Pandemic H1N1 influenza virus causes a stronger inflammatory response than seasonal H1N1 influenza virus in ferrets

A 2009 H1N1 influenza virus pandemic, which had its origin in swine, caused severe illness and mortality in humans. Inflammatory responses may be responsible for pathogenesis caused by infection with influenza viruses. To better understand the pathogenic mechanism, clinical signs and inflammatory responses in ferrets infected with the pandemic H1N1 were compared with those caused by seasonal H1N1 influenza virus. Ferrets infected with the 2009 pandemic H1N1 virus displayed higher body temperatures, greater reduction in body weight, and higher viral titers in the tracheae and lungs. Levels of inflammatory cytokines, including interleukin-6, interferon-alpha, and tumor necrosis factor-alpha, were higher in the lungs of ferrets infected with the 2009 pandemic H1N1. The data support the idea that increased pathogenesis caused by the 2009 pandemic H1N1 influenza virus may have been partially mediated by a higher induction of pro-inflammatory cytokines in the lungs of affected humans or animals.     

Previous H1N1 influenza A viruses circulating in the Mongolian population

Summary Four influenza A viruses of the subtype H1N1, isolated from Mongolian patients in Ulaanbaatar between 1985 and 1991, were analysed by sequencing of various RNA segments. The isolate from 1985 was found to be highly related in all genes sequenced to strains isolated from camels in the same region and at about the same time. These camel isolates were presumably derived from a UV-light inactivated reassortant vaccine (PR8 × USSR/77) prepared in Leningrad in 1978 and used in the Mongolian population at that time [19]. The human isolate from 1988 was also found to be a derivative of a reassortant between PR8 and USSR/77; in contrast to the 1985 isolate, however, it contained an HA closely related to PR8. One of the Mongolian isolates from 1991 (111/91) was in all genes sequenced closely related to PR8, while the other isolate from 1991 (162/91) was closely related to H1N1 strains isolated around 1986 in other parts of the world. About 12% of 235 convalescent sera collected in various parts of Mongolia contained antibodies against PR8, while none of German control sera contained such antibodies. The mutational and evolutionary rates of the Mongolian strains seem to be significantly lower when compared to the rates of human influenza A strains isolated in other parts of the world. This might indicate that these rates depend to a certain extent on the population density. Thus, viruses from remote areas might keep the potential to reappear in the human population after several years to cause a pandemic as it had happened in 1977.     

96例甲型H1N1流感的临床分析

Objective To review the epidemiologic and clinical characteristics of 96 cases with novel H1N1 influenza A.and improve the diagnosis and treatment level of novel H1N1 influenza A.Methods 96 cases of novel H1N1 influenza A admitted to the isolation wards from Oct 20 to Sep 23,2009 were studied.Their epidemioiogic,clinical,laboratory,and radiologic characteristics were analyzed. Results The median age of the 96 patients was 26.52 ±10.62 years(range,5 to 60 years).Sixty-four of the 96 patients had a close contact with novel H1N1 influenza A patients.The main symptoms included fever 100%,cough 86.4%,sore throat 66.6% and myalgia 32.3%.Conclusion The clinical presentation of novel H1N1 infection is largely indistinguishable from that of seasonal influenza.Combines beth a symptom complex with the epidemiological investigation and laboratory characteristics can improve the accuracy of diagnosis of novel H1N1 influenza A.     

96例甲型H1N1流感的临床分析

目的 通过回顾性分析96例甲型HlNl流感的流行病学及临床特点,提高对甲型H1Nl流感的诊断和治疗水平.方法 收集2009年8月20日至9月23日于杭州市第六人民医院住院治疗的96例甲型H1N1流感患者的临床资料,对其流行病学及临床特点进行分析.结果 患者平均年龄（26.52±10.62）岁,64例有甲流接触史.以发热100%、咳嗽86.4%、咽痛66.6%和肌肉酸痛32.3%为主要临床表现.结论 甲型H1N1流感临床表现与季节性流感很难区分,需结合流行病学调查与实验室检查及时诊断隔离治疗.     

96例甲型H1N1流感的临床分析

Objective To review the epidemiologic and clinical characteristics of 96 cases with novel H1N1 influenza A.and improve the diagnosis and treatment level of novel H1N1 influenza A.Methods 96 cases of novel H1N1 influenza A admitted to the isolation wards from Oct 20 to Sep 23,2009 were studied.Their epidemioiogic,clinical,laboratory,and radiologic characteristics were analyzed. Results The median age of the 96 patients was 26.52 ±10.62 years(range,5 to 60 years).Sixty-four of the 96 patients had a close contact with novel H1N1 influenza A patients.The main symptoms included fever 100%,cough 86.4%,sore throat 66.6% and myalgia 32.3%.Conclusion The clinical presentation of novel H1N1 infection is largely indistinguishable from that of seasonal influenza.Combines beth a symptom complex with the epidemiological investigation and laboratory characteristics can improve the accuracy of diagnosis of novel H1N1 influenza A.     

Differential susceptibility of different cell lines to swine-origin influenza A H1N1, seasonal human influenza A H1N1, and avian influenza A H5N1 viruses

BackgroundThe novel swine-origin influenza A H1N1 virus (S-OIV) causes the current pandemic. Its tissue tropism and replication in different cell lines are not well understood.ObjectiveCompare the growth characteristics of cell lines infected by S-OIV, seasonal influenza A H1N1 (sH1N1) and avian influenza A H5N1 (H5N1) viruses and the effect of temperature on viral replication.Study designCytopathic effect (CPE), antigen expression by immunofluorescence (IF) and viral load profile by quantitative RT-PCR in 17 cell lines infected by S-OIV, sH1N1 and H5N1 were examined. Comparison of their replication efficiency in chick embryo was performed. The effect of temperature on viral replication in Madin–Darby canine kidney (MDCK) cells was determined by TCID₅₀ at 33 °C, 37 °C and 39 °C for 5 consecutive days.ResultsS-OIV replicated in cell lines derived from different tissues or organs and host species with comparable viral load to sH1N1. Among 13 human cell lines tested, Caco-2 has the highest viral load for S-OIV. S-OIV showed a low viral load with no CPE or antigen expression in pig kidney cell PK-15, H5N1 demonstrated the most diverse cell tropism by CPE and antigen expression, and the highest viral replication efficiency in both cell lines and allantoic fluid. All three viruses demonstrated best growth at 37 °C in MDCK cells.ConclusionCell line growth characteristics of S-OIV, sH1N1 and H5N1 appear to correlate clinically and pathologically with involved anatomical sites and severity. Low replication of S-OIV in PK-15 suggests that this virus is more adapted to human than swine.     

我国人群中一种新重配的H1N2亚型流感病毒株的发现

１９９６年１月太原铁路卫生防疫站从上感患者中分离到３株流感病毒。经血清学鉴定，它们不同于１９８９和１９９２年所发现的Ｈ１Ｎ２亚型毒株，其ＨＡ的抗原性类似于Ａ／ＰＲ／８／３４（Ｈ１Ｎ１）病毒，而明显不同于当前人群中流行的Ｈ１Ｎ１亚型毒株。病毒粒不同基因节段迁移率比较表明，它们的１～４基因节段迁移率接近于Ａ／ＰＲ／８／３４（Ｈ１Ｎ１）毒株，５～６基因节段迁移率类似于Ａ／武汉／３５９／９５（Ｈ３Ｎ２）病毒，而７～８两节段既不同于Ａ／ＰＲ／８／３４（Ｈ１Ｎ１），又不同于Ａ／武汉／３５９／９５（Ｈ３Ｎ２）病毒。故可认为它们是一种新重配的Ｈ１Ｎ２亚型毒株。     

Update on influenza diagnostics: lessons from the novel H1N1 influenza A pandemic

The menu of diagnostic tools that can be utilized to establish a diagnosis of influenza is extensive and includes classic virology techniques as well as new and emerging methods. This review of how the various existing diagnostic methods have been utilized, first in the context of a rapidly evolving outbreak of novel influenza virus and then during the different subsequent phases and waves of the pandemic, demonstrates the unique roles, advantages, and limitations of each of these methods. Rapid antigen tests were used extensively throughout the pandemic. Recognition of the low negative predictive values of these tests is important. Private laboratories with preexisting expertise, infrastructure, and resources for rapid development, validation, and implementation of laboratory-developed assays played an unprecedented role in helping to meet the diagnostic demands during the pandemic. FDA-cleared assays remain an important element of the diagnostic armamentarium during a pandemic, and a process must be developed with the FDA to allow manufacturers to modify these assays for detection of novel strains in a timely fashion. The need and role for subtyping of influenza viruses and antiviral susceptibility testing will likely depend on qualitative (circulating subtypes and their resistance patterns) and quantitative (relative prevalence) characterization of influenza viruses circulating during future epidemics and pandemics.     

一起新型甲型H1N1流感疫情的临床和实验室特点

目的分析一起新型甲型H1N1流感疫情患者的临床和实验室特点。方法某部集体发病患者35例,均由核酸检测方法确诊;回顾调查患者的密切接触史、实验室检测结果、治疗和预后等。结果本次疫情收治的35人,密切接触平均1．7d出现临床症状,首发症状为发热占97．1％（34／35）,咽痛65．7％（23／35）,咳嗽51．4％（16／35）,肌肉酸痛31．4％（11／35）,头痛28．6％（10／35）。病毒核酸检测阴性的平均天数为4．5d,奥司他韦治疗,5d标准疗程,所有患者7d后痊愈出院。结论本次疫情为比较典型的新型甲型H1N1流感。     

Innate immune response to H3N2 and H1N1 influenza virus infection in a human lung organ culture model

We studied cytokine responses to influenza virus PR8 (H1N1) and Oklahoma/309/06 (OK/06, H3N2) in a novel human lung tissue model. Exposure of the model to influenza virus rapidly activated the mitogen-activated protein kinase signaling (MAPK) pathways ERK, p38 and JNK. In addition, RNase protection assay demonstrated the induction of several cytokine and chemokine mRNAs by virus. This finding was reflected at the translational level as IL-6, MCP-1, MIP-1α/β, IL-8 and IP-10 proteins were induced as determined by ELISA. Immunohistochemistry for IP-10 and MIP-1α revealed that alveolar epithelial cells and macrophages were the source of these two cytokines. Taken together, both PR8 and OK/06 cause similar induction of cytokines in human lung, although OK/06 is less effective at inducing the chemokines MCP-1 and IL-8. This human organ culture model should thus provide a relevant platform to study the biological responses of human lung to influenza virus infection.     

Contemporary seasonal influenza A (H1N1) virus infection primes for a more robust response to split inactivated pandemic influenza A (H1N1) Virus vaccination in ferrets

Human influenza pandemics occur when influenza viruses to which the population has little or no immunity emerge and acquire the ability to achieve human-to-human transmission. In April 2009, cases of a novel H1N1 influenza virus in children in the southwestern United States were reported. It was retrospectively shown that these cases represented the spread of this virus from an ongoing outbreak in Mexico. The emergence of the pandemic led to a number of national vaccination programs. Surprisingly, early human clinical trial data have shown that a single dose of nonadjuvanted pandemic influenza A (H1N1) 2009 monovalent inactivated vaccine (pMIV) has led to a seroprotective response in a majority of individuals, despite earlier studies showing a lack of cross-reactivity between seasonal and pandemic H1N1 viruses. Here we show that previous exposure to a contemporary seasonal H1N1 influenza virus and to a lesser degree a seasonal influenza virus trivalent inactivated vaccine is able to prime for a higher antibody response after a subsequent dose of pMIV in ferrets. The more protective response was partially dependent on the presence of CD8(+) cells. Two doses of pMIV were also able to induce a detectable antibody response that provided protection from subsequent challenge. These data show that previous infection with seasonal H1N1 influenza viruses likely explains the requirement for only a single dose of pMIV in adults and that vaccination campaigns with the current pandemic influenza vaccines should reduce viral burden and disease severity in humans.     

A novel reassortant H1N2 virus related to the pandemic H1N1 2009 influenza virus isolated from Korean pigs

Since the discovery of the pandemic H1N1 (pH1N1) virus in 2009, a novel reassortant H1N2 virus (A/Swine/Korea/VDS1/2010) containing the pH1N1 segments has been detected in Korean pig populations. The hemagglutinin and neuraminidase genes of this virus are derived from reassortant H1N1- and H1N2-group viruses, respectively, identified in Korean pigs, while other genes originate from contemporary circulating pH1N1 viruses. The antigenic and biological properties of this novel virus, as determined by clinical, pathological, serological, and genetic analyses, are similar to those of pH1N1 viruses, which infect swine easily (Weingartl et al. J Virol 84:2245–2256, 2010; Brookes et al. PLoS one 5:e9068, 2010; Lange et al. J Gen Virol 90:2119–2123, 2009). Determining whether this virus will become established and pose a threat to mammalian populations requires further investigation.     

Functional immune response to influenza H1N1 in children and adults after live attenuated influenza virus vaccination

Influenza virus is a major respiratory pathogen, and vaccination is the main method of prophylaxis. In 2012, the trivalent live attenuated influenza vaccine (LAIV) was licensed in Europe for use in children. Vaccine‐induced antibodies directed against the main viral surface glycoproteins, haemagglutinin (HA) and neuraminidase (NA) play important roles in limiting virus infection. The objective of this study was to dissect the influenza‐specific antibody responses in children and adults, and T cell responses in children induced after LAIV immunization to the A/H1N1 virus. Blood samples were collected pre‐ and at 28 and 56 days post‐vaccination from 20 children and 20 adults. No increase in micro‐neutralization (MN) antibodies against A/H1N1 was observed after vaccination. A/H1N1 stalk‐specific neutralizing and NA‐inhibiting (NI) antibodies were boosted in children after LAIV. Interferon γ‐producing T cells increased significantly in children, and antibody‐dependent cellular‐mediated cytotoxic (ADCC) cell activity increased slightly in children after vaccination, although this change was not significant. The results indicate that the NI assay is more sensitive to qualitative changes in serum antibodies after LAIV. There was a considerable difference in the immune response in children and adults after vaccination, which may be related to priming and previous influenza history. Our findings warrant further studies for evaluating LAIV vaccination immunogenicity.     

Serological study of the 2009 pandemic due to influenza A H1N1 in the metropolitan French population

We looked for evidence of antibodies to the 2009 influenza A/H1N1 pandemic virus in panels of sera from individuals living in metropolitan France, obtained either before, during or after the epidemic, using standard haemagglutination inhibition and microneutralization tests. The difference between seroprevalence values measured in post- and pre-epidemic panels was used as an estimate of seroconversion rate in different age groups (23.4% (0–24 years, age-group 0); 16.5% (25–34); 7.9% (35–44); 7.2% (45–54); 1.6% (55–64); and 3.1% (<65)), confirming that the distribution of cases in different age groups was similar to that of the seasonal H1N1 virus. During the pre-pandemic period low-titre cross-reactive antibodies were present in a large proportion of the population (presumably acquired against seasonal H1N1) whereas cross-reactive antibodies were detected in individuals over the age of 65 years with significantly higher prevalence and serological titres (presumably acquired previously against Spanish flu-related H1N1 strains). Clinical data and analysis of post-pandemic seroprevalence showed that few of these latter patients were infected by the influenza virus during the epidemic. In contrast, the majority of both clinical cases and seroconversions were recorded in the 0–24 age group and a global inverse relationship between prevalence of antibodies to pH1N1 in the pre-pandemic period and rate of seroconversion was observed amongst age groups. Our results emphasize the complex relationships involved in antigenic reactivity to pandemic and seasonal H1N1 viral antigens; hence the difficulty in distinguishing between low-titre specific and cross-reactive antibodies, establishing precise seroprevalence numbers and fully understanding the relationship between previous immunity to seasonal viruses and protection against the novel variant.     

A seroepidemiological study of pandemic A/H1N1(2009) influenza in a rural population of Mali

The swine-origin H1N1 influenza A virus (pH1N1(2009)) started to circulate worldwide in 2009, and cases were notified in a number of sub-Saharan African countries. However, no epidemiological data allowing estimation of the epidemic burden were available in this region, preventing comprehensive comparisons with other parts of the world. The CoPanFlu-Mali programme studied a cohort of 202 individuals living in the rural commune of Dioro (southern central Mali). Pre-pandemic and post-pandemic paired sera (sampled in 2006 and April 2010, respectively) were tested by the haemagglutination inhibition (HI) method. Different estimates of pH1N1(2009) infection during the 2009 first epidemic wave were used (increased prevalence of HI titre of ≥ 1/40 or ≥ 1/80, seroconversions) and provided convergent attack rate values (12.4-14.9%), the highest values being observed in the 0-19-year age group (16.0-18.4%). In all age groups, pre-pandemic HI titres of ≥1/40 were associated with complete absence of seroconversion; and geometric mean titres were <15 in individuals who seroconverted and >20 in others. Important variations in seroconversion rate existed among the different villages investigated. Despite limitations resulting from the size and composition of the sample analysed, this study provides strong evidence that the impact of the pH1N1(2009) first wave was more important than previously believed, and that the determinants of the epidemic spread in sub-Saharan populations were quite different from those observed in developed countries.     

Impaired dendritic cell maturation in response to pandemic H1N109 influenza virus

BackgroundInfection with pandemic A/H1N1/2009 influenza virus led to hospitalisation of patients not expected to be at risk of severe disease from seasonal influenza infection.ObjectivesWe sought to establish whether (i) DC maturation was compromised in patients experiencing severe pandemic influenza infection, (ii) the pandemic virus differed from seasonal influenza virus in its ability to induce DC maturation and (iii) there was an associated inability to activate memory B cells or induce antibody.Study designPeripheral blood mononuclear (PBMCs) cells were sampled from individuals with confirmed acute pandemic A/H1N1/2009 influenza infection or from healthy vaccinated controls. DCs were differentiated from the PBMC and tested for their ability to mature following stimulation with pandemic virus, seasonal H3N2 influenza virus or LPS. Serum samples from the patients were used to assess seroconversion to influenza and the levels of influenza specific memory B cells in PBMC were also determined.ResultsDCs obtained from all individuals exhibited negligible maturation marker upregulation when exposed to pandemic A/H1N1/2009 virus but showed a strong response to the seasonal H3N2 virus and LPS. Robust levels of memory B cell were obtained in both groups and patients seroconverted to the virus.ConclusionsOverall, the ability of patient's DC to mature in response to different stimuli was no different to that of control subjects DCs. Importantly, panH1N109 virus failed to induce substantial DC maturation in any individual, contrasting with seasonal virus, but this did not result in failure to mount memory B cell and antibody responses to the virus.