Novel H1N1 influenza A virus infection in a patient with acute rejection after liver transplantation

BACKGROUND:The 2009 H1N1 influenza A virus was first identified in April 2009 and rapidly evolved into a pandemic. Recipients of solid-organ transplants have a higher risk for severe infection because of immunosuppression.There are limited reports of 2009 H1N1 influenza in liver transplant recipients,especially in China. METHODS:We present a case of a 48-year-old male liver transplant recipient with 2009 H1N1 influenza A virus.He received therapy for acute rejection after transplantation and was confirmed with H1N1 virus infection. RESULTS:The patient was started on oseltamivir(75 mg, orally twice daily)and had a benign hospital course,with defervescence and resolution of symptoms within 72 hours. The follow-up chest radiograph after discharge was normal. CONCLUSIONS:The 2009 H1N1 influenza in this hospitalized transplant recipient was relatively mild,and prolonged viral shedding was not noted.Oseltamivir can be a valid measure in immunocompromised individuals.

Traditional Chinese medicine as monotherapy or combined with oseltamivir in the treatment of H1N1 influenza:a systematic review and meta-analysis

Objective This study aimed to assess the efficacy and safety of traditional Chinese medicine,alone or in combination with oseltamivir,in patients with H1N1 Influenza.Methods In the present study,we searched the Cochrane Central Register of Controlled Trials,PUBMED,EMBASE,Chinese Biomedical Literature Database,China Science and Technology Journal Database,China National Knowledge Infrastructure Database,and WanFang Data for studies published in or before February 8,2022.Data were extracted and checked by two investigators.Review Manager 5.4 and STATA statistical software 16.0 were used for the data analysis.Results We identified 22 individual studies reporting data from 2292 individuals with H1N1 influenza.Compared with oseltamivir,the fever clearance duration[MD=-3.99,95%CI(-6.89,-1.09)]and sore throat relief time[MD=-5.39,95%CI(-10.19,-0.59)]in the intervention group of traditional Chinese medicine monotherapy or combined with oseltamivir were shorter.Maxingshigan was the primary component of Lianhuaqingwen.The subgroup analyses indicated that Maxingshigan shortened fever clearance time[MD=-3.23,95%CI(-5.60,-0.85)],and also had certain advantages in relieving sore throat[RR=-4.55,95%CI(-10.04,0.95)].However,as for the effective rate,fever duration,cough disappearance time,hospital length of stay,clinical symptoms time as well as viral shedding duration,there were no significant differences between the two groups.Besides,no serious adverse effects were reported in the included studies.Conclusion Although we couldn’t get a definitive conclusion due to the small sample sizes and high risk of bias in the included studies,most traditional Chinese medicine showed similar effects to oseltamivir in treating H1N1 influenza.Some were showed to have a statistically significant shorter time of fever clearance and sore throat remission when they were used alone or in combination with oseltamivir and were well-tolerated treatment,such as Maxingshigan.

The interaction between the 2009 H1N1 influenza A hemagglutinin and neuraminidase: mutations, co-mutations, and the NA stalk motifs

As the world is closely watching the current 2009 H1N1 pandemic unfold, there is a great interest and need in understanding its origin, genetic structures, virulence, and pathogenicity. The two surface proteins, hemagglutinin (HA) and neuraminidase (NA), of the influenza virus have been the focus of most flu research due to their crucial biological functions. In our previous study on 2009 H1N1, three aspects of NA were investigated: the mutations and co-mutations, the stalk motifs, and the phylogenetic analysis. In this study, we turned our attention to HA and the interaction between HA and NA. The 118 mutations of 2009 H1N1 HA were found and mapped to the 3D homology model of H1, and the mutations on the five epitope regions on H1 were identified. This information is essential for developing new drugs and vaccine. The distinct response patterns of HA to the changes of NA stalk motifs were discovered, illustrating the functional dependence between HA and NA. With help from our previous results, two co-mutation networks were uncovered, one in HA and one in NA, where each mutation in one network co-mutates with the mutations in the other network across the two proteins HA and NA. These two networks residing in HA and NA separately may provide a functional linkage between the mutations that can impact the drug binding sites in NA and those that can affect the host immune response or vaccine efficacy in HA. Our findings demonstrated the value of conducting timely analysis on the 2009 H1N1 virus and of the integrated approach to studying both surface proteins HA and NA together to reveal their interdependence, which could not be accomplished by studying them individually.

Subtle differences in receptor binding specificity and gene sequences of the 2009 pandemic H1N1 influenza virus

A recent phylogenetic inference indicated that the 2009 pandemic H1N1 strains circulating from March 2009 to September 2009 could be divided into two closely related but distinct clusters. Cluster one contained most strains from Mexico, Texas, and California, and cluster two had most strains from New York, both of which were reported to co-circulate in all continents. The same study further revealed nine nucleotide changes in six gene segments of the new virus specific for the two clusters. In the current study, the informational spectrum method (ISM), a bioinformatics technique, was employed to study the receptor binding patterns of the two clusters. It discovered that while both groups shared the same primary human binding affinity, their secondary binding preferences were different. Cluster one favored swine binding as its secondary binding pattern, whereas cluster two mostly exhibited the binding specificity of A/South Carolina/1/18 (H1N1) (one of the 1918 flu pandemic strains) as its secondary binding pattern. Besides all the nine nucleotide changes found in the previous study, Random Forests were applied to uncover several new nucleotide polymorphisms in 10 genes of the strains between the two clusters, and several amino acid changes in the HA protein that might be accountable for the discrepancy of the secondary receptor binding patterns of the two clusters. Finally, entropy analysis was conducted to present a global view of gene sequence variations between the two clusters, which illustrated that cluster one had much higher genetic divergence than cluster two. Furthermore, it suggested a significant overall correspondence between the nucleotide positions of high importance in differentiating the two clusters and nucleotide positions of high entropy in cluster one.

Gene Expression Profiles Comparison between 2009 Pandemic and Seasonal H1N1 Influenza Viruses in A549 Cells

Objective To perform gene expression profiles comparison so that to identify and understand the potential differences in pathogenesis between the pandemic and seasonal A （H1N1） influenza viruses. Methods A549 cells were infected with A/California/07/09 （H1N1） and A/GuangdongBaoan/51/08 （H1N1） respectively at the same MOI of 2 and collected at 2, 4, 8, and 24 h post infection （p.i.）. Gene expression profiles of A549 cells were obtained using the 22 K Human Genome Oligo Array, and differentially expressed genes were analyzed at selected time points. Results Microarrays results indicated that both of the viruses suppressed host immune response related pathways including cytokine production while pandemic H1N1 virus displayed weaker suppression of host immune response than seasonal H1N1 virus. Observation on similar anti-apoptotic events such as activation of apoptosis inhibitor and down-regulation of key genes of apoptosis pathways in both infections showed that activities of promoting apoptosis were different in later stage of infection. Conclusion The immuno-suppression and anti-apoptosis events of pandemic H1N1 virus were similar to those seen by seasonal H1N1 virus. The pandemic H1N1 virus had an ability to inhibit biological pathways associated with cytokine responses, NK activation and macrophage recognition .

Successful High-Dosage Dexamethasone Treatment of H1N1 Influenza A Pneumonia Complicated with Acute Necrotizing Encephalitis in an HIV-Infected Adult

Neurological manifestations in H1N1 influenza A infection are very rare, especially in adults, and its mechanism of action is still uncertain. Here, we reported the case of a 53-year-old woman with human immunodeficiency virus infection (HIV) who had H1N1 influenza A pneumonia complicated with very rare acute necrotizing encephalitis, although the HIV was under control. With prompt identification and administration of high dosage of dexamethasone, her mental status improved from stupor to clear, with minimal right hemiparesis. Further, brain magnetic resonance image revealed great resolution of mass effect. This dramatic improvement in response to the treatment may improve our understanding of the pathophysiology between H1N1 influenza A infection and acute necrotizing encephalitis.

Higher Viral Load and Prolonged Viral Shedding Period is Associated with Impaired Th17 Cell Response in Patients with H1N1 Influenza A

Objective To explore whether age,disease severity,cytokines and lymphocytes in H1N1 influenza A patients correlate with viral load and clearance.Methods Total of 70 mild and 16 severe patients infected with H1N1 influenza A virus were enrolled in this study.Results It was found that the patients under 14 years old and severe patients displayed significantly higher viral loads and prolonged viral shedding periods compared with the patients over 14 years old and mild patients,respectively(P < 0.05).Moreover,the patients under 14 years old and severe patients displayed significantly lower Th17 cell frequency than the patients over 14 years old and mild patients(P < 0.01).The viral shedding period inversely correlated with the frequency of IL-17+IFN-γ-CD4+ T cells.Additionally,the decreased concentration of serum TGF-β correlated with the decreased frequency of IL-17+IFN-γ-CD4+ T cells.Conclusions Both younger and severe patients are associated with higher viral loads and longer viral shedding periods,which may partially be attributed to the impaired Th17 cell response.

Novel host markers in the 2009 pandemic H1N1 influenza a virus

The winter of 2009 witnessed the concurrent spread of 2009 pandemic H1N1 with 2009 seasonal H1N1. It is clinically important to develop knowledge of the key features of these two different viruses that make them unique. A robust pattern recognition technique, Random Forests, was employed to uncover essential amino acid markers to differentiate the two viruses. Some of these markers were also part of the previously discovered genomic signature that separate avian or swine from human viruses. Much research to date in search of host markers in 2009 pandemic H1N1 has been primarily limited in the context of traditional markers of avian-human or swine-human host shifts. However, many of the molecular markers for adaptation to human hosts or to the emergence of a pandemic virus do not exist in 2009 pandemic H1N1, implying that other previously unrecognized molecular determinants are accountable for its capability to infect humans. The current study aimed to explore novel host markers in the proteins of 2009 pandemic H1N1 that were not present in those classical markers, thus providing fresh and unique insight into the adaptive genetic modifications that could lead to the generation of this new virus. Random Forests were used to find 18 such markers in HA, 15 in NA, 9 in PB2, 11 in PB1, 13 in PA, 10 in NS1, 1 in NS2, 11 in NP, 3 in M1, and 1 in M2. The amino acids at many of these novel sites in 2009 pandemic H1N1 were distinct from those in avian, human, and swine viruses that were identical at these positions, reflecting the uniqueness of these novel sites.

Screening of Potent Inhibitor of H1N1 Influenza NS1 CPSF30 Binding Pocket by Molecular Docking

The swine flu, H1N1 virus was outbroken in Mexico and the United States in April 2009 and then rapidly spread worldwide. The World Health Organization declared that the outbreak of influenza is caused by a new subtype of influenza H1N1 influenza virus. And researchers have isolated some oseltamivir resistance strains in 2009 swine flu which makes the imminency of research and development of new anti influenza drug. The CPSF30 binding pocket of effector domain in NS1 protein is very important in the replication of influanza A virus and is a new attractive anti flu drug target. But up to now there is no antiviral drug target this pocket. Here we employ molecular docking to screening of about 200,000 compounds. We find four novel compounds with high binding energy. Binding comformation analysis revealed that these small molecules can interact with the binding pocket by some strong hydrophobic interaction. This study find some novel small molecules can be used as lead compounds in the development of new antiinfluenza drug based on CPSF30 pocket.

Intergenic subset organization within a set of geographically-defined viral sequences from the 2009 H1N1 influenza A pandemic

We report a bioinformatic analysis of the datasets of sequences of all ten genes from the 2009 H1N1 influenza A pandemic in the state of Wisconsin. The gene with the greatest summed information entropy was found to be the hemagglutinin (HA) gene. Based upon the viral ID identifier of the HA gene sequence, the sequences of all of the genes were sorted into two subsets, depending upon whether the nucleotide occupying the position of maximum entropy, position 658 of the HA sequence, was either A or U. It was found that the information entropy (H) distributions of subsets differed significantly from each other, from H distributions of randomly generated subsets and from the H distributions of the complete datasets of each gene. Mutual information (MI) values facilitated identification of nine nucleotide positions, distributed over seven of the influenza genes, at which the nucleotide subsets were disjoint, or almost disjoint. Nucleotide frequencies at these nine positions were used to compute mutual information values that subsequently served as weighting factors for edges in a graph net-work. Seven of the nucleotide positions in the graph network are sites of synonymous mutations. Three of these sites of synonymous mutation are within a single gene, the M1 gene, which occupied the position of greatest graph centrality. It is proposed that these bioinformatic and network graph results may reflect alterations in M1-mediated viral packaging and exteriorization, known to be susceptible to synonymous mutations.

Distinctive clinical and laboratory features of COVID-19 and H1N1 influenza infections among hospitalized pediatric patients

Background It had been documented in many studies that pediatric coronavirus disease 2019(COVID-19)is characterized by low infectivity rates,low mortalities,and benign disease course.On the other hand,influenza type A viruses are recognized to cause severe and fatal infections in children populations worldwide.This study is aimed to compare the clinical and laboratory characteristics of COVID-19 and H1N1 influenza infections.Methods A retrospective study comprising 107 children hospitalized at Abha Maternity and Children Hospital,Southern region of Saudi Arabia,with laboratory-confirmed COVID-19 and H1N1 influenza infections was carried out.A complete follow-up for all patients from the hospital admission until discharge or death was made.The clinical data and laboratory parameters for these patients were collected from the medical records of the hospital.Results Out of the total enrolled patients,73(68.2%)were diagnosed with COVID-19,and 34(31.8%)were diagnosed with H1N1 influenza.The median age is 12 months for COVID-19 patients and 36 months for influenza patients.A relatively higher number of patients with influenza had a fever and respiratory symptoms than COVID-19 patients.In contrast,gastrointestinal symptoms were observed in a higher number of COVID-19 patients than in influenza patients.A statistically significant increase in white cell counts is noted in COVID-19 but not in influenza patients(P<0.05).There are no obvious variations in the mean period of duration of hospitalization between COVID-19 and influenza patients.However,the total intensive care unit length of stay was longer for influenza compared to COVID-19 patients.Conclusions A considerable number of children infected with COVID-19 and H1N1 influenza were noted and reported in this study.There were no significant variations in the severity of the symptomatology and laboratory findings between the two groups of patients.Significant differences between these patients in some hospitalization factors and diagnosis upon admission also were not observed.However,more severe clinical manifestations and serious consequences were observed among pediatric patients hospitalized with influenza infections than among those with COVID-19.

Insights from investigating the interactions of natural product inhibitors with neuraminidase of the 2009 H1N1 influenza virus

Neuraminidase(NA) plays a biologically vital role in the replication of influenza virus, and NA inhibitors(NAIs) are most widely used in the clinical anti-flu therapy. NA of 2009 H1 N1 influenza virus(09 N1) possesses a different substrate-binding cavity compared with other NA subtypes, making 09 N1 a more appropriate starting point for the discovery of potent 09 N1 inhibitors. As natural products are of great structural diversity, research on the interaction between natural NAIs and 09 N1 can throw light on the design of new structural NAIs. In this study, we, for the first time, conducted molecular docking procedure with GOLD on 10 natural inhibitors to 09 N1, and acquired their binding modes with 09 N1. The docking results showed that the active site S1 was important in the binding of NAIs to 09 N1. Then five scaffolds were extracted from these NAIs with interactions to site S1, and these could be used in the structural modification of NAIs. Besides, we found that the addition of H-bonding interaction with the active site could improve the NA inhibitory activity of NAIs, and it might be the reason why the approved NAIs showed high efficiency. Two terminal hydrophobic sites(Terminal 1 and Terminal 2) with no interactions to the approved NAI zanamivir were found in the 09 N1 active cavity, and four NAIs were first found to bind with the terminals. Till now, there are few studies on the meaning of Terminal 2 in the binding of NAI to NA, which could be a new direction for the rational design of NAIs.

Detection and pathogenesis of a novel swine H3N2 influenza virus containing three genes from the 2009 pandemic H1N1 influenza viruses in Korea in 2015

Dear Editor,Influenza A viruses cause pandemics at an interval of approximately 10-40 years,and pigs are regarded as a＂mixing vessel＂because they are easily infected with avian and human influenza viruses（Ito et al.,1998）.According to previous studies,H3N2,H1N2,and H1N1 subtypes o（swine influenza viruses have been detected in Korean pigs （Pascua et al., 2013; Kim et al., 2014; Song et al., 2007）. Moreover, a novel H3N2 influenza virus containing the matrix （34） gene from a 2009 pandemic influenza virus was detected in Korean pigs in 2013 （Pascua et al., 2013）, an H1N2 influenza virus con- taining the internal genes from a 2009 pandemic influ- enza virus was found in Korean pigs in 2014 （Kim et al., 2014）, and an H1N1 influenza virus containing all genes from the classical swine influenza viruses was isolated from Korean pigs in 2007 （Song et al., 2007）.

感冒清热片防治甲型H1N1流感病毒感染的药效学研究

目的探讨感冒清热片防治甲型H1N1流感病毒感染的药效学情况。方法108例甲型H1N1流感病毒感染患者,随机分为观察组和对照组,每组54例。对照组使用磷酸奥司他韦治疗,观察组在对照组治疗基础上使用感冒清热片治疗。比较两组患者临床疗效、症状改善指标(起效时间、退热时间、止咳时间、痊愈时间)、以及治疗前后的中医证候(发热、咽痛、头痛、鼻塞流涕、咳嗽、乏力)积分、血清炎性因子[C反应蛋白(CRP)、肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)]。结果观察组治疗总有效率96.30%高于对照组的77.78%(P<0.05)。治疗后,观察组患者发热、咽痛、头痛、鼻塞流涕、咳嗽、乏力评分分别为(0.83±0.24)、(1.12±0.30)、(1.07±0.29)、(1.03±0.32)、(1.05±0.33)、(1.09±0.28)分,均低于对照组的(1.46±0.37)、(2.31±0.58)、(2.14±0.52)、(2.25±0.63)、(2.36±0.61)、(2.42±0.65)分(P<0.05)。观察组起效时间(1.03±0.34)d、退热时间(1.56±0.40)d、止咳时间(4.35±0.68)d、痊愈时间(6.63±1.26)d均短于对照组的(2.32±0.57)、(2.63±0.74)、(7.24±1.16)、(9.38±1.72)d(P<0.05)。治疗后,观察组CRP(2.11±0.56)mg/L、TNF-α(19.34±2.20)μg/L、IL-6(4.02±0.54)ng/L均低于对照组的(4.83±0.69)mg/L、(25.89±2.73)μg/L、(6.13±0.62)ng/L(P<0.05)。结论感冒清热片防治甲型H1N1流感病毒感染的疗效显著,能有效缓解证候,加快症状缓解速度,提高抗炎作用。

射干止咳胶囊体内外抗甲型H1N1流感病毒作用初探

目的初步探讨射干止咳胶囊对甲型H1N1流感病毒的体内外抗病毒作用。方法通过细胞病变效应(cytopathic effect,CPE)结合MTT法考察射干止咳胶囊体外抑制甲型H1N1流感病毒活性的作用;采用小鼠滴鼻法感染甲型H1N1流感病毒为模型,观察射干止咳胶囊对甲型H1N1流感病毒的体内抗病毒作用。结果体外抗病毒试验结果显示,射干止咳胶囊对甲型H1N1流感病毒的最高抑制率为11.86%,提示在攻毒剂量为100TCID_(50)的条件下,射干止咳胶囊在体外对甲型H1N1流感病毒的抑制作用不明显。小鼠滴鼻感染5LD_(50)的甲型H1N1流感病毒,射干止咳胶囊低、中、高剂量组小鼠的死亡率分别为30%、10%、30%,死亡保护率达50%、70%、50%,平均生存天数分别为13 d、14 d、13 d,说明各给药组对攻毒小鼠具有较好的死亡保护作用。结论射干止咳胶囊体外抗甲型H1N1流感病毒的作用不显著,但射干止咳胶囊各剂量组对甲型H1N1流感病毒攻毒致小鼠死亡却具有明显的保护作用。

甲型流感病毒H1N1型灭活病毒标准物质研究

以人工培养真实病毒为原料,研制了一种甲型流感病毒H1N1型灭活病毒标准物质。该标准物质经化学灭活及灭活效果验证后,联合多家实验室采用数字PCR定量检测方法对拷贝数浓度进行定值,对标准物质的均匀性、稳定性及不确定度进行了评估。结果显示该标准物质均匀性良好,在-80~-70℃保存条件下稳定保存5个月,标准物质最终定值结果为(1384.5±249.3)copies/μL(k=2)。该标准物质可为H1N1型甲型流感病毒检测的全过程提供计量溯源和质量控制支撑,有助于提升相关核酸检测结果的准确性和可靠性。

The novel H1N1 Influenza A global airline transmission and early warning without travel containments

A novel influenza A (H1N1) has been spreading worldwide. Early studies implied that international air travels might be key cause of a severe potential pandemic without appropriate containments. In this study, early outbreaks in Mexico and some cities of United States were used to estimate the preliminary epidemic parameters by applying adjusted SEIR epidemiological model, indicating transmissibility infectivity of the virus. According to the findings, a new spatial allocation model totally based on the real-time airline data was established to assess the potential spreading of H1N1 from Mexico to the world. Our estimates find the basic reproductive number R0 of H1N1 is around 3.4, and the effective reproductive number fall sharply by effective containment strategies. The finding also implies Spain, Canada, France, Panama, Peru are the most possible country to be involved in severe endemic H1N1 spreading.

Two cases of exudative retina detachment and uveitis following H1N1 influenza vaccination

Uveitis was a rare adverse event of vaccination. We met two cases of acute uveitis with exudative retinal detachment following vaccination of H1 N1 influenza. Case 1 was a 10-year-old boy who was admitted for bilateral blurred vision at 10 days after vaccination of H1N1 influenza. Vitreous opacity was obvious in both eyes. Broad exudative retinal detachment was observed in the right eye. Case 2 was a 47-year-old female who suffered from an acute high fever at 2 days after the vaccination of H1 N1 influenza. Later, she encountered bilateral headache and decreasing vision. In both eyes, mutton fat keratic precipitates, positive Tyndall phenomenon, congestion of optic disc and exudative retinal detachment were observed.

Clinical Predictors for Diagnosing Pandemic (H1N1) 2009 and Seasonal Influenza (H3N2) in Fever Clinics in Beijing,China

Objective Symptomatic predictors of influenza could assess risks and improve decisions about isolation and outpatient treatment. To develop such predictors, we undertook a prospective analysis of pandemic （HIN1） 2009 and seasonal influenza （H3N2） in patients attending fever clinics. Methods From 1 May 2009 to 1 January 2010, all adult patients admitted to fever clinics for suspected influenza, confirmed by real time RT-PCR, were enrolled. Predictors of influenza virus infection were selected with logistic regression models. Measures of sensitivity, specificity, positive and negative likelihood ratios （LRs） were calculated to identify the best predictors. Results The clinical features and routine blood test results of influenza （H1N1） 2009 and seasonal influenza were similar. The positive and negative LRs of current US CDC influenza-like illness （ILl） criteria were modest in predicting influenza infection. Our modified clinic predictors improved the ability of the positive and negative LRs to recognize pandemic （HIN1） 2009 and seasonal influenza. The revised criteria are： fever ~ 38 ~C accompanied by at least one of the following--cough, arthralgia or relative iymphopenia. Conclusion Patients with symptoms and signs that meet the new criteria are likely to have influenza and timely antiviral therapy may be appropriate. In addition, physicians should ascertain if influenza is circulating within the community or if there is a contact history of influenza and combine this information with the newly developed criteria to clinically diagnose influenza.

Localization Analysis of Heterophilic Antigen Epitopes of H1N1 Influenza Virus Hemagglutinin

Previous studies have indicated that two monoclonal antibodies(mAbs;A1-10 and H1-84)of the hemagglutinin(HA)antigen on the H1N1 influenza virus cross-react with human brain tissue.It has been proposed that there are heterophilic epitopes between the HA protein and human brain tissue(Guo et al.in Immunobiology 220:941-946,2015).However,characterisation of the two mAbs recognising the heterophilic epitope on HA has not yet been performed.In the present study,the common antigens of influenza virus HA were confirmed using indirect enzyme-linked immunosorbent assays and analysed with DNAMAN software.The epitopes were localized to nine peptides in the influenza virus HA sequence and the distribution of the peptides in the three-dimensional structure of HA was determined using PyMOL software.Key amino acids and variable sequences of the antibodies were identified using abYsis software.The results demonstrated that there were a number of common antigens among the five influenza viruses studied that were recognised by the mAbs.One of the peptides,P2(LVLWGIHHP191-199),bound both of the mAbs and was located in the head region of HA.The key amino acids of this epitope and the variable regions in the heavy and light chain sequences of the mAbs that recognised the epitope are described.A heterophilic epitope on H1N1 influenza virus HA was also introduced.The existence of this epitope provides a novel perspective for the occurrence of nervous system diseases that could be caused by influenza virus infection,which might aid in influenza prevention and control.