1株为H275Y突变, 另1株为S247N突变; H3N2中未发现NAI抗性突变。   结论   H1N1pdm09和H3N2是2014-2019年青岛市季节性IAV中的主要组分, 进化迅速。加强流感监测和研究十分必要, 流感疫苗株需要及时更新。     

Age-related changes in memory and effector T cells responding to influenza A/H3N2 and pandemic A/H1N1 strains in humans

Age-dependent changes in the cellular immune response have been mainly described in CD8+ T cells, with relative sparing of CD4+ T cells. We show that in older compared to young adults, effector memory and effector CD8+ T-cell subsets responding to influenza A/H3N2 challenge have diminished cytolytic activity. In contrast, effector CD4+ T-cell subsets in older adults share similar phenotypic and functional characteristics with those from young adults. Further, we observed a diminished cytolytic T-cell response to both seasonal influenza A/H3N2 and pandemic H1N1 (pH1N1) strains in older compared to young adults who had received seasonal influenza vaccine. These results are consistent with the observed rates of serious complications from seasonal and pandemic influenza infections in different age groups, and suggest that CD4+ T cells may provide a compensatory response to influenza infection when CD8+ T cells become compromised during the aging process.     

Impacts on influenza A(H1N1)pdm09 infection from cross-protection of seasonal trivalent influenza vaccines and A(H1N1)pdm09 vaccines: systematic review and meta-analyses

Cross-protection by seasonal trivalent influenza vaccines (TIVs) against pandemic influenza A H1N1 2009 (now known as A[H1N1]pdm09) infection is controversial; and the vaccine effectiveness (VE) of A(H1N1)pdm09 vaccines has important health-policy implications. Systematic reviews and meta-analyses are needed to assess the impacts of both seasonal TIVs and A(H1N1)pdm09 vaccines against A(H1N1)pdm09.We did a systematic literature search to identify observational and/or interventional studies reporting cross-protection of TIV and A(H1N1)pdm09 VE from when the pandemic started (2009) until July 2011. The studies fulfilling inclusion criteria were meta-analysed. For cross-protection and VE, respectively, we stratified by vaccine type, study design and endpoint. Seventeen studies (104,781 subjects) and 10 studies (2,906,860 subjects), respectively, reported cross-protection of seasonal TIV and VE of A(H1N1)pdm09 vaccines; six studies (17,229 subjects) reported on both. Thirteen studies (95,903 subjects) of cross-protection, eight studies (859,461 subjects) of VE, and five studies (9,643 subjects) of both were meta-analysed and revealed: (1) cross-protection for confirmed illness was 19% (95% confident interval=13-42%) based on 13 case-control studies with notable heterogeneity. A higher cross-protection of 34% (9-52%) was found in sensitivity analysis (excluding five studies with moderate/high risk of bias). Further exclusion of studies that recruited early in the pandemic (when non-recipients of TIV were more likely to have had non-pandemic influenza infection that may have been cross-protective) dramatically reduced heterogeneity. One RCT reported cross-protection of 38% (19-53%) for confirmed illness. One case-control study reported cross-protection of 50% (40-59%) against hospitalisation. (2) VE of A(H1N1)pdm09 for confirmed illness was 86% (73-93%) based on 11 case-control studies and 79% (22-94%) based on two cohort studies; VE against medically-attended ILI was 32% (8-50%) in one cohort study. TIVs provided moderate cross-protection against both laboratory-confirmed A(H1N1)pdm09 illness (based on eight case-control studies with low risk of bias and one RCT) and also hospitalisation. A finding of increased risk from seasonal vaccine was limited to cases recruited early in the pandemic. A(H1N1)pdm09 vaccines were highly effective against confirmed A(H1N1)pdm09 illness. Although cross-protection was less than the direct effect of strain-specific vaccination against A(H1N1)pdm09, TIV was generally beneficial before A(H1N1)pdm09 vaccine was available.     

Live attenuated seasonal and pandemic influenza vaccine in school-age children: A randomized controlled trial

Background

The novel influenza A(H1N1pdm09) virus emerged in North America in early 2009 and rapidly spread worldwide. In this study we report the efficacy of the live attenuated monovalent H1N1pdm09 vaccine and 2009–10 seasonal influenza vaccine in a randomized double-blind placebo-controlled trial.

Methods

We enrolled 703 children aged 7–11. Each child was randomly allocated in the ratio 3:2 to receive one dose of live attenuated monovalent H1N1pdm09 vaccine or saline placebo between November 2009 and January 2010, followed after 3–10 weeks by independent random allocation to one dose of live attenuated trivalent 2009–10 seasonal influenza vaccine or saline placebo in the same ratio. Children were followed up through September 2010 with biweekly telephone calls and symptom diaries. Seasonal and pandemic influenza infections were confirmed by virologic testing of nose and throat swabs collected during acute respiratory illnesses.

Results

Overall, 30 children had confirmed influenza including 3 (0.43%) H1N1pdm09, 10 (1.4%) seasonal A(H3N2), and 17 (2.4%) influenza B. There were no significant differences in incidence rates of H1N1pdm09 or A(H3N2) between the four study arms, but receipt of the seasonal influenza vaccine was associated with a significant reduction in risk of influenza B (p < 0.01). Vaccine efficacy against confirmed H1N1pdm09 infection associated with receipt of the monovalent H1N1pdm09 vaccine was 65% (95% confidence interval, CI: −281%, 97%). Vaccine efficacies against confirmed seasonal influenza A(H3N2) and B infection associated with receipt of the seasonal influenza vaccine were 31% (95% CI: −138%, 80%) and 96% (95% CI: 67%, 99%) respectively.

Conclusions

Vaccine efficacy was consistent with other studies of the monovalent H1N1pdm09 vaccine and seasonal influenza vaccines. Our study was underpowered to provide precise estimates of vaccine efficacy due to low incidence of influenza A viruses during the study period.     

Haemagglutinin stability was not the primary cause of the reduced effectiveness of live attenuated influenza vaccine against A/H1N1pdm09 viruses in the 2013–2014 and 2015–2016 seasons

During the 2013–2014 influenza season, the quadrivalent live attenuated influenza vaccine (QLAIV), had lower than expected vaccine effectiveness (VE) against circulating A/H1N1pdm09 viruses in the USA. The underlying reason proposed for this was that the A/H1N1pdm09 vaccine strain, A/California/07/2009 (A/CA09), had a thermally unstable haemagglutinin (HA) protein. Consequently, a new A/H1N1pdm09 candidate strain, A/Bolivia/559/2013 (A/BOL13), was developed for inclusion in the 2015–2016 QLAIV. A key parameter for selection of A/BOL13 was its more thermostable HA phenotype compared with A/CA09. During the 2015–2016 season, QLAIV containing A/BOL13 was found in some studies to have improved, but still with suboptimal, VE against circulating A/H1N1pdm09 viruses and was not recommended for use by the CDC in the US market in the 2016–2017 influenza season. This suggested that improved HA thermostability had not entirely resolved the reduced VE observed. One hypothesis for this was that, by improving thermostability, the A/BOL13 HA protein had been over-stabilised, compromising its activation at the low endosomal pH required for successful viral entry. Here we demonstrate that, while the A/BOL13 HA protein is more stable than that of A/CA09, its thermal and pH stability were comparable with historically efficacious LAIV strains, suggesting that the HA had not been over-stabilised. Furthermore, studies simulating potential heat exposure during distribution by exposing QLAIV nasal sprayers to 33 °C for 4 h showed that, while remaining within product specification, A/CA09 viral potency was statistically decreased after 12 weeks at 2–8 °C. These data suggest that although unfavourable HA protein stability may have contributed to the reduced VE of A/CA09 in 2013–2014, it was unlikely to have affected A/BOL13 in 2015–2016. We conclude that HA stability was not the primary cause of the reduced effectiveness of LAIV against A/H1N1pdm09 viruses in the 2013–2014 and 2015–2016 seasons.     

2018-2019年宁夏甲型H1N1流感病毒血凝素基因组序列分析

目的 了解宁夏2018-2019流感监测年度流感病毒病原学检测情况,分析甲型H1N1流感病毒血凝素（HA）基因特征。方法 采用real time RT-PCR方法对流感监测哨点医院采集的流感样病例（ILI）标本进行核酸检测;对阳性标本进行毒株分离;提取甲型H1N1毒株的RNA,采用RT-PCR方法扩增HA片段并测序,利用生物信息软件对测序结果进行比对分析。结果 宁夏流感网络实验室检测咽拭子标本共5214份,核酸检测阳性数为760份,其中甲型H1N1阳性数为485份,占总阳性数的63.82%,分离出甲型H1N1毒株 161株。宁夏分离毒株与疫苗株A/Califaoria/07/2009不在同一进化分支,同源性为92.6%~96.3%;与疫苗株A/Michigan/45/2015(H1N1)为同一进化分支,同源性为96.6%~98.1%。与疫苗株A/Califaoria/07/2009比较,抗原位点、受体结合位点及其他位点均有变异,除毒株 A/Ningxia_Xixia/SWL1176/2019(H1N1)第222位氨基酸发生D222G变异外,其他甲型H1N1流感毒株均未发生D222G变异。所有毒株增加糖基化位点162NQT,个别毒株糖基化位点增加2~3个。结论 宁夏2018 -2019年度流感优势毒株为甲型H1N1毒株。序列分析表明甲型H1N1病毒发生了不同程度的变异,在抗原特异性、毒力和感染性上有可能已经发生变化,需要及时更换疫苗株成分。     

Squalene-containing licensed adjuvants enhance strain-specific antibody responses against the influenza hemagglutinin and induce subtype-specific antibodies against the neuraminidase

While seasonal influenza vaccines are usually non-adjuvanted, H1N1pdm09 vaccines were formulated with different squalene-containing adjuvants, to enable the reduction of antigen content thus increasing the number of doses available. To comparatively assess the effects of these adjuvants on antibody responses against matched and mismatched strains, and to correlate antibody levels with protection from disease, ferrets were immunized with 2 μg of commercial H1N1pdm09 vaccine antigen alone or formulated with different licensed adjuvants. The use of squalene-containing adjuvants increased neutralizing antibody responses around 100-fold, and resulted in a significantly reduced viral load after challenge with a matched strain. While all animals mounted strong total antibody responses against the homologous H1N1 hemagglutinin (HA) protein, which correlated with the respective neutralizing antibody titers, no reactivity with the divergent H3, H5, H7, and H9 proteins were detected. Only the adjuvanted vaccines also induced antibodies against the neuraminidase (NA) protein, which were able to also recognize NA proteins from other N1 carrying strains. These findings not only support the use of squalene-containing adjuvants in dose-sparing strategies but also support speculations that the induction of NA-specific responses associated with the use of these adjuvants may confer partial protection to heterologous strains carrying the same NA subtype.     

New genetic variants of influenza A(H1N1)pdm09 detected in Cuba during 2011–2013

Influenza A(H1N1)pdm09 virus has evolved continually since its emergence in 2009. For influenza virus strains, genetic changes occurring in HA1 domain of the hemagglutinin cause the emergence of new variants. The aim of our study is to establish genetic associations between 35 A(H1N1)pdm09 viruses circulating in Cuba in 2011–2012 and 2012–2013 seasons, and A/California/07/2009 strain recommended by WHO as the H1N1 component of the influenza vaccine. The phylogenetic analysis revealed the circulation of clades 3, 6A, 6B, 6C and 7. Mutations were detected in the antigenic site or in the receptor-binding domains of HA1 segment, including S174P, S179N, K180Q, S202T, S220T and R222K. Substitutions S174P, S179N, K180Q and R222K were detected in Cuban strains for the first time.     

Pandemic and seasonal H1N1 influenza hemagglutinin-specific T cell responses elicited by seasonal influenza vaccination

Understanding whether seasonal influenza vaccines can elicit antibody and T cell responses against the 2009 pandemic H1N1 strain is important. We compared T cell and antibody responses elicited by trivalent inactivated influenza vaccine (TIV) and live attenuated influenza vaccine (LAIV) in healthy adults. Both vaccines boosted pre-existing T cells to the seasonal and pandemic hemagglutinin (HA) but responses were significantly greater following immunization with LAIV. Antibody titers were significantly boosted only by TIV. The relationship between antibody and T cell responses and the effect of the magnitude of pre-existing immunity on vaccine-induced responses were also evaluated. Cross reactive T cell responses to the pandemic H1N1 HA existed among the cohort before the circulation of the virus to varying degrees and these responses were boosted by seasonal vaccination.     

A Historical Perspective of Influenza A(H1N2) Virus

The emergence and transition to pandemic status of the influenza A(H1N1)A(H1N1)pdm09) virus in 2009 illustrated the potential for previously circulating human viruses to re-emerge in humans and cause a pandemic after decades of circulating among animals. Within a short time of the initial emergence of A(H1N1)pdm09 virus, novel reassortants were isolated from swine. In late 2011, a variant (v) H3N2 subtype was isolated from humans, and by 2012, the number of persons infected began to increase with limited person-to-person transmission. During 2012 in the United States, an A(H1N2)v virus was transmitted to humans from swine. During the same year, Australia recorded its first H1N2 subtype infection among swine. The A(H3N2)v and A(H1N2)v viruses contained the matrix protein from the A(H1N1)pdm09 virus, raising the possibility of increased transmissibility among humans and underscoring the potential for influenza pandemics of novel swine-origin viruses. We report on the differing histories of A(H1N2) viruses among humans and animals.