Human adenovirus (HAdV) type 14 had been infrequently associated with outbreaks of febrile respiratory illness (FRI) until the HAdV-14p1 emerged in 2006 and rapidly spread in the United States. Here, we report an outbreak of FRI caused by HadV-14p1 that occurred in 2011 at a primary and middle school in China.

Laboratory-confirmed deaths grossly underestimate influenza mortality burden, so that reliable burden estimates are derived from indirect statistical studies, which are scarce in low- and middle-income settings.

The present report compares prognosis in hospitalized cases with the H1N1 pandemic virus in two seasons.

Reassortment of influenza A viruses can give rise to viral ribonucleoproteins (vRNPs) with elevated polymerase activity and the previous three pandemic influenza viruses contained reassorted vRNPs of different origins. These suggest that reassorted vRNP may be one of the factors leading to a pandemic virus. In this study, we reconstituted chimeric vRNPs with three different viral strains isolated from avian, human and swine hosts. We applied a statistical strategy to identify the effect that the origin of a single vRNP protein subunit or the interactions between these subunits on polymerase activity.

Neuraminidase (NA) inhibitors (NAIs) are currently the only antivirals effective against influenza infections due to widespread resistance to M2 inhibitors.

Non-replicating respiratory syncytial virus (RSV) vaccine candidates could potentially prime for enhanced respiratory disease (ERD) due to a T-cell-mediated immunopathology, following RSV infection. Vaccines with built-in immune response modifiers, such as Toll-like receptor (TLR) ligands, may avoid such aberrant imprinting of the immune system.

Effectiveness of pandemic plans and community compliance was extensively researched following the H1N1 pandemic. This systematic review examined community response studies to determine whether behavioural responses to the pandemic were related to level of knowledge about the pandemic, perceived severity of the pandemic and level of concern about the pandemic.

To evaluate the safety of CSL's split-virion inactivated trivalent 2009 Southern Hemisphere formulation influenza vaccine (TIV) in children.

We investigated the roles of Toll-like receptors (TLRs) in naturally occurring influenza.

Seasonal influenza activity varies with geography and time of year.

H3N2 influenza viruses circulating in humans and European pigs originate from the pandemic A/Hong Kong/68 virus. Because of slower antigenic drift in swine, the antigenic divergence between swine and human viruses has been increasing. It remains unknown to what extent this results in a reduced cross-protection between recent human and swine H3N2 influenza viruses.

Characterization of the human respiratory syncytial virus (HRSV) season at the local level has important implications for appropriate decisions on the time period for administration of specific prophylaxis.

Influenza vaccines are effective in protecting against illness and death caused by this seasonal pathogen. The potency of influenza vaccines is measured by single radial immunodiffusion (SRID) assay that quantifies antigenic forms of hemagglutinin (HA). Hydrostatic pressure results in loss of binding of influenza virus to red blood cells, but it is not known whether this infers loss of potency.

During summer 2009, a US Navy ship experienced an influenza-like illness outbreak with 126 laboratory-confirmed cases of pandemic influenza A (H1N1) 2009 virus among the approximately 2000-person crew.

Laboratory testing results are often used to monitor influenza illness in populations, but results may not be representative of illness burden and distribution, especially in populations that are geographically, socioeconomically, and racially/ethnically diverse.

Epidemiological and clinical data of human coronaviruses (HCoVs) infections are restricted to span 1–3 years at most. We conducted a comprehensive 9-year study on HCoVs by analyzing 1137 respiratory samples from four subsets of patients (asymptomatic, general community, with comorbidities, and hospitalized) in São Paulo, Brazil.

Influenza C virus can cause both upper and lower respiratory tract infections and has been reported to be prevalent in children. However, these infections have been under-diagnosed, and epidemiological data available are limited due to the lack of convenient detection assays.

Delays in the release of national vital statistics hinder timely assessment of influenza severity, especially during pandemics. Inpatient mortality records could provide timelier estimates of influenza-associated mortality.

Denmark experienced three waves of the new pandemic influenza A (H1N1)pdm09 from July 2009 to February 2011. The aim of the study was to describe the epidemiology and clinical characteristics of hospitalized patients in a defined population of North Denmark Region with a mixed urban and rural community of 579 000 inhabitants.

To experimentally determine the survival kinetics of influenza virus on personal protective equipment (PPE) and to evaluate the risk of virus transfer from PPE, it is important to compare the effects on virus recovery of the method used to contaminate the PPE with virus and the type of eluent used to recover it.

Interferon alpha (IFNα) is a known antiviral agent. A double-blind, placebo-controlled clinical trial was conducted investigating the use of low-dose oral interferon alpha for preventing acute viral respiratory illnesses.

Acute lung injury is an important cause of death in humans infected with H5N1. It has been found that oxygen free radicals (OFRs) are elevated in lung tissue during influenza virus infections. In this study, we used a mouse model to explore the role of OFRs in acute lung injury caused by H5N1 viral infection.

The global impact of the 2009 influenza A(H1N1) pandemic (H1N1pdm) is not well understood.

Equine influenza virus (EIV) epizootics affect 2·1 million Mongolian horses approximately every 10 years and critically impact economy and nomadic livelihood of Mongolia.

Influenza A viruses circulating in pigs in Brazil are still not characterized, and only limited data are available about swine influenza epidemiology in the country. Therefore, we characterized the hemagglutinin (HA) and neuraminidase (NA) genes of influenza viruses isolated from Brazilian pigs. We also evaluated one case of probable swine-to-human transmission.

The duration of viral shedding is an important determinant of infectivity and transmissibility and provides vital information for effective infection prevention and control. However, few studies have evaluated viral shedding in patients admitted to hospital with 2009 H1N1 influenza and treated with oseltamivir.

During an evolving public health crisis, news organizations disseminate information rapidly, much of which is uncertain, dynamic, and difficult to verify. We examine factors related to international news coverage of H1N1 during the first month after the outbreak in late April 2009 and consider the news media's role as an information source during an emerging pandemic.

Influenza virus A(H1N1)pdm09 first appeared in Israel in late April 2009, disappeared in mid-March 2010, and reappeared in late October 2010. Symptoms were mostly mild without need for medical care.

Background  Influenza-associated lower respiratory tract hemorrhage (LRTH) has been reported in previous pandemics and is a rare complication of seasonal influenza virus infection. We describe patients with LRTH associated with 2009 pandemic influenza A (H1N1) (pH1N1) virus infection identified from April 2009 to April 2010 in the United States.

Methods  We ascertained patients with pH1N1-associated LRTH through state and local surveillance, the Emerging Infections Program, and CDCs Infectious Diseases Pathology Branch. All patients had influenza A, evidence of pneumonia, and evidence of LRTH.

Results  We identified 44 cases; the median number of days from illness onset to clinical signs of LRTH was one. Hemoptysis or respiratory tract bleeding was documented in 40% of pH1N1-associated LRTH cases, often present early during the course of illness. Twenty-one (48%) patients with LRTH had no other hemorrhagic diatheses. Seven (23%) patients with LRTH received antiviral treatment within two days of illness onset.

Conclusions  During influenza season, clinicians should consider influenza infection in the differential diagnosis for patients presenting with hemoptysis or other signs or symptoms of LRTH. While the impact of timing of antiviral therapy on this complication has not been studied, the rapid progression of LRTH may support use of early empiric therapy. Continued investigation is necessary to betterdefine the clinical spectrum of both seasonal influenza- and pH1N1-associated LRTH.

Human Rhinovirus (HRV) classification is an evolving process. New genotypes have been described within HRV-A and HRV-C species, but only one has been accepted related to HRV-B. From 2003 to 2010, a total of 3987 nasopharyngeal aspirate samples were taken from pediatric patients admitted to the Severo Ochoa Hospital in Madrid (Spain). After viral analysis, 949 (23·8%) tested positive to HRV. A random selection of 397 (42%) positive samples showed that 39 (9·8%) were HRV-B. The sequencing of partial VP4/VP2 coding region revealed the spread of 13 of 25 defined HRV-B serotypes and three putative new genotypes. Such results remark the high diversity of HRV-B.

Background  We estimated rates of influenza-associated deaths and hospitalizations in Argentina, a country that recommends annual influenza vaccination for persons at high risk of complications from influenza illness.

Methods  We identified hospitalized persons and deaths in persons diagnosed with pneumonia and influenza (P&I, ICD-10 codes J10-J18) and respiratory and circulatory illness (R&C, codes I00-I99 and J00-J99). We defined the influenza season as the months when the proportion of samples that tested positive for influenza exceeded the annual median. We used hospitalizations and deaths during the influenza off-season to estimate, using linear regression, the number of excess deaths that occurred during the influenza season. To explore whether excess mortality varied by sex and whether people were age <65 or ≥65 years, we used Poisson regression of the influenza-associated rates.

Results  During 2002–2009, 2411 P&I and 8527 R&C mean excess deaths occurred annually from May to October. If all of these excess deaths were associated with influenza, the influenza-associated mortality rate was 6/100 000 person-years (95% CI 4–8/100 000 person-years for P&I and 21/100 000 person-years (95% CI 12–31/100 000 person-years) for R&C. During 2005–2008, we identified an average of 7868 P&I excess hospitalizations and 22 994 R&C hospitalizations per year, resulting in an influenza-associated hospitalization rate of 2/10 000 person-years (95% CI 1–3/10 000 person-years) for P&I and 6/10 000 person-years (95% CI 3–8/10 000 person-years) for R&C.

Conclusion  Our findings suggest that annual rates of influenza-associated hospitalizations and death in Argentina were substantial and similar to neighboring Brazil.

In the present study, we analyzed the presence of antibodies to four different influenza viruses (pH1N1, hH1N1, swH1N1, and swH3N2) in the sera of 2094 backyard pigs from Mexico City. The sera were obtained between 2000 and 2009.

Objective  Influenza causes severe morbidity and mortality. This systematic review aimed to assess the incidence, etiology, and resource usage for influenza in Latin America and the Caribbean.

Design  Meta-analytic systematic review. Arcsine transformations and DerSimonian Laird random effects model were used for meta-analyses.

Setting  A literature search from 1980 to 2008 in MEDLINE, Cochrane Library, EMBASE, LILACS, Ministries of Health, PAHO, proceedings, reference lists, and consulting experts.

Sample  We identified 1092 references, of which 31 were finally included, in addition to influenza surveillance reports. We also used information from the 10 reports from the collaborative group for epidemiological surveillance of influenza and other respiratory virus (GROG), and information retrieved from the WHO global flu database FLUNET.

Main outcome measures  Incidence, percentage of influenza specimens out of the total received by influenza centers and resource-use outcomes.

Results  A total of 483 130 specimens of patients with influenza were analyzed. Meta-analysis showed an annual rate of 36 080 (95%CI 28 550 43 610) influenza-like illness per 100 000 persons-years. The percentage of influenza out of total specimens received by influenza centers ranged between 4.66% and 15.42%, with type A the most prevalent, and A subtype H3 predominating. The mean length of stay at hospital due to influenza ranged between 5.8 12.9 days, total workdays lost due to influenza-like illnesses were 17 150 days, and the mean direct cost of hospitalization was US$575 per laboratory-confirmed influenza case.

Conclusions  Our data show that seasonal influenza imposes a high morbidity and economic burden to the region. However, the vaccine-uptake rate has been low in this region. Population-based cohort studies are required to improve the knowledge about incidence and resource utilization, which would inform healthcare authorities for decision making.

From June 2009 through December 2009, Haiti conducted sentinel surveillance for influenza. 499 samples were collected and tested using real-time RT-PCR. 197 (39.5%) were positive for influenza, including 95 (48%) pandemic (H1N1) 2009, 57 (29%) seasonal influenza A and 45 (23%) influenza B. The median age of pandemic (H1N1) 2009 cases was 21.7; two-thirds of pandemic (H1N1) 2009 cases were in patients aged 6 years – 35 years. Pandemic activity peaked in September and co-circulated with other influenza subtypes. The age distribution and seasonality of pandemic (H1N1) 2009 in Haiti were similar to other countries in the Caribbean region.

Background:  Pediatric oncology patients represent a cohort of individuals uniquely at risk of complications from influenza, yet less likely to respond to the vaccine. It is not yet clear how to best protect this vulnerable population.

Methods:  We performed a prospective analysis of 177 pediatric oncology patients to define the predictors of influenza vaccine responses. Each variable was examined over three time points and a repeated measure analysis was performed.

Results:  Patients with ALL vaccinated during induction phase had superior influenza vaccine responses than those subjects vaccinated during post-induction or maintenance phases (P = 0·0237). Higher aggregate HAI titer responses were associated with a higher baseline B-cell count (P = 0·0240), and higher CD4 and CD8 influenza-specific T-cell responses, suggesting prior antigen exposure is a significant contributor. The solid tumor cohort had equivalent responses during all time frames of chemotherapy.

Discussion:  The optimal protection from influenza of pediatric patients on chemotherapy should include vaccination, but it is clear that not all patients produce high titers of antibodies after vaccination. This study identified biomarkers that could be used to individualize vaccine approaches. Immunologic predictors might have a role in targeting resources, as B-cell counts predicted of vaccine responses among the patients with ALL.

Coronaviruses have been demonstrated to contribute substantially to respiratory tract infections among the child population. Though infected children commonly present mild upper airway symptoms, in high-risk patients with underlying conditions, particularly in immunocompromised children these pathogens may lead to severe lung infection and extrapulmonary disorders. In this paper, we provide the first report of the case of a 15-month-old child with severe combined immunodeficiency and coronavirus HKU1-related pneumonia with fatal respiratory distress syndrome.

Background & objective  During the temperate out-of-season months in Australia in late 2010 and early 2011, an unprecedented high number of influenza notifications were recorded. We aimed to assess the significance of these notifications.

Methods  For Australia, we used laboratory-confirmed cases notified to the WHO FluNet surveillance tool; the percentage of these that were positive; notifications by state and influenza type and subtype; and surveillance data from Google FluTrends. For the state of Victoria, we used laboratory-confirmed notified cases and influenza-like illness (ILI) proportions. We compared virus characterisation using haemagglutination-inhibition assays and phylogenetic analysis of the haemagglutinin gene for seasonal and out-of-season notifications.

Results  The increase in notifications was most marked in tropical and subtropical Australia, but the number of out-of-season notifications in temperate Victoria was more than five times higher than the average of the previous three seasons. However, ILI proportions in spring-summer were not different to previous years. All out-of-season viruses tested were antigenically and genetically similar to those tested during either the 2010 or 2011 influenza seasons. An increase in the number of laboratories testing for influenza has led to an increase in the number of tests performed and cases notified.

Conclusion  An increase in influenza infections in spring-summer of 2010–11 in tropical and temperate Australia was not associated with any differences in virus characterisation compared with viruses that circulated in the preceding and following winters. This increase probably reflected a natural variation in out-of-season virus circulation, which was amplified by increased laboratory testing.

Background  School closures were widely implemented in Argentina during the 2009 H1N1 influenza virus pandemic.

Objectives  To assess the economic impact of school closures on households, their effectiveness in preventing children from engaging in social group activities, and parental attitudes toward them.

Methods  Three schools that closed for 2 weeks in response to the pandemic were identified in two socioeconomically distinct cities in Argentina. All households with children enrolled in these schools were surveyed. Direct and indirect costs attributable to closures were estimated from the household perspective. Other information collected included children activities during the closures and parental attitudes toward the intervention.

Results  Completed questionnaires were returned by 45% of surveyed households. Direct and indirect costs due to closures represented 11% of imputed monthly household income in the city with lower socioeconomic status, and 3% in the other city (P = 0·01). Non-childcare expenses and loss of workdays were more common in the city with lower socioeconomic status. Childcare expenses were less common and were experienced by a similar percentage of households in both cities. About three-quarters of respondents in both cities agreed with the closures. The main concern among those who disagreed with closures was their negative impact on education. Children in more than two-thirds of affected households left their home at least once during the closures to spend time in public places.

Conclusion  School closures may more significantly impact low-income households. Authorities should consider the range of economic impacts of school closures among families when planning their implementation.

Background  Highly pathogenic avian influenza A/H5N1 virus remains a potential pandemic threat, and it is essential to continue vaccine development against this subtype. A local mucosal immune response in the upper respiratory tract may stop influenza transmission. It is therefore important to develop effective intranasal pandemic influenza vaccines that induce mucosal immunity at the site of viral entry.

Objectives  We evaluated the humoral and cellular immune responses of two promising mucosal adjuvants (Chitosan and c-di-GMP) for intranasal influenza H5N1 vaccine in a murine model. Furthermore, we evaluated the concept of co-adjuvanting an experimental adjuvant (c-di-GMP) with chitosan.

Methods  BALB/c mice were intranasally immunised with two doses of subunit NIBRG-14 (H5N1) vaccine (7·5, 1·5 or 0·3 μg haemagglutinin (HA) adjuvanted with chitosan (CSN), c-di-GMP or both adjuvants.

Results  All adjuvant formulations improved the serum and local antibody responses, with the highest responses observed in the 7·5 μg HA CSN and c-di-GMP-adjuvanted groups. The c-di-GMP provided dose sparing with protective single radial haemolysis (SRH), and haemagglutination inhibition (HI) antibody responses found in the 0·3 μg HA group. CSN elicited a Th2 response, whereas c-di-GMP induced higher frequencies of virus-specific CD4⁺ T cells producing one or more Th1 cytokines (IFN-γ⁺, IL-2⁺, TNF-α⁺). A combination of the two adjuvants demonstrated effectiveness at 7·5 μg HA and triggered a more balanced Th cytokine profile.

Conclusion  These data show that combining adjuvants can modulate the Th response and in combination with ongoing studies of adjuvanted intranasal vaccines will dictate the way forward for optimal mucosal influenza vaccines.

The 2009 pandemic served as a strong reminder that influenza-induced disease can have a great impact on certain at-risk populations and that pregnant women are one such important population. The increased risk of fatal and severe disease in these women was appreciated more than 500 years ago, and during the last century, pregnant women and their newborns have continued to be greatly affected by both seasonal and pandemic influenza. In this review, we briefly discuss the data collected both before and after the 2009 pandemic as it relates to the impact of influenza on pregnant women and their fetuses/newborns, as well as risk variables, clinical features, clues to pathophysiologic mechanisms, and approaches to treatment and prevention.

Purpose  To examine whether apparent advantages following training in meditation over exercise can be attributed to specific symptoms, functional impairments, or quality-of-life indicators assessed by the Wisconsin Upper Respiratory Symptom Survey (WURSS-24).

Methods  Results from the randomized controlled trial “Meditation or Exercise for Preventing Acute Respiratory Illness” showed mean global severity and total days of illness were worse in control (358, 8·9) compared with exercise (248, 5·1) or meditation (144, 5·0). Global severity of illness was estimated using area under the curve from daily self-reported severity scores on the WURSS-24. For this project, we estimated within-group WURSS item-level severity and between-group effect sizes (Cohen’s “d” statistic) relative to control. The item-level effect sizes were grouped into (i) symptom and (ii) function and quality of life domains.

Results  Among the three groups, mediators showed the lowest severity estimates for 21 of 22 WURSS items. Item-level Cohen’s “d” indicated most benefit was evident in WURSS items representing function and quality of life. Compared with exercise, meditation fostered larger reductions in illness severity, although due mostly to improved function and the quality of life domain (d = −0·33, P < 0·001) compared with symptom domain (d = −0·22, P < 0·001).

Conclusions  The apparent advantage of training in meditation over exercise for reducing cold and flu illness is explained more by improved function and quality of life than by a reduction in symptom severity.

Background  Outbreaks of influenza-like illness (ILI) are common in long-term care facilities (LTCFs) and result in significant morbidity and mortality among residents.

Objectives  We describe patterns of reported ILI outbreaks in LTCFs in Winnipeg, Canada, and examine LTCF and outbreak characteristics that influence the clinical outcomes of these outbreaks.

Methods  We analyzed the electronic records of all ILI outbreaks reported by LTCFs in Winnipeg from 2003 to 2011. Outbreak duration, ILI attack rates among staff and residents, and residents’ death rates were calculated by presumed viral etiology, staff vaccination rates, type of influenza chemoprophylaxis used, and time to notification to public health.

Results  Of a total of 154 reported outbreaks, most (N = 80) were attributed to influenza, and these outbreaks tended to have higher attack and death rates among LTCF residents compared with outbreaks caused by other respiratory viruses (12) or those of unknown etiology (62). About 92% of residents and 38% of staff of the average LTCFs were vaccinated. Chemoprophylaxis was used in 57·5% of influenza outbreaks. Regardless of presumed viral etiology, outbreaks reported within 3 days of onset ended sooner and had lower attack and mortality rates among residents.

Conclusions  Influenza-like illness outbreaks still occur among highly immunized LTCF residents, so in addition to vaccination of staff and residents, it is important to maintain competent infection control practices. Early identification and notification to public health authorities and possibly early initiation of control measures could improve clinical outcomes of ILI outbreaks.

Recent research involving lab-modified H5N1 influenza viruses with increased transmissibility and the ongoing evolution of the virus in nature should remind us of the continuing importance of preparedness for a severe influenza pandemic. Current vaccine technology and antiviral supply remain inadequate, and in a severe pandemic, most low-resource communities will fail to receive adequate medical supplies. However, with suitable guidance, these communities can take appropriate actions without substantial outside resources to reduce influenza transmission and care for the ill. Such guidance should be completed, and support provided to developing countries to adapt it for their settings and prepare for implementation.

Background  Highly pathogenic avian influenza (H5N1) virus continues to cause infections in Egypt. This study describes the practices associated with raising and slaughtering household poultry to identify risk factors for H5N1 infection and reasons for non-compliance with preventive measures.

Methods  An investigation was conducted of 56 households with household flocks (19 households with human H5N1 cases, 19 with poultry H5N1 cases, and 18 with no reported poultry or human H5N1 cases). Data were collected via structured observations and in-depth interviews.

Results  Half of the households kept at least some free-range poultry and mixed at least some different species of poultry as it was considered beneficial for the poultry. Feeding and cleaning practices exposed children to contact with poultry; slaughtering contaminated homes; use of personal protective barriers was not a norm; waste management exposed the communities to slaughtering waste and dead chickens; and reporting of sick and dead poultry was not a practice. Only minor changes in poultry-handling took place following H5N1 virus outbreaks.

Discussion  H5N1 virus prevention in Egypt represents both an epidemiological and socio-cultural challenge. Traditional poultry-rearing practices that likely increase exposures to H5N1-infected poultry are common throughout Egypt. Despite education campaigns following sporadic H5N1 outbreaks, no differences in these practices could be detected between households with previous H5N1 human or poultry cases and those households with any previous experience with H5N1. Development of H5N1 infection–related education campaign strategies should focus on perceptions underlying traditional practices in order to tailor public awareness messages that are meaningful for communities.

Background  Vietnam is currently developing domestic capability to manufacture influenza vaccines but information on the genetic and antigenic characteristics of locally circulating seasonal influenza viruses is limited. To assess the relevance of WHO recommended vaccine strains to the situation in Vietnam, we analyzed the genetic relatedness of the hemagglutinin (HA) gene of seasonal influenza A viruses circulating in Vietnam from 2001 to 2009 to WHO recommended vaccine strains over the same period.

Methods and Principal findings  We sequenced the HA gene of 32 H1N1 and 44 H3N2 seasonal influenza A isolates from laboratory-based sentinel surveillance sites in Hanoi from 2001 to 2005 and from a national influenza surveillance system from 2005 to 2009. H1 and H3 HA phylogenetic trees rooted to vaccine strains A/Beijing/295/1995 (H1N1) and A/Moscow/10/1999 (H3N2), respectively, were constructed with contemporary HA sequences of isolates from neighboring countries. We found some genetic differences between seasonal influenza H3N2 viruses and three WHO influenza vaccine strains recommended for use in the Northern and Southern Hemispheres for the 2001–2004 and 2007–2008 seasons and close genetic identity of circulating H3N2 strains with the recommended WHO Southern Hemisphere vaccine strains for 2004 and 2009 seasons. The genetic similarity of circulating H1N1 strains with the WHO recommended vaccine strains are described for the study period 2001–2009.

Conclusions  The HA gene of seasonal influenza virus strains in Vietnam (especially influenza A/H3N2) showed varying degrees of genetic identity compared with those of the Northern or Southern Hemisphere vaccine strains recommended by WHO. The close relatedness of the HA of Vietnamese strains and contemporary strains from nearby countries indicate a good genetic match of circulating strains during study period. Greater representation of virus isolates from South East Asia in the vaccine strain selection process is desirable of influenza vaccine development in Vietnam.

The trends of influenza infection in Suriname were assessed from February 2010 through February 2011. Testing of 393 patients with symptoms of acute respiratory infection (ARI) revealed 15·3% Influenza B and 18·6% could be identified as influenza A positive, consisting of 56% influenza A(H1N1)pdm09 and 44% seasonal A(H3N2). Influenza infection occurred throughout the year, and all three influenza types affected young children as the primary population. The annual incidence of A(H1N1)pdm09 was 6·88 per 100 000 inhabitants [CI] 4·87–9·45. The spread of influenza could neither be linked to tourist flow from the Netherlands nor to contact rates related to school schedules.

Background  Few drugs are currently licensed to treat influenza A infection, and new therapies are needed, especially for highly pathogenic strains. Traditional medicinal plants, such as Lycoris radiata, are a potential source of new antiviral agents.

Objective  To test 15 Amaryllidaceae alkaloids isolated from the bulbs of L. radiata in vitro for antiviral activities against influenza virus type A, A/Chicken/GuangDong/178/2004 (H5N1, 178).

Methods  Antiviral activities of the compounds were tested in time-of-addition assays, hemagglutination inhibition (HI) assays, neuraminidase (NA) activity assays, and viral entry inhibition assays using H5N1-HIV pseudoviruses. Effects of the compounds on localization and activity of the viral ribonucleoprotein (RNP) were determined by immunofluorescence and an RNP minigenome assay, respectively.

Results  Among the alkaloids, lycorine (AA1), hippeastrine (AA2), hemanthamine (AA3) and 11-hydroxy vittatine (AA4) exhibited antiviral activities, with EC₉₀ values of 0·52, 82·07, 4·15, and 13·45 μm, respectively. These compounds did not affect the function of the outer membrane proteins or the viral entry process and viral RNP activity. As AA1 and AA3 exhibited stronger antiviral activities, they were further analyzed. Intracellular nucleoprotein (NP) localization showed that AA1 and AA3 inhibited the RNP complex in the nucleus at an early stage of a single-round and multi-round of replication.

Conclusion  Four Amaryllidaceae alkaloids were first determined that could exert anti-influenza activities after virus entry into cells. Furthermore, AA1 and AA3 could inhibit nuclear-to-cytoplasmic export of the RNP complex of virus replication. Thus, these compounds may be developed further as anti-influenza drug candidates.

Background  Findings from studies examining the association between obesity and acute respiratory infection are inconsistent. Few studies have assessed the relationship between obesity-related behavioral factors, such as diet and exercise, and risk of acute respiratory infection.

Objective  To determine whether community prevalence of obesity, low fruit/vegetable consumption, and physical inactivity are associated with influenza-related hospitalization rates.

Methods  Using data from 274 US counties, from 2002 to 2008, we regressed county influenza-related hospitalization rates on county prevalence of obesity (BMI ≥ 30), low fruit/vegetable consumption (<5 servings/day), and physical inactivity (<30 minutes/month recreational exercise), while adjusting for community-level confounders such as insurance coverage and the number of primary care physicians per 100 000 population.

Results  A 5% increase in obesity prevalence was associated with a 12% increase in influenza-related hospitalization rates [adjusted rate ratio (ARR) 1·12, 95% confidence interval (CI) 1·07, 1·17]. Similarly, a 5% increase in the prevalence of low fruit/vegetable consumption and physical inactivity was associated with an increase of 12% (ARR 1·12, 95% CI 1·08, 1·17) and 11% (ARR 1·11, 95% CI 1·07, 1·16), respectively. When all three variables were included in the same model, a 5% increase in prevalence of obesity, low fruit/vegetable consumption, and physical inactivity was associated with 6%, 8%, and 7% increases in influenza-related hospitalization rates, respectively.

Conclusions  Communities with a greater prevalence of obesity were more likely to have high influenza-related hospitalization rates. Similarly, less physically active populations, with lower fruit/vegetable consumption, tended to have higher influenza-related hospitalization rates, even after accounting for obesity.

Background  The constant-phase model (CPM) is commonly fit to respiratory system input impedance (Z_rs) to estimate lung mechanics. Driving signal frequencies and the method of model fitting may influence the results, especially in cases of severe lung disease or under severe bronchoconstriction.

Objective  To illustrate the effects of different CPM fits to Z_rs data using a mouse model of influenza-induced lung disease.

Methods  BALB/c mice infected with influenza (or control) were challenged with methacholine. The CPM was fitted to Z_rs, measured between 0·25 and 19·625 Hz, using both unweighted and weighted fits. The effect of different lowest frequencies was assessed.

Results and Conclusions  For influenza-infected mice, the unweighted fit was poor, and airway resistance (R_aw) was often biologically impossible. The weighted fit provided more realistic estimates of R_aw. Different model fits and minimal frequencies had little effect on tissue mechanics.

Objectives  The goal of this double-blind, randomized, controlled clinical trial was to assess the safety and immunogenicity of two different doses of a monovalent split-virion 2009 pandemic influenza A/H1N1 vaccine without adjuvant in Chinese infants aged 6-35 months.

Design and setting  Subjects were randomly assigned to receive either a 2009 pandemic (H1N1) vaccine containing 7.5 or 15 μg haemagglutinin (HA) or a seasonal influenza vaccine. 2 doses of the H1N1 vaccines or the seasonal influenza vaccine were given 21 days apart in younger infants aged 6-23 months or older infants aged 24-35 months.

Sample  Serum samples were collected immediately before the first injection and before and 21 days after the second injection.

Main outcome measures  Primary outcomes were haemagglutinin inhibition (HI) antibody responses 21 days following each vaccination. Safety was monitoring throughout the study.

Results  The first vaccination of 7.5 μg and 15 μg H1N1 vaccine induced seroprotective antibody titers (HI titers ≥ 1: 40) in 42.9-57.4% of younger infants and 49.1-61.0% older infants. Immune responses after completion of the two dose schedule were comparable in both age groups with seroprotective rates of 91-98% in each vaccine and age group and GMTs of 173-263. The H1N1 vaccine elicited similar rates of local and systemic adverse reactions as the seasonal influenza vaccine.

Conclusions  The 2009 pandemic influenza A /H1N1 vaccine were highly immunogenic in infants aged 6-35 months, and displayed a safety and reactogenicity profile similar to the seasonal influenza vaccine.

Trial registration  ClinicalTrial.gov identifier: NCT01047202

Background  The clinical presentation of influenza in infancy may be similar to serious bacterial infection and be investigated with invasive procedures like lumbar puncture (LP), despite very limited evidence that influenza occurs concomitantly with bacterial meningitis, perhaps because the diagnosis of influenza is very often not established when the decision to perform LP is being considered.

Methods  A retrospective medical record review was undertaken in all children presenting to the Children’s Hospital at Westmead, Sydney, Australia, in one winter season with laboratory-confirmed influenza or other respiratory virus infections (ORVIs) but excluding respiratory syncytial virus, to compare the use of, and reflect on the need for, the performance of invasive diagnostic procedures, principally LP, but also blood culture, in influenza and non-influenza cases. We also determined the rate of concomitant bacterial meningitis or bacteraemia.

Findings  Of 294 children, 51% had laboratory-confirmed influenza and 49% had ORVIs such as parainfluenza viruses (34%) and adenoviruses (15%). Of those with influenza, 18% had a LP and 71% had a blood culture performed compared with 6·3% and 55·5% in the ORVI group (for both P < 0·01). In multivariate analysis, diagnosis of influenza was a strong independent predictor of both LP (P = 0·02) and blood culture (P = 0·05) being performed, and, in comparison with ORVIs, influenza cases were almost three times more likely to have a LP performed on presentation to hospital. One child with influenza (0·9%) had bacteraemia and none had meningitis.

Interpretation  Children with influenza were more likely to undergo LP on presentation to hospital compared with those presenting with ORVIs. If influenza is confirmed on admission by near-patient testing, clinicians may be reassured and less inclined to perform LP, although if meningitis is clinically suspected, the clinician should act accordingly. We found that the risk of bacterial meningitis and bacteraemia was very low in hospitalised children with influenza and ORVIs. A systematic review should be performed to investigate this across a large number of settings.

Abstract Background  Adjuvant formulations are critical components of modern vaccines based on recombinant proteins, which are often poorly immunogenic without additional immune stimulants. Oil-in-water emulsions comprise an advanced class of vaccine adjuvants that are components of approved seasonal and pandemic influenza vaccines. However, few reports have been published that systematically evaluate the in vitro stability and in vivo adjuvant effects of different emulsion components.

Objectives  To evaluate distinct classes of surfactants, oils, and excipients, for their effects on emulsion particle size stability, antigen structural interactions, and in vivo activity when formulated with a recombinant H5N1 antigen.

Methods  Emulsions were manufactured by high pressure homogenization and characterized alone or in the presence of vaccine antigen by dynamic light scattering, zeta potential, viscosity, pH, hemolytic activity, electron microscopy, fluorescence spectroscopy, and SDS-PAGE. In vivo vaccine activity in the murine model was characterized by measuring antibody titers, antibody-secreting plasma cells, hemagglutination inhibition titers, and cytokine production.

Results  We demonstrate that surfactant class and presence of additional excipients are not critical for biological activity, whereas oil structure is crucial. Moreover, we report that simplified two-component emulsions appear more stable by particle size than more complex formulations.Finally, differences in antigen structural interactions with the various emulsions do not appear to correlate with in vivo activity.

Conclusions  Oil-in-water emulsions can significantly enhance antibody and cellular immune responses to a pandemic influenza antigen. The dramatic differences in adjuvant activity between squalene-based emulsion and medium chain triglyceride-based emulsion are due principally to the biological activity of the oil composition rather than physical interactions of the antigen with the emulsion.

The prevalence and timing of emergence of oseltamivir-resistant seasonal and pandemic influenza A (H1N1) viruses in Myanmar in 2008 and 2009 are described in this report. In 2008, the oseltamivir-resistant seasonal H1N1 virus was detected at a lower rate (6%) and emerged at least 2 months later when compared with neighboring countries. Similarly, the prevalence of pandemic H1N1 virus was low (3%) and the timing of emergence was late (August 2009) in Myanmar. Interestingly, we detected three isolates that were resistant to both amantadine and oseltamivir. Limited movement of people into the country is attributed to the delayed emergence of drug-resistant seasonal and pandemic A(H1N1) viruses.

Background  The number of admissions to hospital for which influenza is laboratory confirmed is considered to be a substantial underestimate of the true number of admissions due to an influenza infection. During the 2009 pandemic, testing for influenza in hospitalized patients was a priority, but the ascertainment rate remains uncertain.

Methods  The discharge abstracts of persons admitted with any respiratory condition were extracted from the Canadian Discharge Abstract Database, for April 2003–March 2010. Stratified, weekly admissions were modeled as a function of viral activity, seasonality, and trend using Poisson regression models.

Results  An estimated 1 out of every 6·4 admissions attributable to seasonal influenza (2003–April 2009) were coded to J10 (influenza virus identified). During the 2009 pandemic (May–March 2010), the influenza virus was identified in 1 of 1·6 admissions (95% CI, 1·5–1·7) attributed to the pandemic strain. Compared with previous H1N1 seasons (2007/08, 2008/09), the influenza-attributed hospitalization rate for persons <65 years was approximately six times higher during the 2009 H1N1 pandemic, whereas for persons 75 years or older, the pandemic rate was approximately fivefold lower.

Conclusions  Case ascertainment was much improved during the pandemic period, with under ascertainment of admissions due to H1N1/2009 limited primarily to patients with a diagnosis of pneumonia.

Introduction  Recent literature suggests that dual or multiple virus infections may affect disease severity. However, few studies have investigated the effect of co-infection with influenza A viruses.

Objectives  To identify the association between influenza A and respiratory viruses co-infections with disease outcome.

Methodology  Data for samples from North West England tested between January 2007 and June 2011 was analysed for patterns of co-infection between influenza A viruses and eight respiratory viruses. Risk of hospitalisation to ICU or general ward in single versus co-infections was assessed using logistic regression.

Results  Of the 25 596 samples analysed for respiratory viruses 40·7% (10 501) were positive for any virus. Co-infections were detected in 4·7% (137/2879) of all patients with influenza A(H1N1)pdm09, and 7·3% (57/779) of those with other influenza A virus infections. Co-infection between seasonal influenza A viruses and influenza B virus was associated with a significant increase in the risk of admission to ICU/death (OR: 22·0, 95% CI: 2·21–219·8, P = 0·008). Respiratory syncytial virus/influenza A (RSV/Flu A) co-infection also increased this risk but was not statistically significant. For influenza A(H1N1)pdm09, RSV and AdV co-infection increased risk of hospitalisation to general ward whereas Flu B increased risk of admission to ICU, but none of these were statistically significant.

Conclusion  Co-infection is a significant predictor of disease outcome; combined treatment, introduction of an integrated vaccine for all respiratory viruses and development of multi-target rapid diagnostic tests is recommended. Integration of respiratory viruses’ co-infections into public health reports could also contribute to the accumulation of evidence.

Background  Patients with end-stage renal disease have a reduced response to vaccination because of the general suppression of the immune system associated with uraemia.

Objectives  We evaluated the immune response and differential factors in the immunogenecity to an adjuvanted A(H1N1) pdm09 vaccine (Pandemrix^®) in four populations of renal patients after one and two doses of vaccine.

Patients Methods  151 patients were included in this study: 58 chronic haemodialysis patients, 52 renal allograft recipients, 14 peritoneal dialysis patients and 27 patients with advanced chronic kidney disease in preparation for kidney replacement therapy. Influenza-specific antibody levels were measured by monitoring A(H1N1) pdm09 titres using a haemagglutination inhibition assay.

Results  The seroconversion rate at 42 days after two vaccine doses was 80% in the haemodialysis group, 64·9% in the renal allograft recipients group, 100% in the advanced chronic kidney disease group and 71·4% in the peritoneal dialysis group (P = 0·041).

Conclusions  Immune response to two doses of the influenza A H1N1 vaccine is dissimilar in the four renal conditions, confirming that seroprotection in pre-dialysis, haemodialysis and peritoneal dialysis is similar to that in the general population vaccinated with one dose. In contrast, renal transplant recipients with good allograft function showed inadequate protection and triple immunosuppressive therapy including calcineurin inhibitors, mycophenolate and steroids negatively influenced seroconversion after vaccination in renal recipients.

Objectives:  To determined the pathogen-specific incidence of respiratory virus infection in Hutterite communities occurring over the 2008–2009 influenza season and assess temporal characteristics of respiratory illness related to infection.

Methods:  3273 participants community members enrolled in a cluster randomized trial of influenza vaccine were studied.

Results:  One hundred forty-nine participants had laboratory-confirmed influenza, and 595 had at least one episode of laboratory-confirmed respiratory viral infection other than influenza. Entero/rhinovirus had the highest incidence among children <5 years.

Conclusions:  A decline in the incidence of infections with age was observed for influenza as well as for most other respiratory viruses.

Background  Influenza vaccines are licensed annually based on immunogenicity studies. We used five sequential years of data to estimate influenza vaccine effectiveness (VE), the critical outcome in the field.

Methods  Between 2007 and 2011, we performed annual prospective test-negative design case–control studies among adults aged 20–64 years recruited from sentinel general practices in the Australian state of Victoria. We used PCR-confirmed influenza as the endpoint to estimate influenza VE for all years. We compared annual VE estimates with the match between circulating and vaccine strains, determined by haemagglutination inhibition assays.

Results  The adjusted VE estimate for all years (excluding 2009) was 62% (95% CI 43, 75). By type and subtype, the point estimates of VE by year ranged between 31% for seasonal influenza A(H1N1) and 88% for influenza A(H1N1)pdm09. In 2007, when circulating strains were assessed as incompletely matched, the point estimate of the adjusted VE against all influenza was 58%. The point estimate was 59% in 2011 when all strains were assessed as well matched.

Conclusion  Trivalent inactivated vaccines provided moderate protection against laboratory-confirmed influenza in adults of working age, although VE estimates were sensitive to the model used. VE estimates correlated poorly with circulating strain match, as assessed by haemagglutination inhibition assays, suggesting a need for VE studies that incorporate antigenic characterization data.

Background  Influenza B strains from two distinct lineages (Yamagata and Victoria) have cocirculated over recent years. Current seasonal vaccines contain a single B lineage resulting in frequent mismatches between the vaccine strain and the circulating strain. An Ann Arbor strain quadrivalent live attenuated influenza vaccine (Q/LAIV) containing B strains from both lineages is being developed to address this issue.

Objectives  The goal of this study was to evaluate whether Q/LAIV administered intranasally as a single dose to a single nostril, using a blow-fill-seal (BFS) delivery system had a similar immunogenicity and safety profile compared with the licensed trivalent vaccine delivered using the Accuspray device.

Patients/Methods  Adults aged 18–49 years were randomized to receive one intranasal dose of Q/LAIV delivered using a BFS device (Q/LAIV-BFS; n = 1202) or one of two trivalent live attenuated influenza vaccines (T/LAIV) containing one of the corresponding B strains (total T/LAIV, n = 598). Primary endpoints were the post-vaccination strain-specific serum hemagglutination inhibition antibody geometric mean titers for each strain. Secondary immunogenicity endpoints, safety, and acceptability of the BFS device were also assessed.

Results  Q/LAIV was immunogenically non-inferior to T/LAIV for all four influenza strains. Secondary immunogenicity outcomes were consistent with the primary endpoint. Solicited symptoms and AEs were comparable in both groups. Subjects considered the BFS device to be acceptable.

Conclusions  Immune responses to vaccination with Ann Arbor strain Q/LAIV-BFS were non-inferior to those with T/LAIV. Q/LAIV may confer broader protection against seasonal influenza B by targeting both major influenza B lineages.

Hand hygiene may be associated with modest protection against some acute respiratory tract infections, but its specific role in influenza transmission in different settings is unclear. We aimed to review evidence that improving hand hygiene reduces primary and secondary transmission of (i) influenza and (ii) acute respiratory tract infections in community settings. We searched Medline, Embase, Global Health and Cochrane databases up to 13 February 2012 for reports in any language of original research investigating the effect of hand hygiene on influenza or acute respiratory tract infection where aetiology was unspecified in community settings including institutions such as schools, and domestic residences. Data were presented and quality rated across outcomes according to the Grading of Recommendations Assessment, Development and Evaluation system. Sixteen articles met inclusion criteria. There was moderate to low-quality evidence of a reduction in both influenza and respiratory tract infection with hand hygiene interventions in schools, greatest in a lower–middle-income setting. There was high-quality evidence of a small reduction in respiratory infection in childcare settings. There was high-quality evidence for a large reduction in respiratory infection with a hand hygiene intervention in squatter settlements in a low-income setting. There was moderate- to high-quality evidence of no effect on secondary transmission of influenza in households that had already experienced an index case. While hand hygiene interventions have potential to reduce transmission of influenza and acute respiratory tract infections, their effectiveness varies depending on setting, context and compliance.

Despite the widespread availability and use of influenza vaccines, influenza still poses a considerable threat to public health. Vaccines against seasonal influenza do not offer protection against pandemic viruses, and vaccine efficacy against seasonal viruses is reduced in seasons when the vaccine composition is not a good match for the predominant circulating viruses. Vaccine efficacy is also reduced in older adults, who are one of the main target groups for vaccination. The continual threat of pandemic influenza, with the known potential for rapid spread around the world and high mortality rates, has prompted researchers to develop a number of novel approaches to providing immunity to this virus, focusing on target antigens which are highly conserved between different influenza A virus subtypes. Several of these have now been taken into clinical development, and this review discusses the progress that has been made, as well as considering the requirements for licensing these new vaccines and how they might be used in the future.

Background  Travellers’ compliance with measures to prevent influenza through the use of antivirals and influenza vaccine remains very poor despite influenza being one of the commonest travel and vaccine-preventable diseases. A study was undertaken to assess travellers’ beliefs, perceptions and intentions to take antivirals for the treatment and prevention of influenza during the H1N1 pandemic.

Methods  A cross-sectional survey (n = 96) of travellers who attended the Royal Free Travel Health Centre, London, UK was undertaken in September 2009. A self-administered questionnaire was completed by a traveller in advance of their pre-travel health consultation. Logistic regression identified variables independently associated with compliance.

Results  Influenza vaccination uptake for the 5 years preceding the study was found to be 20·8%. This was statistically significantly higher for older travellers and those with underlying health conditions (P < 0·005). Mean intention to comply with antiviral drugs on a preventive and therapeutic basis was 58% and 72%, respectively, and this varied markedly with age and with dispensed antimalarial chemoprophylaxis.

Conclusion  This study identifies some beliefs and perceptions travellers consider with regard to the therapeutic and preventive influenza use of antivirals during the H1N1 pandemic; it underscores the importance of travellers receiving hemisphere appropriate influenza vaccination. The external validity of these study findings requires further corroboration involving other travel clinics and different cohorts of travellers during seasonal activity or outbreaks of influenza. These findings could guide the development of future strategies for the prevention of influenza in travellers.

Background  Exposure to diesel exhaust particles (DEP) is thought to exacerbate many pre-existing respiratory diseases, including asthma, bronchitis and chronic obstructive pulmonary disease, however, there is a paucity of data on whether DEP exacerbates illness due to respiratory viral infection.

Objectives  To assess the physiological consequences of an acute DEP exposure during the peak of influenza-induced illness.

Methods  We exposed adult female BALB/c mice to 100 μg DEP (or control) 3·75 days after infection with 10^4·5 plaque forming units of influenza A/Mem71 (or control). Six hours, 24 hours and 7 days after DEP exposure we measured thoracic gas volume and lung function at functional residual capacity. Bronchoalveolar lavage fluid was taken for analyses of cellular inflammation and cytokines, and whole lungs were taken for measurement of viral titre.

Results  Influenza infection resulted in significantly increased inflammation, cytokine influx and impairment to lung function. DEP exposure alone resulted in less inflammation and cytokine influx, and no impairment to lung function. Mice infected with influenza and exposed to DEP had higher viral titres and neutrophilia compared with infected mice, yet they did not have more impaired lung mechanics than mice infected with influenza alone.

Conclusions  A single dose of DEP is not sufficient to physiologically exacerbate pre-existing respiratory disease caused by influenza infection in mice.

Background  The performance of rapid influenza diagnostic tests (RIDTs) that detect influenza viral nucleoprotein (NP) antigen has been reported to be variable. Recent human infections with variant influenza A viruses that are circulating in pigs prompted the investigation of the analytical reactivity of RIDTs with these variant viruses.

Objectives  To determine analytical reactivity of seven FDA-cleared RIDTs with influenza A variant viruses in comparison with the reactivity with recently circulating seasonal influenza A viruses.

Methods  Tenfold serial dilutions of cell culture–grown seasonal and variant influenza A viruses were prepared and tested in duplicate with seven RIDTs.

Results  All RIDTs evaluated in this study detected the seasonal influenza A(H3N2) virus, although detection limits varied among assays. All but one examined RIDT identified the influenza A(H1N1)pdm09 virus. However, only four of seven RIDTs detected all influenza A(H3N2)v, A(H1N2)v, and A(H1N1)v viruses. Reduced sensitivity of RIDTs to variant influenza viruses may be due to amino acid differences between the NP proteins of seasonal viruses and the NP proteins from viruses circulating in pigs.

Conclusions  Clinicians should be aware of the limitations of RIDTs to detect influenza A variant viruses. Specimens from patients with influenza-like illness in whom H3N2v is suspected should be sent to public health laboratories for additional diagnostic testing.

Background:  Estimating influenza incidence in outpatient settings is challenging. We used outpatient healthcare practice populations as a proxy to estimate community incidence of influenza-like illness (ILI) and laboratory-confirmed influenza-associated ILI.

Methods:  From October 2009 to July 2010, 38 outpatient practices in seven jurisdictions conducted surveillance for ILI (fever with cough or sore throat for patients ≥2 years; fever with ≥1 respiratory symptom for patients <2 years). From a sample of patients with ILI, respiratory specimens were tested for influenza.

Results:  During the week of peak influenza activity (October 24, 2009), 13% of outpatient visits were for ILI and influenza was detected in 72% of specimens. For the 10-month surveillance period, ILI and influenza-associated ILI incidence were 20·0 (95% CI: 19·7, 20·4) and 8·7/1000 (95% CI: 8·2, 9·2) persons, respectively. Influenza-associated ILI incidence was highest among children aged 2–17 years. Observed trends were highly correlated with national ILI and virologic surveillance.

Conclusions:  This is the first multistate surveillance system demonstrating the feasibility of using outpatient practices to estimate the incidence of medically attended influenza at the community level. Surveillance demonstrated the substantial burden of pandemic influenza in outpatient settings and especially in children aged 2–17 years. Observed trends were consistent with established syndromic and virologic systems.

Annual influenza vaccination rates among hospital healthcare workers (HCW) are almost universally low despite recommendations from WHO and public health authorities in many countries. To assist in the development of successful vaccination programmes, we reviewed studies where interventions aimed to increase the uptake of influenza vaccination among hospital HCW. We searched PUBMED from 1990 up to December 2011 for publications with predetermined search strategies and of pre-defined criteria for inclusion or exclusion. We evaluated a large number of ‘intervention programmes’ each employing one or more ‘intervention components’ or strategies, such as easy access to vaccine or educational activities, with the goal to raise influenza vaccine uptake rates in hospital HCW during one influenza season. Included studies reported results of intervention programmes and compared the uptake with the season prior to the intervention (historical control) or to another intervention programme within the same season that started from the same set of baseline activities. Twenty-five studies performed in eight countries met our selection criteria and described 45 distinct intervention programmes. Most studies used their own facility as historical control and evaluated only one season. The following elements were used in intervention programmes that increased vaccine uptake: provision of free vaccine, easy access to the vaccine (e.g. through mobile carts or on-site vaccination), knowledge and behaviour modification through educational activities and/or reminders and/or incentives, management or organizational changes, such as the assignment of personnel dedicated to the intervention programme, long-term implementation of the strategy, requiring active declination and mandatory immunization policies. The number of these components applied appeared to be proportional to the increase in uptake. If influenza uptake in hospital HCW is to be increased on sustained basis, hospital managers need to be committed to conduct a well-designed long-term intervention programme that includes a variety of co-ordinated managerial and organizational elements.

Objective  To determine immunogenicity and safety of intradermal (ID) influenza vaccines compared with intramuscular (IM) administration and effect of dose and age.

Design  Meta-anlysis.

Setting  Systematic review and meta-analysis of randomized controlled trials on influenza vaccines.

Sample  Randomized, controlled trials comparing ID seasonal split-virus influenza vaccines with 15 μg IM control in subjects 18 years of age or older and assessed antibody response at 21–28 days post-vaccination were considered for inclusion.

Results  A total of 13 trials were included. The pooled immunogenicity outcomes did not differ significantly between the IM and ID vaccine groups for the H1N1 (ratio of GMTR: 0·92, 95% confidence interval 0·77–1·09; seroconversion: 0·94, 0·86–1·02; seroprotection: 0·97, 0·94–1·00) and B strains (GMTR: 0·93, 0·80–1·08; seroconversion: 0·91, 0·80–1·04; seroprotection: 0·97, 0·91–1·03). For the H3N2 strain, there was no significant difference in GMTR (0·97, 0·80–1·18); however, there was a lower pooled seroconversion (0·89, 0·80–0·99) and seroprotection rate (0·98, 0·96–0·99) for ID recipients. There was a statistically significant association between increasing doses of the ID vaccination with increasing immunogenicity response (P = 0·01). There were no differences in adverse event rates within 3 days post-vaccination for ID versus IM. But for adverse events occurring 7 days post-vaccination, ID vaccination was associated with a greater incidence of local events but not systemic events.

Conclusions  There was no significant difference in immunologic response when comparing ID with IM administration of the influenza vaccination in the overall population, but higher doses of ID vaccine in the older adult population produced a better response.

Background  Wild birds are the natural hosts for influenza A viruses (IAVs) and provide a niche for the maintenance of this virus.

Objectives  This study was undertaken to analyze nucleotide sequences of the matrix (M) gene of AIVs isolated from wild birds and live bird markets (LBMs) to index the changes occurring in this gene.

Methods  M-gene of 229 avian influenza virus (AIV) isolates obtained from wild birds and LBMs was amplified and sequenced. Full-length sequences (∼900 nt.) thus obtained were analyzed to identify changes that may be associated with resistance to adamantanes. Phylogenetic analysis of all sequences was performed using clustalw, and evolutionary distances were calculated by maximum composite likelihood method using mega (ver. 5.0) software.

Results  Twenty-seven different viral subtypes were represented with H3N8 being the most dominant subtype in wild birds and H7N2 being the predominant subtype among isolates from LBMs. Phylogenetic analysis of the M-gene showed a high degree of nucleotide sequence identity with US isolates of AIVs but not with those of Asian or European lineages. While none of the isolates from wild birds had any antiviral resistance–associated mutations, 17 LBM isolates carried polymorphisms known to cause reduced susceptibility to antiviral drugs (adamantanes). Of these 17 isolates, 16 had S31N change and one isolate had V27A mutation.

Conclusions  These results indicate independent evolution of M-gene in the absence of any antiviral drugs leading to mutations causing resistance indicating the need for continued active surveillance of AIVs.

Background  Influenza vaccination is the primary method for preventing influenza and its severe complications. An accurate rapid method to determine hemagglutinin (HA) concentration would facilitate reference antigen preparation and consequently expedite availability of seasonal as well as pandemic vaccines.

Objective  The goal of this study was to develop a label-free mass spectrometry (MS) based method that enables simultaneous identification and quantification of HA, neuraminidase (NA), and other viral proteins and protein contaminations in influenza vaccine or virus preparations.

Methods  The method presented is based on LC/MSE analysis of vaccine or virus preparations tryptic digests spiked with a known amount of protein standard from which a universal response factor is generated and applied to calculate the concentration of proteins identified in the mixture.

Results  We show that, with the use of an appropriate internal standard, the label-free MS-based protein quantification method is applicable for simultaneous identification and absolute quantification of HA and identification and relative quantification of other influenza proteins as well as protein impurities in influenza vaccines and virus preparations. We show that different subtype recombinant HA is preferred internal standard that provides the most accurate results in absolute quantification of HAs and other influenza proteins. We applied this method to measure the absolute quantity of HA as well as relative quantities of other viral proteins and impurities in preparations of whole virus and monovalent vaccine, providing data to demonstrate strain-dependent differences in the amount of NA.

Conclusion  The label-free MS method presented here is ideally suited for timely preparation of reference material needed for potency testing of seasonal and pandemic vaccines.

Background  Modern molecular techniques reveal new information on the role of respiratory viruses in community-acquired pneumonia. In this study, we tried to determine the prevalence of respiratory viruses and bacteria in patients with community-acquired pneumonia who were admitted to the hospital.

Methods  Between April 2008 and April 2009, 408 adult patients (aged between 20 and 94 years) with community-acquired pneumonia were tested for the presence of respiratory pathogens using bacterial cultures, real-time PCR for viruses and bacteria, urinary antigen testing for Legionella and Pneumococci and serology for the presence of viral and bacterial pathogens.

Results  Pathogens were identified in 263 (64·5%) of the 408 patients. The most common single organisms in these 263 patients were Streptococcus pneumoniae (22·8%), Coxiella burnetii (6·8%) and influenza A virus (3·8%). Of the 263 patients detected with pathogens, 117 (44·5%) patients were positive for one or more viral pathogens. Of these 117 patients, 52 (44·4%) had no bacterial pathogen. Multiple virus infections (≥2) were found in 16 patients.

Conclusion  In conclusion, respiratory viruses are frequently found in patients with CAP and may therefore play an important role in the aetiology of this disease.

Background  Timely influenza surveillance is important to monitor influenza epidemics.

Objectives  (i) To calculate the epidemic threshold for influenza-like illness (ILI) and acute respiratory infections (ARI) in 19 countries, as well as the thresholds for different levels of intensity. (ii) To evaluate the performance of these thresholds.

Methods  The moving epidemic method (MEM) has been developed to determine the baseline influenza activity and an epidemic threshold. False alerts, detection lags and timeliness of the detection of epidemics were calculated. The performance was evaluated using a cross-validation procedure.

Results  The overall sensitivity of the MEM threshold was 71·8% and the specificity was 95·5%. The median of the timeliness was 1 week (range: 0–4·5).

Conclusions  The method produced a robust and specific signal to detect influenza epidemics. The good balance between the sensitivity and specificity of the epidemic threshold to detect seasonal epidemics and avoid false alerts has advantages for public health purposes. This method may serve as standard to define the start of the annual influenza epidemic in countries in Europe.

Background  Highly pathogenic H5N1 influenza viruses reemerged in humans in 2003 and have caused fatal human infections in Asia and Africa as well as ongoing outbreaks in poultry. These viruses have evolved substantially and are now so antigenically varied that a single vaccine antigen may not protect against all circulating strains. Nevertheless, studies have shown that substantial cross-reactivity can be achieved with H5N1 vaccines. These studies have not, however, addressed the issue of duration of such cross-reactive protection.

Objectives  To directly address this using the ferret model, we used two recommended World Health Organization H5N1 vaccine seed strains – A/Vietnam/1203/04 (clade 1) and A/duck/Hunan/795/02 (clade 2.1) – seven single, double, or triple mutant viruses based on A/Vietnam/1203/04, and the ancestral viruses A and D, selected from sequences at nodes of the hemagglutinin and neuraminidase gene phylogenies to represent antigenically diverse progeny H5N1 subclades as vaccine antigens.

Results  All inactivated whole-virus vaccines provided full protection against morbidity and mortality in ferrets challenged with the highly pathogenic H5N1 strain A/Vietnam/1203/04 5 months and 1 year after immunization.

Conclusion  If an H5N1 pandemic was to arise, and with the hypothesis that one can extrapolate the results from three doses of a whole-virion vaccine in ferrets to the available split vaccines for use in humans, the population could be efficiently immunized with currently available H5N1 vaccines, while the homologous vaccine is under production.

Background  Respiratory viruses are known to cocirculate but this has not been described in detail during an influenza pandemic.

Objectives  To describe respiratory viruses, including co-infection and associated attributes such as age, sex or comorbidity, in patients presenting with influenza-like illness to a community sentinel network, during the pandemic A(H1N1)pdm09 in Ontario, Canada.

Methods  Respiratory samples and epidemiologic details were collected from 1018 patients with influenza-like illness as part of respiratory virus surveillance and a multiprovincial case–control study of influenza vaccine effectiveness.

Results  At least one virus was detected in 668 (65·6%) of 1018 samples; 512 (50·3%) had single infections and 156 (15·3%) co-infections. Of single infections, the most common viruses were influenza A in 304 (59·4%) samples of which 275 (90·5%) were influenza A(H1N1)pdm09, and enterovirus/rhinovirus in 149 (29·1%) samples. The most common co-infections were influenza A and respiratory syncytial virus B, and influenza A and enterovirus/rhinovirus. In multinomial logistic regression analyses adjusted for age, sex, comorbidity, and timeliness of sample collection, single infection was less often detected in the elderly and co-infection more often in patients <30 years of age. Co-infection, but not single infection, was more likely detected in patients who had a sample collected within 2 days of symptom onset as compared to 3–7 days.

Conclusions  Respiratory viral co-infections are commonly detected when using molecular techniques. Early sample collection increases likelihood of detection of co-infection. Further studies are needed to better understand the clinical significance of viral co-infection.

Background  Healthcare workers (HCWs) universally have a poor uptake of influenza vaccination. However, no data are available from India.

Objective  To explore knowledge, attitudes, and practices associated with influenza vaccination in HCWs in a temperate climate area in northern India.

Patients and Methods  A self-administered questionnaire was offered to all HCWs in three major hospitals of Srinagar and information sought on motivations, perceptions, preferences and practices regarding influenza vaccination.

Results  Of the 1750 questionnaires received, 1421 (81%) were returned. Only 62 (4·4%) HCWs had ever received influenza vaccination even as 1348 (95%) believed that influenza poses adverse potential consequences for themselves or their contacts; 1144 (81%) were aware of a vaccine against influenza and 830 (58%) of its local availability. Reasons cited by 1359 participants for not being vaccinated included ignorance about vaccine availability (435; 32%), skepticism about efficacy (248; 18%), busy schedule (166; 12%), fear of side effects (70; 4%), and a perception of not being-at-risk (82; 6%). Sixty-one percent (865) believed that vaccine programs are motivated by profit. Eighty-eight percent opined for mandatory vaccination for HCWs caring for the high-risk patients, as a part of ‘employee health program’. Most of the participants intended to get vaccinated in the current year even as 684 (48%) held that vaccines could cause unknown illness and 444 (31%) believed their adverse effects to be underreported.

Conclusion  Influenza vaccination coverage among HCWs is dismally low in Srinagar; poor knowledge of vaccine availability and misperceptions about vaccine effectiveness, fear of adverse effects and obliviousness to being-at-risk being important barriers. Multifaceted, adaptable measures need to be invoked urgently to increase the coverage.

Background  The effects of individual lifestyle factors on the mortality risk after influenza infection have not been explored.

Objectives  In this study, we assessed the modifying effects of cigarette smoking on mortality risks associated with influenza in a cohort of Hong Kong elders with a follow-up period of 1998–2009.

Methods  We used the Cox proportional hazards model with time-dependent covariates of weekly proportions of specimens positive for influenza (termed as influenza virus activity), to calculate the hazard ratio of mortality associated with a 10% increase in influenza virus activity for never, ex- and current smokers. Other individual lifestyle and socioeconomic factors as well as seasonal confounders were also added into the models.

Results  The overall hazard ratio associated with influenza was 1·028 (95% confidence interval, 1·006, 1·051) for all natural cause mortality and 1·035 (1·003, 1·068) for cardiovascular and respiratory mortality. We found that influenza-associated hazard ratio was greater in current and ex-smokers than in never smokers for mortality of all natural causes, cardiovascular and respiratory diseases.

Conclusions  The findings suggest that smoking might increase influenza-associated mortality risks among elders.

Background  PB1F2 is the 11th protein of influenza A virus translated from +1 alternate reading frame of PB1 gene. Since the discovery, varying sizes and functions of the PB1F2 protein of influenza A viruses have been reported. Selection of PB1 gene segment in the pandemics, variable size and pleiotropic effect of PB1F2 intrigued us to analyze amino acid sequences of this protein in various influenza A viruses.

Methods  Amino acid sequences for PB1F2 protein of influenza A H5N1, H1N1, H2N2, and H3N2 subtypes were obtained from Influenza Research Database. Multiple sequence alignments of the PB1F2 protein sequences of the aforementioned subtypes were used to determine the size, variable and conserved domains and to perform mutational analysis.

Results  Analysis showed that 96·4% of the H5N1 influenza viruses harbored full-length PB1F2 protein. Except for the 2009 pandemic H1N1 virus, all the subtypes of the 20th-century pandemic influenza viruses contained full-length PB1F2 protein. Through the years, PB1F2 protein of the H1N1 and H3N2 viruses has undergone much variation. PB1F2 protein sequences of H5N1 viruses showed both human- and avian host-specific conserved domains. Global database of PB1F2 protein revealed that N66S mutation was present only in 3·8% of the H5N1 strains. We found a novel mutation, N84S in the PB1F2 protein of 9·35% of the highly pathogenic avian influenza H5N1 influenza viruses.

Conclusions  Varying sizes and mutations of the PB1F2 protein in different influenza A virus subtypes with pandemic potential were obtained. There was genetic divergence of the protein in various hosts which highlighted the host-specific evolution of the virus. However, studies are required to correlate this sequence variability with the virulence and pathogenicity.

Objective  The objective of this study was to estimate the diagnostic sensitivity (Se) and specificity (Sp) of the HI test and six other diagnostic assays for the detection of AI antibodies without assuming a gold standard.

Methods  We applied a Bayesian version of latent class analysis to compare the results of multiple tests from different study settings reported in the literature.

Results  The results showed that the HI test has nearly perfect accuracy (i.e. 98·8% sensitivity and 99·5% specificity). It performed well in both chickens and turkeys and yet was less accurate in experimentally infected poultry, compared to naturally infected. Blocking ELISA and the indirect immunofluorescence assay also performed very well.

Conclusions  Given its very high Se and Sp, the HI test may be effectively considered a gold standard. In the framework of LPAI surveillance, where large numbers of samples have to be processed, the blocking ELISA could be a valid alternative to the HI test, in that it is almost as sensitive and specific as the HI test yet quicker and easier to automate.

Objective  This study investigated the inter- and intraspecies transmission of CIV among dogs, cats, and ferrets, under laboratory conditions, to determine whether transmission of the virus was possible between as well as within these domestic animal species.

Method  The transmission routes for inter- and intraspecies transmission were airborne and direct contact, respectively. Transmission was conducted through intranasal infection of dogs followed by exposure to either cats or ferrets and by comingling infected and naïve animals of the same species.

Results  The interspecies transmission of CIV H3N2 via airborne was only observed from dogs to cats and not from dogs to ferrets. However, direct intranasal infection of either cats or ferrets with CIV could induce influenza-like clinical signs, viral shedding, and serological responses. Additionally, naïve cats and ferrets could be infected by CIV via direct contact with infected animals of the same species.

Conclusion  Cats appear to be another susceptible host of CIV H3N2, whereas ferrets are not likely natural hosts. The molecular-based mechanism of interspecies and intraspecies transmission of CIV H3N2 should be further studied.

Methods  A total of 1147 sera obtained from pigs in Cambodia were tested by haemagglutination inhibition (HI) assays for antibody to human influenza A viruses along with both HI and microneutralization (MN) tests to assess immunological responses to H5N1 virus. The results were compared by year, age, and province.

Results  Antibodies against a human influenza A virus were detected in 14·9% of samples. A(H1N1)pdm09 virus were dominant over the study period (23·1%), followed by those to human H1N1 (17·3%) and H3N2 subtypes (9·9%). No pigs were serologically positive for avian H5 influenza viruses. The seroprevalence of human H1N1 and H3N2 influenza viruses peaked in 2008, while that of A(H1N1)pdm09 reached a peak in 2010. No significant differences in seroprevalence to human influenza subtypes were observed in different age groups.

Conclusions  Cambodian pigs were exposed to human strains of influenza A viruses either prior to or during this study. The implications of these high prevalence rates imply human-to-swine influenza virus transmission in Cambodia. Although pigs are mostly raised in small non-commercial farms, our preliminary results provide evidence of sustained human influenza virus circulation in pig populations in Cambodia.

Methods  From 2003–2007, we conducted surveillance for hospitalized respiratory illness in rural Thailand. We performed reverse-transcriptase polymerase chain reaction on nasopharyngeal specimens and enzyme immunoassay on paired sera

Results  Of 10,097 patients enrolled, 573 (5%) of all ages and 370 (9%) of children <5 years of age had evidence of HPIV infection (HPIV1=189, HPIV2=54, HPIV3=305, untyped=27). Average adjusted annual incidence of HPIV-associated hospitalized respiratory illness was greatest in children aged <1 year (485 per 100,000 person years).

Conclusions  In Thailand, HPIV caused substantial illnesses requiring hospitalization in young children.

Design:  Through mathematical and statistical modelling, we analysed Italian epidemiological and virological surveillance data collected together with serological data derived from influenza vaccine clinical trials performed in Italy.

Results:  Depending on the season, the reporting rate estimates ranged between 20% and 33% of the total incidence with higher reporting rates in seasons dominated by A/H3N2 virus. Despite a generally higher number of individuals immune against A/H3N2 viruses, effective reproduction ratios were quite similar in all seasons varying between 1·2 and 1·4. We observed an age-dependent transmissibility for different subtypes: susceptible children were more likely than susceptible adults and elderly to get infected when A/H1N1 or B strains were circulating, while no clear age-dependence was found for A/H3N2. We also perform sensitivity analysis under different assumptions for vaccine effectiveness, generation time (GT) and model variants; we found that the overall results in predicted patterns were extremely similar, with a slightly better fit obtained with shorter GTs.

Conclusions:  Our results provide relevant information on the influenza dynamics to fine-tune intervention strategies and for data collection improvement.

Objectives  We describe 8 years of sentinel surveillance data and the contribution of influenza and other viruses to medically attended influenza-like illness (ILI) in Côte d’Ivoire.

Methods  Sentinel surveillance was established in 2003. Nasopharyngeal (NP) specimens and epidemiologic data are collected from persons of all ages presenting with ILI at sentinel sites. Respiratory specimens have been tested for influenza using various viral and molecular diagnostic methods. A subset of 470 specimens collected from children aged 0–5 years were tested for multiple respiratory viruses using RT-PCR.

Results  From 2003 to 2010, 5074 NP specimens were collected from patients with ILI. Overall, 969/5074 (19%) of these specimens tested positive for influenza. Seasonal influenza A(H1N1) viruses predominated during 5 years and influenza A(H3N2) viruses predominated during 3 years. Influenza B viruses cocirculated with influenza A viruses during each year from 2004 to 2010. Seasonal peaks in influenza circulation were observed during the months of May, June, and October, with the largest peak corresponding with the primary rainfall season. Of 470 specimens collected from children under aged 5 who were tested for multiple respiratory viruses, a viral respiratory pathogen was detected in 401/470 (85%) of specimens. Commonly detected viruses were RSV (113 of 470 specimens, 24%), rhinoviruses (85/470, 18%), influenza (77/470, 16%), and parainfluenza (75/470, 16%).

Conclusion  In Côte d’Ivoire, there is a significant annual contribution of influenza and other respiratory viruses to medically attended ILI.

Objectives  To describe epidemiological and virologic characteristics of influenza in Lao PDR to recommend public health interventions, including improvements in surveillance and response.

Patients/Methods  We performed a descriptive analysis of samples taken from patients with influenza-like-illness (ILI) (fever >38°C with cough and/or sore throat) presenting at seven sentinel hospitals in three regions of Lao PDR, January 2008–December 2010. A nasopharyngeal (NP) swab or combined nasal with oropharyngeal swab was collected from patients with ILI. Samples were tested for influenza by either Luminex RVP, conventional reverse transcriptase PCR (RT-PCR) (January 2008–2009), or by real-time PCR (rRT-PCR) using US CDC reagents (February 2009 onward).

Results  Of 2346 samples tested from patients with ILI, 523 (22%) were positive for influenza. The median age of those positive was 12 years (range, <1–60 year). The percentage of samples that were influenza positive was similar over the 3 years (20–23%). Each year 3–4 types/subtypes cocirculated with differing predominant type/subtype. Influenza was detected year-round with the highest proportion of positive specimens in the 3rd and 4th quarter.

Conclusions  Similar to other countries in the region, we found that influenza is present year-round and has a peak activity from July to December. Dominant types or subtypes vary by year. A large proportion of patients with ILI are not influenza positive. ILI surveillance is critical for weighing disease burden, both morbidity and mortality, against the costs of advancing influenza vaccine delivery strategy.

Methodology  Demographic, clinical data, and respiratory specimens were collected for 4236 ILI patients tested for influenza virus infection by RT-PCR and viral culture.

Principal Findings  Influenza A and B viruses co-circulated year-round with seasonal peaks in influenza A virus activity during the rainy season (December–January). During 2003–2007, influenza viruses were identified in 20·1% (4236/21 030) of ILI patients, including 20·1% (4015/20 012) of outpatients, and 21·7% (221/1018) of inpatients. One H5N1 case was identified retrospectively in an outpatient with ILI. Antigenic drift in circulating influenza A and B virus strains was detected during the surveillance period in Indonesia. In a few instances, antigenically drifted viruses similar to the World Health Organization (WHO) vaccine strains were detected earlier than the date of their designation by WHO.

Conclusions  Influenza A and B virus infections are an important cause of influenza-like illness among outpatients and hospitalized patients in Indonesia. While year-round circulation of influenza viruses occurs, prevention and control strategies should be focused upon the seasonal peak during rainy season months. Ongoing virologic surveillance and influenza disease burden studies in Indonesia are important priorities to better understand the public health impact of influenza in South-East Asia and the implications of influenza viral evolution and global spread.

Design, setting and sample:  During July 2009–August 2011, we collected clinical data and specimens (nasal and throat swabs) from rural patients hospitalized for acute medical illnesses. Specimens were tested by rRT-PCR for influenza viruses.

Main outcome measures:  Case definitions evaluated the following: influenza-like illness (ILI: measured fever plus cough or sore throat); severe acute respiratory illness (SARI: ILI with difficulty breathing in ≥5 years, Integrated Management of Childhood Illness–defined pneumonia or severe pneumonia, or physician diagnosed lower respiratory infection in <5 years); acute respiratory infection (ARI: ≥1 of cough, nasal discharge, difficulty breathing or sore throat); febrile acute respiratory illness (FARI: fever plus either cough, sore throat, runny nose, difficulty breathing, or earache). Variants that included “reported fever” and additional sign–symptom combinations were also evaluated.

Results:  We enrolled 1043 hospitalized patients, including 257 children <5 years of age (range 1 day–86 years). Seventy-four patients tested influenza virus positive (including 28 A(H1N1)pdm09). Sensitivity(95% CI) and specificity (95% CI) for influenza infection were 78% (67–87) and 60% (57–63) for ILI (measured/reported fever); 37% (26–49) and 78% (75–80) for SARI (measured/reported fever); 82% (72–90) and 57% (54–60) for FARI (measured/reported fever); 88% (78–94) and 45% (42–49) for ARI; and 74% (63–84) and 61% (58–64) for measured/reported fever plus cough. Case definitions including only measured fever had lower sensitivity.

Conclusion:  ILI and FARI with measured/reported fever provided good balance between sensitivity and specificity among hospitalized patients. The simpler case definition of measured/reported fever plus cough is suited for field surveillance.

Objectives  We looked for respiratory viruses in a population-based sample of healthy adults with influenza-like illness (ILI). We investigated host and spatio-temporal associations with virus isolation and host, spatio-temporal and virus associations with self-reported symptoms.

Patients/Methods  We recruited 586 participants experiencing 651 illness episodes from a population of healthy adults enrolled in an influenza vaccine effectiveness trial. At ILI assessment visits, a respiratory swab was collected and tested for viruses using a combination of polymerase chain reaction (PCR) assays. Participants also completed a questionnaire detailing their clinical course in 336 episodes.

Results  Of 643 samples analysed, a virus was identified in 44%. Half were picornaviruses, with influenza and coronaviruses the next most common. Individuals with influenza were significantly less likely to have been immunised than the reference (virus negative) population (OR = 0·52 (0·31, 0·87) P = 0·01).

The mean symptom score (95% CI) reported by individuals with influenza was significantly higher than in all other episodes [Influenza: 10·2 (9·4, 10·9); Other: 7·4 (7·2, 7·7); Difference (95% CI): 2·5 (1·5, 3·5); P < 0·001]. In an analysis restricted to influenza-positive cases, the symptom score was not attenuated by vaccination.

Conclusions  Our findings indicate that a greater number of symptoms are displayed by individuals presenting with influenza confirmed ILI compared with other agents that cause ILI. While influenza vaccination reduced the probability of influenza virus detection, symptom score for influenza-positive ILI was not attenuated.

Objective  In this study, we developed HPAI virus-like particle (VLP) vaccine as a candidate poultry vaccine and evaluated its protective efficacy and possible application for differentiating infected from vaccinated animals (DIVA).

Methods  Specific pathogen-free chickens received a single injection of HPAI H5N1 VLP vaccine generated using baculovirus expression vector system. Immunogenicity of VLP vaccines was determined using hemagglutination inhibition (HI), neuraminidase inhibition (NI), and ELISA test. Challenge study was performed to evaluate efficacy of VLP vaccines.

Results and Conclusions  A single immunization with HPAI H5N1 VLP vaccine induced high levels of HI and NI antibodies and protected chickens from a lethal challenge of wild-type HPAI H5N1 virus. Viral excretion from the vaccinated and challenged group was strongly reduced compared with a mock-vaccinated control group. Furthermore, we were able to differentiate VLP-vaccinated chickens from vaccinated and then infected chickens with a commercial ELISA test kit, which offers a promising strategy for the application of DIVA concept.

Methods  Major databases, including MEDLINE, EMBASE, and the Cochrane Library, were searched for studies published between January 1966 and May 2009 that evaluated PCT as a marker for diagnosing bacterial infections in patients with influenza infections and that provided sufficient data to construct two-by-two tables.

Results  Six studies were selected that included 137 cases with bacterial coinfection and 381 cases without coinfection. The area under a summary ROC curve was 0·68 (95% CI: 0·64–0·72). The overall sensitivity and specificity estimates for PCT tests were 0·84 (95% CI: 0·75–0·90) and 0·64 (95% CI: 0·58–0·69), respectively. These studies reported heterogeneous sensitivity estimates ranging from 0·74 to 1·0. The positive likelihood ratio for PCT (LR+ = 2·31; 95% CI: 1·93–2·78) was not sufficiently high for its use as a rule-in diagnostic tool, while its negative likelihood ratio was reasonably low for its use as a rule-out diagnostic tool (LR− = 0·26; 95% CI: 0·17–0·40).

Conclusions  Procalcitonin tests have a high sensitivity, particularly for ICU patients, but a low specificity for identifying secondary bacterial infections among patients with influenza. Because of its suboptimal positive likelihood ratio and good negative likelihood ratio, it can be used as a suitable rule-out test but cannot be used as a standalone rule-in test.

Objective  This study examines socio-demographic differences in opinions about 2009 pandemic influenza A (H1N1) (pH1N1) and seasonal influenza disease and vaccines and the association with receipt of influenza vaccinations during the 2009–2010 influenza season. Changes in opinions over the course of the pH1N1 pandemic were also examined.

Methods  Data from the 2009 National H1N1 Flu Survey (NHFS) were analyzed. The NHFS was a CDC-sponsored telephone survey initiated in response to the 2009 pH1N1 pandemic to obtain weekly within-season estimates of vaccination coverage, opinions, and other information.

Results  Opinions about influenza vaccine and disease varied significantly by race/ethnicity, income, and education level. In multivariable logistic regression analysis, adjusted 2009 H1N1 vaccination coverage was most strongly associated with opinions about the effectiveness of the vaccine and personal risk of disease, varying from 7 to 11% among adults who believed the vaccine to have low effectiveness and themselves at low risk of influenza, to 50–53% among those who thought vaccine effectiveness to be high and themselves at high risk of influenza.

Conclusion  Improving communication about personal risk and the effectiveness of influenza vaccines may improve vaccination coverage. The findings of difference in opinions could be used to target communication.

Objectives To use pig ex vivo organ explants to assess the susceptibility of pigs to infection with contemporary viruses, for which there is evidence of human infection and that are thought to pose the greatest threat to pig and human populations.

Methods  Pig tracheal, bronchi and lung ex vivo organ explants were infected with both highly pathogenic and low pathogenic avian influenza (AI) virus and the pandemic H1N1 [A(H1N1)pdm/09] virus. Successful infection of explants was detected using a positive-sense RNA real-time RT-PCR assay and anti-nucleoprotein immunohistochemistry. The distribution of cell-surface α2-3- and α2-6-linked sialic acid receptors, the avian- and mammalian influenza A virus–preferred host receptors, respectively, was also characterised for the ex vivo organ cultures and uninfected pig material following necropsy.

Results  The α2-3 and α2-6 sialic acid receptor staining on tracheal, bronchi and lung organ explant sections showed similar distributions to those seen for pig tissue following necropsy. While the pig ex vivo organ cultures were susceptible to nearly all viruses tested, lower levels of virus were detected in trachea and bronchi after infection.

Conclusion  These results confirm that pigs are susceptible to contemporary viruses that may threaten both veterinary and human health and contribute to the ecology of influenza A viruses.

Objectives  To assess the diagnostic accuracy of these case definitions and to attempt to improve on them using a scoring system based on clinical findings at presentation.

Patients/Methods  This study is a retrospective case–control study across three metropolitan Melbourne hospitals and one associated community-based clinic during the influenza season, 2009. Patients presenting with influenza-like illness who were tested for H1N1/09 influenza A were administered a standard questionnaire of symptomatology, comorbidities, and risk factors. Patients with a positive test were compared to those with a negative test. Logistic regression was performed to examine for correlation of clinical features with disease. A scoring system was devised and compared with case definitions used during the pandemic. The main outcome measures were the positive and negative predictive values of our scoring system, based on real-life data, versus the mandated case definitions’.

Results  Both the devised scoring system and the case definitions gave similar positive predictive values (38–58% using ascending score groups, against 39–44% using the various case definitions). Negative predictive values were also closely matched (ranging from 94% to 73% in the respective score groups against 83–84% for the case definitions).

Conclusions  Accurate clinical diagnosis of H1N1/09 influenza A was difficult and not improved significantly by a structured scoring system. Investment in more widespread availability of rapid and sensitive diagnostic tests should be considered in future pandemic planning.

Methods  Using medical charts, we collected data on 242 patients who were hospitalized with confirmed laboratory results (defined as positive by specific PCR for pandemic 2009 influenza A H1N1).

Results  During the study period, there were 242 cases of hospitalization or death. Of the 242, 46% were admitted to an intensive care unit (ICU) and 33·5% died. The mean age was 27·8 years for surviving and 39·6 for those who died. Twenty-eight percent of hospitalizations involved persons under the age of 15 years; 33% of the patients were between the age of 15 and 44 years; and only 3·3% were 65 years of age or older. Sixty-seven percent had an underlying medical conditions, including diabetes, obesity, heart and lung diseases, and pregnancy. Of the 242 patients, 68% had findings consistent with pneumonia. Treatment with oseltamivir was administered to 227 (93·8%) patients from which 38 received oseltamivir within 48 hours after the onset of symptoms.

Conclusions  The pandemic strain caused severe illness, including pneumonia and acute respiratory distress syndrome, and resulted in ICU admissions in 46% of patients and death in 33·5%. The mean age of hospitalized infected cases was lower than is common with seasonal influenza. Underlying medical conditions were common in the 67% the evaluated patients. Patients who died had a higher prevalence of comorbidities (86·4%) than those who survived (57%), suggesting that the presence of chronic illness may increase the likelihood of death. However, the severe illness was also identified among young, healthy persons.

Methods  We identified index patients with laboratory confirmed influenza infection and interviewed household members after illness day 8 of the index patient. Outcome was influenza-like illness (ILI) in a household contact.

Results  We included 108 households. Overall secondary attack rate was 10·1% (27/267) and decreased with increasing age. Apart from being in close daily proximity with the index patient for at least 9 hours, no other behavioral risk factor was associated with secondary ILI. Of all index patients and household contacts, 49% and 55%, respectively, cleaned their hands more often in the week after symptom onset of the index patient (in comparison with 2008/2009 P-value for both <0·01).

Conclusions  While the study was hampered by its relatively limited size, data suggest that a significantly larger proportion of influenza households practiced good hand hygiene compared to the last pre-pandemic season. This may have led to a different risk factor profile and a delay of the time threshold necessary for transmission among household members with close contact.

Design  Observational study from sentinel surveillance sites.

Setting  Bhutan remains isolated, with only one to two flights a day at the lone airport, no trains, and only three major roads that enter from India.

Main outcome measures  PCR positive human respiratory samples

Results  The first case of A(H1N1)pdm09 infection was detected in Bhutan in July 2009, 3 months after the virus was first reported in Mexico in April 2009. During the official WHO pandemic period (11 June 2009 to 8 August 2010), a total of 2149 samples were collected and tested by RT-PCR of which 22.7% (487) were confirmed A(H1N1)pdm09; H3N2, H1N1, and B were positive in 2.2%, 1.1%, and 7.2%, respectively. The highest rate of A(H1N1)pdm09 cases (57.4%) was detected in the 6-20 year-old age group. Importantly, Bhutan increased from 3 sentinel sites in April 2009 to 11 a year later, and in April 2010 established PCR capability for influenza.

Conclusions  Despite relative isolation, the A(H1N1)pdm09 reached Bhutan within 3 months of identification in Mexico. The H1N1 pandemic has made Bhutan more prepared for epidemics in the future.

Objectives  Influenza vaccine-induced humoral immunity against several isolates of influenza A(H1N1)pdm09 virus was analyzed in healthcare professionals. Age-dependent responses were also analyzed.

Methods  Influenza viruses were selected to represent viruses that circulated in Finland during two consecutive influenza epidemic seasons 2009–2010 and 2010–2011. Serum samples from vaccinated volunteers, age 20–64 years, were collected before and after vaccination with AS03-adjuvanted pandemic and non-adjuvanted trivalent seasonal influenza vaccine that was given 1 year later.

Results  Single dose of pandemic vaccine induced a good albeit variable antibody response. On day 21 after vaccination, depending on the virus strain, 14–75% of vaccinated had reached antibody titers (≥1:40) considered seroprotective. The booster vaccination 1 year later with a seasonal vaccine elevated the seroprotection rate to 57–98%. After primary immunization, younger individuals (20–48 years) had significantly higher antibody titers against all tested viruses than older persons (49–64 years) but this difference disappeared after the seasonal booster vaccination.

Conclusions  Even a few amino acid changes in influenza A HA may compromise the vaccine-induced antibody recognition. Older adults (49 years and older) may benefit more from repeated influenza vaccinations.

Methods  Observational retrospective study using data collected by the Spanish National Hospital Discharge Database. We selected all admissions with diagnosis ICD-9-CM code 488·1 [A(H1N1)pdm09]. Discharges were grouped as follows: no diabetes, Type1 and Type 2 diabetes. Underlying medical conditions and risk factors included all those that constitute an indication for annual influenza vaccination, pregnancy, and obesity. The outcome variables analyzed were in-hospital case fatality risk, length of hospital stay, and costs.

Results  The total number of persons hospitalized with A(H1N1)pdm09 was 11 499. Of those, 97 suffered Type 1 and 936 Type 2, giving an overall prevalence of diabetes of 9%. The most common underlying medical condition among Type 2 subjects was obesity (26·8%), and for Type 1 renal disease (10·3%). In-hospital mortality was 2·1% among Type 1 patients, 3·8% among Type 2 patients, and 2·3% among non-diabetics; after multivariate analysis, diabetes was not a factor independently associated with dying during hospitalization for A(H1N1)pdm09. Independent factors increasing the risk of death among diabetic patients included age (OR 1·03; 95% CI1·01–1·05), hematological disorders (OR 3·49; 95% CI, 1·46–8·37), and obesity (OR 1·88; 95% CI1·07–3·92).

Conclusions  Among individuals hospitalized in Spain with A(H1N1)pdm09 infections, the age-specific prevalence of diabetes was higher than the general population in most age groups. The results of multivariate analysis suggest that possibly concomitant conditions such as obesity increase the risk of dying from the infection, but not diabetes itself.

Methods  The clinical symptoms and duration of fever (body temperature ≥37·5°C) after the first dose of an NAI (oseltamivir, zanamivir, laninamivir) were analyzed. PCR was carried out for 365 patients with A(H1N1)pdm09 in the 2009–2010 season and for 388 patients with one of the three types of influenza circulating in the 2010–2011 season. IC₅₀ for the three NAIs was also analyzed in 51 patients in the 2010–2011 season.

Results  The peak body temperature was significantly higher in 2010–2011 than in 2009–2010 for patients under 20 years with A(H1N1)pdm09, and in the 2010–2011 season for children 15 years or younger with A(H1N1)pdm09 than for those with other virus types. The percentage of A(H1N1)pdm09 patients with loss of appetite or fatigue was significantly higher in 2010–2011 than in the previous season. The duration of fever was not affected by the kind of NAI or by age in multiple regression analysis. The percentage of patients afebrile at 48 hours after the first dose of NAI was significantly higher for A(H1N1)pdm09 than for A(H3N2) (laninamivir) or B (oseltamivir and laninamivir).

Conclusion  Although the clinical symptoms of A(H1N1)pdm09 were slightly more severe in the 2010–2011 season, the effectiveness of the NAIs remained high in comparison with 2009–2010 and with other types of seasonal influenza.

Objective  To examine immunogenicity of influenza A(H1N1)pdm09 vaccine in patients with liver disease and to explore the associated factors on lowered immune response.

Patients/Methods  A single subcutaneous dose of monovalent inactivated unadjuvanted split-virus influenza A(H1N1)pdm09 vaccination was performed in 80 patients with chronic hepatitis C virus infection at Osaka City University Hospital in Japan. To measure the hemagglutination inhibition antibody titer, serum samples were collected before and 3 weeks after vaccination.

Results  No serious adverse events were observed. After vaccination, antibody titers ≥1:40 were observed in 56 patients (71%). The corresponding seroconversion proportion was 72%, and the mean fold rise was 10·3. Immune responses were robust regardless of severity of liver disease or existence of probable cirrhosis. However, patients with older age, lower body mass index, or receiving Stronger Neo-Minophagen C tended to show lower antibody responses to A(H1N1)pdm09 vaccine. In addition, reduced immune responses were observed in patients who had received the 2009/10 seasonal vaccination prior to A(H1N1)pdm09 vaccination.

Conclusions  Single dose of A(H1N1)pdm09 vaccine achieved a sufficient level of immunity among patients with chronic hepatitis C. Antibody response may be affected by age, body mass index, Stronger Neo-Minophagen C administration, and recent seasonal influenza vaccination.

Objectives  The aim of this study was to examine factors associated with the transmission of pandemic (H1N1) 2009 among HCWs.

Methods  A 1:4 matched case–control study by hospital, ward, age, and gender was conducted in HCWs from hospitals in Beijing during February 2010. Cases were diagnosed with pandemic (H1N1) 2009, and controls had neither influenza-like illness nor diagnosis with pandemic (H1N1) 2009 during August 2009 to January 2010. Information during 7 days before symptom onset of case was collected, and controls were queried about the same period.

Results  A total of 51 cases identified via National Notifiable Infectious Disease Surveillance System participated in this study. Controls were matched to cases for a total of 255 individuals. About 19·6% (10/51) of cases and 26·0% (53/204) of controls recalled they had conducted a high-risk procedure on a patient with pandemic (H1N1) 2009. 72·5% (37/51) of cases and 71·6% (146/204) of controls stated they wore medical masks in ≥80% of working time. Only 5·9% (3/51) and 36·3% (74/204) of cases and controls, respectively, reported receiving pandemic vaccination. Participants receiving pandemic vaccination had a significantly lower risk of infection during the pandemic (OR = 0·150, 95% CI: 0·047–0·479, P = 0·001). The estimated vaccine effectiveness was 85·0%.

Conclusions  We showed a high vaccine effectiveness of the pandemic vaccine and that vaccination was the only factor significantly associated with risk of infection in HCWs.

Methods  To develop a method for determining P_aO₂:F_iO₂ ratios, uninfected mice were anesthetized with pentobarbital, diazepam/ketamine, or inhaled isoflurane. Subsequently, they were allowed to breathe spontaneously or were mechanically ventilated. After 15 minutes exposure to room air (F_iO₂ = 0·21) or 100% O₂ (F_iO₂ = 1·0), carotid P_aO₂ was measured. To determine influenza effects on P_aO₂:F_iO₂, mice were challenged with 10 000 p.f..u./mouse influenza A/WSN/33.

Results  P_aO₂:F_iO₂ ratios were abnormally low (≤400 mmHg) in spontaneously breathing mice. Mechanical ventilation with positive end-expiratory pressure was required to obtain P_aO₂:F_iO₂ ratios in uninfected mice consistent with normal values in humans (≥600 mmHg). At day 2 following infection P_aO₂:F_iO₂ ratios indicated the onset of acute lung injury. By day 6, P_aO₂:F_iO₂ ratios were <200 mmHg, indicating progression to ARDS. Impaired gas exchange in influenza-infected mice was accompanied by progressive hemoglobin desaturation, hypercapnia, uncompensated respiratory acidosis, hyperkalemia, and polycythemia.

Conclusions  Influenza infection of mice results in impairment of alveolar gas exchange consistent with rapid development of acute lung injury and progression to ARDS. P_aO₂:F_iO₂ ratios may be of utility as clinically relevant and predictive outcome measures in influenza pathogenesis and treatment studies that use mouse models.

Results and Conclusions  As the panel of mAbs included antibodies with hemagglutination inhibition (HI) and virus neutralizing activities, this allowed identification and characterization of potentially important antigenic and neutralizing epitopes of the A/California/04/2009 HA and comparison of those epitopes with the HAs of other influenza viruses including seasonal H1N1 viruses as well as the A/South Carolina/1918 and A/New Jersey/1976 H1N1 viruses. Three mAbs with the highest HI and neutralizing titers were able to provide passive protection against virus challenge. Two other mAbs without HI or neutralizing activities were able to provide partial protection against challenge. HA epitopes recognized by the strongest neutralizing mAbs in the panel were identified by isolation and selection of virus escape mutants in the presence of individual mAbs. Cloned viruses resistant to HI and antibody neutralization were sequenced to identify mutations, and two unique mutations (D127E and G155E) were identified, both near the antigenic site Sa. Using human post-vaccination sera, however, there were no differences in HI titer between A/California/04/2009 and either escape mutant, suggesting that these single mutations were not sufficient to abrogate a protective antibody response to the vaccine.