Since 2020, there has been unprecedented global spread of highly pathogenic avian influenza A(H5N1) in wild bird populations with spillover into a variety of mammalian species and sporadically humans(1). In March 2024, clade 2.3.4.4b A(H5N1) virus was first detected in dairy cattle in the U.S., with subsequent detection in numerous states(2), leading to over a dozen confirmed human cases(3,4). In this study, we employed the ferret model, a well-characterized species that permits concurrent investigation of viral pathogenicity and transmissibility(5) in the evaluation of A/Texas/37/2024 (TX/37) A(H5N1) virus isolated from a dairy farm worker in Texas(6). Here, we show that the virus has a remarkable ability for robust systemic infection in ferrets, leading to high levels of virus shedding and spread to naïve contacts. Ferrets inoculated with TX/37 rapidly exhibited a severe and fatal infection, characterized by viremia and extrapulmonary spread. The virus efficiently transmitted in a direct contact setting and was capable of indirect transmission via fomites. Airborne transmission was corroborated by the detection of infectious virus shed into the air by infected animals, albeit at lower levels compared to the highly transmissible human seasonal and swine-origin H1 subtype strains. Our results show that despite maintaining an avian-like receptor binding specificity, TX/37 displays heightened virulence, transmissibility, and airborne shedding relative to other clade 2.3.4.4b virus isolated prior to the 2024 cattle outbreaks(7), underscoring the need for continued public health vigilance.

BACKGROUND: Anopheles stephensi is an invasive malaria vector in Africa that threatens to put an additional 126 million people at risk of malaria if it continues to spread. The island nation of Mauritius is highly connected to Asia and Africa and is at risk of introduction due to this connectivity. For early detection of An. stephensi, the Vector Biology and Control Division under the Ministry of Health in Mauritius, leveraged a well-established Aedes program, as An. stephensi is known to share Aedes habitats. These efforts triggered multisectoral coordination and cascading benefits of integrated vector and One Health approaches. METHODS: Beginning June 2021, entomological surveys were conducted at points of entry (seaport, airport) and on ships transporting livestock in collaboration with the Civil Aviation Department, the Mauritian Port Authority and National Veterinary Services. A total of 18, 39, 723 mosquito larval surveys were respectively conducted in the airport, seaport, and other localities in Mauritius while two, 20, and 26 adult mosquito surveys were respectively conducted in the airport, seaport, and twenty-six animal assembly points. Alongside adult mosquito surveys, surveillance of vectors of veterinary importance (e.g.- Culicoides spp.) was also carried out in collaboration with National Parks and Conservation Service and land owners. RESULTS: A total of 8,428 adult mosquitoes were collected and 1,844 larval habitats were positive for mosquitoes. All collected mosquitoes were morphologically identified and 151 Anopheles and 339 Aedes mosquitoes were also molecularly characterized. Mosquito species detected were Aedes albopictus, Anopheles arabiensis, An. coustani, An. merus, Culex quinquefasciatus, Cx. thalassius and Lutzia tigripes. Anopheles stephensi was not detected. The One Health approach was shared with the French Agricultural Research Centre for International Development (CIRAD), strengthening collaboration between Mauritius and Réunion Island on vector surveillance at entry points and insecticide resistance monitoring. The Indian Ocean Commission (IOC) was also alerted to the risk of An. stephensi, leading to regional efforts supporting trainings and development of a response strategy to An. stephensi bringing together stakeholders from Comoros, Madagascar, Mauritius, Réunion Island and Seychelles. CONCLUSIONS: Mauritius is a model system showing how existing public health entomology capabilities can be used to enhance vector surveillance and control and create multisectoral networks to respond to any emerging public and veterinary health vector-borne disease threat.

Chronic disease affects 6 in 10 adults in the US, while 4 in 10 adults live with multiple chronic diseases (1). Chronic diseases represent one of the nation’s leading causes of disability and drivers of the nation’s $4.1 trillion in annual health care spending (1). Chronic conditions including heart disease, cancer, stroke, diabetes, and chronic kidney disease dominate the leading causes of death. Furthermore, leading lifestyle risk factors in the US include tobacco use, poor nutrition, physical inactivity, and excessive alcohol use (1). | | Chronic disease prevention and control necessitates a comprehensive strategy to prevent disease (2–4), which is needed now more than ever (5). Information systems innovations are needed to advance health activities and outcomes and to allow decision makers and practitioners to act (4,6–8). Chronic disease data are a foundation that can inform interventions to promote healthy communities, support healthy behaviors and lifestyles, and facilitate effective and coordinated chronic disease prevention and health promotion (5,9). The benefits of an improved chronic disease data landscape include improved management of chronic disease programs, enhanced communication, data exchange, and coordination between federal, state, tribal, local, and territorial health departments and their partners. Additionally, efforts aimed at enhancing chronic disease surveillance practices will better enable a learning health system, precision public health, and improved situational awareness that will ultimately allow people across the US to live longer, healthier lives (10–12).

The globular head domain of influenza virus surface protein hemagglutinin (HA1) is the major target of neutralizing antibodies elicited by vaccines. As little as one amino acid substitution in the HA1 can result in an antigenic drift of influenza viruses, indicating the dominance of some epitopes in the binding of HA to polyclonal serum antibodies. Therefore, identifying dominant binding epitopes of HA is critical for selecting seasonal influenza vaccine viruses. In this study, we have developed a biolayer interferometry (BLI)-based assay to determine dominant binding epitopes of the HA1 in antibody response to influenza vaccines using a panel of recombinant HA1 proteins of A(H1N1)pdm09 virus with each carrying a single amino acid substitution. Sera from individuals vaccinated with the 2010-2011 influenza trivalent vaccines were analyzed for their binding to the HA1 panel and hemagglutination inhibition (HI) activity against influenza viruses with cognate mutations. Results revealed an over 50% reduction in the BLI binding of several mutated HA1 compared to the wild type and a strong correlation between dominant residues identified by the BLI and HI assays. Our study demonstrates a method to systemically analyze antibody immunodominance in the humoral response to influenza vaccines.

Generations of medical educators have recommended including public and population health (PPH) content in the training of U.S. physicians. The COVID-19 pandemic, structural racism, epidemic gun violence, and the existential threats caused by climate change are currently unsubtle reminders of the essential nature of PPH in medical education and practice. To assess the state of PPH content in medical education, the authors reviewed relevant guidance, including policies, standards, and recommendations from national bodies that represent and oversee medical education for physicians with MD degrees. Findings confirm that guidance across the medical education continuum, from premedical education to continuing professional development, increasingly includes PPH elements that vary in specificity and breadth. Graduate medical education policies present the most comprehensive approach in both primary care and subspecialty fields. Behavioral, quantitative, social, and systems sciences are represented, although not uniformly, in guidance for every phase of training. Quantitative PPH skills are frequently presented in the context of research, but not in relation to the development of population health perspectives (e.g., evidence-based medicine, quality improvement, policy development). The interdependence between governmental public health and medical practice, environmental health, and the impact of structural racism and other systems of oppression on health are urgent concerns, yet are not consistently or explicitly included in curricular guidance. To prepare physicians to meet the health needs of patients and communities, educators should identify and address gaps and inconsistencies in PPH curricula and related guidance. Re-examinations of public health and health care systems in the wake of the COVID-19 pandemic support the importance of PPH in physician training and practice, as physicians can help to bridge clinical and public health systems. This review provides an inventory of existing guidance (presented in the appendices) to assist educators in establishing PPH as an essential foundation of physician training and practice.

BACKGROUND: SARS-CoV-2 and influenza viruses continue to co-circulate, representing two major public health threats from respiratory infections with similar clinical presentations. SARS-CoV-2 and influenza vaccines can also now be co-administered. However, data on antibody responses to SARS-CoV-2 and influenza co-infection, and vaccine co-administration remains limited. METHODS: We developed a 41-plex antibody immunity assay that can simultaneously characterize antibody landscapes to SARS-CoV-2/influenza/common human coronaviruses. We analyzed sera from 840 individuals (11-93 years), including sera from reverse transcription polymerase chain reaction (RT-PCR) confirmed SARS-CoV-2 positive (n = 218) and negative (n = 120) cases, paired sera from SARS-CoV-2 vaccination (n = 29) and infection (n = 11), and paired sera from influenza vaccination (n = 56) and RT-PCR confirmed influenza infection (n = 158) cases. Lastly, we analyzed sera collected from 377 individual that exhibited acute respiratory illness (ARI) in 2020. RESULTS: This 41-plex assay has high sensitivity and specificity in detecting SARS-CoV-2 infections. It differentiated SARS-CoV-2 vaccination (antibody responses only to spike protein) from infection (antibody responses to both spike and nucleoprotein). No cross-reactive antibodies were detected to SARS-CoV-2 from influenza vaccination and infection, and vice versa, suggesting no interaction between SARS-CoV-2 and influenza antibody responses. However, cross-reactive antibodies were detected between spike proteins of SARS-CoV-2 and common human coronaviruses that were removed by serum adsorption. Among 377 individual who exhibited ARI in 2020, 129 were influenza positive, none had serological evidence of SARS-CoV-2/influenza co-infections. CONCLUSIONS: Multiplex detection of antibody landscapes can provide in-depth analysis of the antibody protective immunity to SARS-CoV-2 in the context of other respiratory viruses including influenza.

To better understand the antibody landscape changes following influenza virus natural infection and vaccination, we developed a high-throughput multiplex influenza antibody detection assay (MIADA) containing 42 recombinant hemagglutinins (rHAs) (ectodomain and/or globular head domain) from pre-2009 A(H1N1), A(H1N1)pdm09, A(H2N2), A(H3N2), A(H5N1), A(H7N7), A(H7N9), A(H7N2), A(H9N2), A(H13N9), and influenza B viruses. Panels of ferret antisera, 227 paired human sera from vaccinees (children and adults) in 5 influenza seasons (2010 to 2018), and 17 paired human sera collected from real-time reverse transcription-PCR (rRT-PCR)-confirmed influenza A(H1N1)pdm09, influenza A(H3N2), or influenza B virus-infected adults were analyzed by the MIADA. Ferret antisera demonstrated clear strain-specific antibody responses to exposed subtype HA. Adults (19 to 49 years old) had broader antibody landscapes than young children (<3 years old) and older children (9 to 17 years old) both at baseline and post-vaccination. Influenza vaccination and infection induced the strongest antibody responses specific to HA(s) of exposed strain/subtype viruses and closely related strains; they also induced cross-reactive antibodies to an unexposed influenza virus subtype(s), including novel viruses. Subsequent serum adsorption confirmed that the cross-reactive antibodies against novel subtype HAs were mainly induced by exposures to A(H1N1)/A(H3N2) influenza A viruses. In contrast, adults infected by influenza B viruses mounted antibody responses mostly specific to two influenza B virus lineage HAs. Median fluorescence intensities (MFIs) and seroconversion in MIADA had good correlations with the titers and seroconversion measured by hemagglutination inhibition and microneutralization assays. Our study demonstrated that antibody landscape analysis by the MIADA can be used for influenza vaccine evaluations and characterization of influenza virus infections.IMPORTANCE Repeated influenza vaccination and natural infections generate complex immune profiles in humans that require antibody landscape analysis to assess immunity and evaluate vaccines. However, antibody landscape analyses are difficult to perform using traditional assays. Here, we developed a high-throughput, serum-sparing, multiplex influenza antibody detection assay (MIADA) and analyzed the antibody landscapes following influenza vaccination and infection. We showed that adults had broader antibody landscapes than children. Influenza vaccination and infection not only induced the strongest antibody responses to the hemagglutinins of the viruses of exposure, but also induced cross-reactive antibodies to novel influenza viruses that can be removed by serum adsorption. There is a good correlation between the median fluorescence intensity (MFI) measured by MIADA and hemagglutination inhibition/microneutralization titers. Antibody landscape analysis by the MIADA can be used in influenza vaccine evaluations, including the development of universal influenza vaccines and the characterization of influenza virus infections.

Curriculum models and training activities in medical education have been markedly enhanced to prepare physicians to address the health needs of diverse populations and to advance health equity. While different teaching and experiential learning activities in the public health and population health sciences have been implemented, there is no existing framework to measure the effectiveness of public and population health (PPH) education in medical education programs. In 2015, the Association of American Medical Colleges (AAMC) established the Expert Panel on Public and Population Health in Medical Education, which convened 20 U.S. medical faculty members whose goal was to develop an evaluation framework adapted from the New World Kirkpatrick Model. Institutional leaders can use this framework to assess the effectiveness of PPH curricula for learners, faculty, and community partners. It may also assist institutions with identifying opportunities to improve the integration of PPH content into medical education programs. In this article, the authors present outcomes metrics and practical curricular or institutional illustrations at each Kirkpatrick training evaluation level to assist institutions with the measurement of (1) reaction to the PPH education content, (2) learning accomplished, (3) application of knowledge and skills to practice, and (4) outcomes achieved as a result of PPH education and practice. A fifth level was added to measure the benefit of PPH curricula on the health system and population health. The framework may assist with developing a locally relevant evaluation to further integrate and support PPH education at U.S. medical schools and teaching hospitals.

OBJECTIVE: To estimate prevalence levels of and time trends for active syphilis, gonorrhoea and chlamydia in women aged 15-49 years in four countries in the Pacific (Fiji, the Federated States of Micronesia [FSM], Papua New Guinea [PNG] and Samoa) to inform surveillance and control strategies for sexually transmitted infections (STIs). METHODS: The Spectrum-STI model was fitted to data from prevalence surveys and screenings of adult female populations collected during 1995-2017 and adjusted for diagnostic test performance and to account for undersampled high-risk populations. For chlamydia and gonorrhoea, data were further adjusted for age and differences between urban and rural areas. RESULTS: Prevalence levels were estimated as a percentage (95% confidence interval). In 2017, active syphilis prevalence was estimated in Fiji at 3.89% (2.82 to 5.06), in FSM at 1.48% (0.93 to 2.16), in PNG at 3.91% (1.67 to 7.24) and in Samoa at 0.16% (0.07 to 0.37). For gonorrhoea, the prevalence in Fiji was 1.63% (0.50 to 3.87); in FSM it was 1.59% (0.49 to 3.58); in PNG it was 11.0% (7.25 to 16.1); and in Samoa it was 1.61% (1.17 to 2.19). The prevalence of chlamydia in Fiji was 24.1% (16.5 to 32.7); in FSM it was 23.9% (18.5 to 30.6); in PNG it was 14.8% (7.39 to 24.7); and in Samoa it was 30.6% (26.8 to 35.0). For each specific disease within each country, the 95% confidence intervals overlapped for 2000 and 2017, although in PNG the 2017 estimates for all three STIs were below the 2000 estimates. These patterns were robust in the sensitivity analyses. DISCUSSION: This study demonstrated a persistently high prevalence of three major bacterial STIs across four countries in WHO's Western Pacific Region during nearly two decades. Further strengthening of strategies to control and prevent STIs is warranted.

Low pathogenicity avian influenza A(H9N2) viruses, enzootic in poultry populations in Asia, are associated with fewer confirmed human infections but higher rates of seropositivity compared to A(H5) or A(H7) subtype viruses. Co-circulation of A(H5) and A(H7) viruses leads to the generation of reassortant viruses bearing A(H9N2) internal genes with markers of mammalian adaptation, warranting continued surveillance in both avian and human populations. Here, we describe active surveillance efforts in live poultry markets in Vietnam in 2018 and compare representative viruses to G1 and Y280 lineage viruses that have infected humans. Receptor binding properties, pH thresholds for HA activation, in vitro replication in human respiratory tract cells, and in vivo mammalian pathogenicity and transmissibility were investigated. While A(H9N2) viruses from both poultry and humans exhibited features associated with mammalian adaptation, one human isolate from 2018, A/Anhui-Lujiang/39/2018, exhibited increased capacity for replication and transmission, demonstrating the pandemic potential of A(H9N2) viruses.IMPORTANCE A(H9N2) influenza viruses are widespread in poultry in many parts of the world, and for over twenty years, have sporadically jumped species barriers to cause human infection. As these viruses continue to diversify genetically and antigenically, it is critical to closely monitor viruses responsible for human infections, to ascertain if A(H9N2) viruses are acquiring properties that make them better suited to infect and spread among humans. In this study, we describe an active poultry surveillance system established in Vietnam to identify the scope of influenza viruses present in live bird markets and the threat they pose to human health. Assessment of a recent A(H9N2) virus isolated from an individual in China in 2018 is also reported and was found to exhibit properties of adaptation to humans and, importantly, show similarities to strains isolated from the live bird markets of Vietnam.

Of the eighteen hemagglutinin (HA) subtypes (H1-H18) that have been identified in bats and aquatic birds, many HA subtypes have been structurally characterized. However, several subtypes (H8, H11 and H12) still require characterization. To better understand all of these HA subtypes at the molecular level, HA structures from an A(H4N6) (A/swine/Missouri/A01727926/2015), an A(H8N4) (A/turkey/Ontario/6118/1968), an A(H11N9) (A/duck/Memphis/546/1974), an A(H14N5) A/mallard/Gurjev/263/1982, and an A(H15N9) (A/wedge-tailed shearwater/Western Australia/2576/1979 were determined by X-ray crystallography at 2.2Å, 2.3Å, 2.8Å, 3.0Å and 2.5Å resolution, respectively. The interactions between these viruses and host receptors were studied utilizing glycan-binding analyses with their recombinant HA. The data show that all avian HAs retain their strict binding preference to avian receptors, whereas swine H4 has a weak human receptor binding. The molecular characterization and structural analyses of the HA from these zoonotic influenza viruses not only provide a deeper appreciation and understanding of the structure of all HA subtypes, but also re-iterate why continuous global surveillance is needed.

BACKGROUND: The development of serologic assays that can rapidly assess human exposure to novel influenza viruses remains a public health need. Previously, we developed an 11-plex magnetic fluorescence microsphere immunoassay (MAGPIX) by using globular head domain recombinant hemagglutinins (rHAs) with serum adsorption using two ectodomain rHAs. METHODS: We compared sera collected from two cohorts with novel influenza exposures: animal shelter staff during an A(H7N2) outbreak in New York City in 2016-2017 (n = 119 single sera) and poultry workers from a live bird market in Bangladesh in 2012-2014 (n = 29 pairs). Sera were analyzed by microneutralization (MN) assay and a 20-plex MAGPIX assay with rHAs from 19 influenza strains (11 subtypes) combined with serum adsorption using 8 rHAs from A(H1N1) and A(H3N2) viruses. Antibody responses were analyzed to determine the novel influenza virus exposure. RESULTS: Among persons with novel influenza virus exposures, the median fluorescence intensity (MFI) against the novel rHA from exposed influenza virus had the highest correlation with MN titers to the same viruses and could be confirmed by removal of cross-reactivity from seasonal H1/H3 rHAs following serum adsorption. Interestingly, in persons with exposures to novel influenza viruses, age and MFIs against exposed novel HA were negatively correlated, whereas in persons without exposure to novel influenza viruses, age and MFI against novel HAs were positively correlated. CONCLUSIONS: This 20-plex high-throughput assay with serum adsorption will be a useful tool to detect novel influenza virus infections during influenza outbreak investigations and surveillance, especially when well-paired serum samples are not available.

BACKGROUND: Human immunodeficiency virus (HIV)-infected persons are at a higher risk of severe influenza. Although we have shown that a standard-dose intradermal influenza vaccine versus a standard-dose intramuscular influenza vaccine does not result in differences in hemagglutination-inhibition titers in this population, a comprehensive examination of cell-mediated immune responses remains lacking. METHODS: Serological, antigen-specific B-cell, and interleukin 2-, interferon gamma-, and tumor necrosis factor alpha-secreting T-cell responses were assessed in 79 HIV-infected men and 79 HIV-uninfected men. RESULTS: The route of vaccination did not affect the immunoglobulin A and immunoglobulin G (IgG) plasmablast or memory B-cell response, although these were severely impaired in the group with a CD4+ T-cell count of <200 cells/muL. The frequencies of IgG memory B cells measured on day 28 after vaccination were highest in the HIV-uninfected group, followed by the group with a CD4+ T-cell count of >/=200 cells/muL and the group with a CD4+ T-cell count of <200 cells/muL. The route of vaccination did not affect the CD4+ or CD8+ T-cell responses measured at various times after vaccination. CONCLUSIONS: The route of vaccination had no effect on antibody responses, antibody avidity, T-cell responses, or B-cell responses in HIV-infected or HIV-uninfected subjects. With the serological and cellular immune responses to influenza vaccination being impaired in HIV-infected individuals with a CD4+ T-cell count of <200 cells/muL, passive immunization strategies need to be explored to protect this population. CLINICAL TRIALS REGISTRATION: NCT01538940.

A(H3N2) virus predominated recent influenza seasons, which has resulted in the rigorous investigation of haemagglutinin, but whether neuraminidase (NA) has undergone antigenic change and contributed to the predominance of A(H3N2) virus is unknown. Here, we show that the NA of the circulating A(H3N2) viruses has experienced significant antigenic drift since 2016 compared with the A/Hong Kong/4801/2014 vaccine strain. This antigenic drift was mainly caused by amino acid mutations at NA residues 245, 247 (S245N/S247T; introducing an N-linked glycosylation site at residue 245) and 468. As a result, the binding of the NA of A(H3N2) virus by some human monoclonal antibodies, including those that have broad reactivity to the NA of the 1957 A(H2N2) and 1968 A(H3N2) reference pandemic viruses as well as contemporary A(H3N2) strains, was reduced or abolished. This antigenic drift also reduced NA-antibody-based protection against in vivo virus challenge. X-ray crystallography showed that the glycosylation site at residue 245 is within a conserved epitope that overlaps the NA active site, explaining why it impacts antibody binding. Our findings suggest that NA antigenic drift impacts protection against influenza virus infection, thus highlighting the importance of including NA antigenicity for consideration in the optimization of influenza vaccines.

The recent reduction of live attenuated influenza vaccine (LAIV) effectiveness in multivalent formulations was particularly associated with the A(H1N1)pdm09 component. In the 2017 the WHO vaccine composition committee changed its recommendations for the A(H1N1)pdm09 component to include an A/Michigan/45/2015-like virus. We evaluated effectiveness and quality of newly developed and previous A(H1N1)pdm09 LAIV reassortants through assessment of their thermal and pH stability, receptor binding specificity and replication fitness in primary human airway epithelial cells of nasal origin (hAECN). Our analysis showed that LAIV expressed hemagglutinin (HA) and neuraminidase (NA) from an A/Michigan/45/2015-like strain A/New York/61/2015 (A/New York/61/2015-CDC-LV16A, NY-LV16A), exhibit higher thermal and pH stability compared to the previous vaccine candidates expressing HA and NA from A/California/07/2009 and A/Bolivia/559/2013 (A17/Cal09 and A17/Bol13). Reassortants A/South Africa/3626/2013-CDC-LV14A (SA-LV14A) and NY-LV16A showed preferential binding to alpha2,6 sialic acid (SA) receptors and replicated at higher titers and more extensively in hAECN compared to A17/Cal09 and A17/Bol13, which had an alpha2,3 SA receptor binding preference. Our data analysis supports selection of A/New York/61/2015-CDC-LV16A for LAIV formulation and the introduction of new assays for LAIV characterization.

Objectives: The Centers for Disease Control and Prevention launched the Temporary Epidemiology Field Assignee (TEFA) Program to help state and local jurisdictions respond to the risk of Ebola virus importation during the 2014-2016 Ebola Outbreak in West Africa. We describe steps taken to launch the 2-year program, its outcomes and lessons learned. Methods: State and local health departments submitted proposals for a TEFA to strengthen local capacity in four key public health preparedness areas: 1) epidemiology and surveillance, 2) health systems preparedness, 3) health communications, and 4) incident management. TEFAs and jurisdictions were selected through a competitive process. Descriptions of TEFA activities in their quarterly reports were reviewed to select illustrative examples for each preparedness area. Results: Eleven TEFAs began in the fall of 2015, assigned to 7 states, 2 cities, 1 county and the District of Columbia. TEFAs strengthened epidemiologic capacity, investigating routine and major outbreaks in addition to implementing event-based and syndromic surveillance systems. They supported improvements in health communications, strengthened healthcare coalitions, and enhanced collaboration between local epidemiology and emergency preparedness units. Several TEFAs deployed to United States territories for the 2016 Zika Outbreak response. Conclusion: TEFAs made important contributions to their jurisdictions' preparedness. We believe the TEFA model can be a significant component of a national strategy for surging state and local capacity in future high-consequence events.

Cold adapted influenza virus A/Leningrad/134/17/57 (H2N2) is a reliable master donor virus (Len/17-MDV) for preparing live attenuated influenza vaccines (LAIV). LAIVs are 6:2 reasortants that contain 6 segments of Len/17-MDV and the hemagglutinin (HA) and neuraminidase (NA) of contemporary circulating influenza A viruses. The problem with the classical reassortment procedure used to generate LAIVs is that there is limited selection pressure against NA of the Len/17-MDV resulting in 7:1 reassortants with desired HA only, which are not suitable LAIVs. The monoclonal antibodies (mAb) directed against the N2 of Len/17-MDV were generated. 10C4-8E7 mAb inhibits cell-to-cell spread of viruses containing the Len/17-MDV N2, but not viruses with the related N2 from contemporary H3N2 viruses. 10C4-8E7 antibody specifically inhibited the Len/17-MDV replication in vitro and in ovo but didn't inhibit replication of H3N2 or H1N1pdm09 reassortants. Our data demonstrate that addition of 10C4-8E7 in the classical reassortment improves efficiency of LAIV production.

Many broadly neutralizing antibodies (bnAbs) bind to conserved areas of the hemagglutinin (HA) stalk region and can inhibit the low pH induced HA conformational changes necessary for viral membrane fusion activity. We developed and evaluated a high-throughput virus-free and cell-free ELISA based low pH induced HA Conformational Change Inhibition Antibody Detection Assay (HCCIA) and a complementary proteinase susceptibility assay. Human serum samples (n = 150) were tested by HCCIA using H3 recombinant HA. Optical density (OD) ratios of mAb HC31 at pH 4.8 to pH 7.0 ranged from 0.87 to 0.09. Our results demonstrated that low pH induced HA conformational change inhibition antibodies (CCI) neutralized multiple H3 strains after removal of head-binding antibodies. The results suggest that HCCIA can be utilized to detect and characterize CCI in sera, that are potentially broadly neutralizing, and serves as a useful tool for evaluating universal vaccine candidates targeting the HA stalk.

The avian influenza A(H7N9) virus continues to cause human infections in China and is a major ongoing public health concern. Five epidemic waves of A(H7N9) infection have occurred since 2013, and the recent fifth epidemic wave saw the emergence of two distinct lineages with elevated numbers of human infection cases and broader geographic distribution of viral diseases compared to the first four epidemic waves. Moreover, highly pathogenic avian influenza (HPAI) A(H7N9) viruses were also isolated during the fifth epidemic wave. Here, we present a detailed structural and biochemical analysis of the surface hemagglutinin (HA) antigen from viruses isolated during this recent epidemic wave. Results highlight that when compared to the 2013 virus HAs, the fifth wave virus HAs remained a weak binder to human glycan receptor analogs. We also studied three mutations, V177K-K184T-G219S, that were recently reported to switch a 2013 A(H7N9)HA to human-type receptor specificity. Our results indicate that these mutations could also switch the H7 HA receptor preference to a predominantly human binding specificity for both fifth wave H7 HAs analyzed in this study.IMPORTANCE The A(H7N9) viruses circulating in China are of great public health concern. Herein, we report a molecular and structural study of the major surface proteins from several recent A(H7N9) influenza viruses. Our results improve the understanding of these evolving viruses and provide important information on their receptor preference that is central to ongoing pandemic risk assessment.

PURPOSE: Sexual violence (SV) perpetration on college campuses is a serious and prevalent public health issue in the U.S. In response, incoming male freshmen are mandated to receive SV prevention programming. To provide a more effective response, however, we need to understand the SV behaviors of male freshmen before they arrive on campus and the associated factors that contribute to risk and that afford protection, areas that have received limited attention. METHODS: Male freshmen (N = 1,133) across 30 selected 4-year colleges and universities throughout the state of Georgia were recruited for a longitudinal study on SV perpetration. Levels of precollege SV as well as a range of covariates were assessed at baseline. Self-reported SV perpetrators were compared with nonperpetrators on demographic and hypothesized covariates deemed either risk or protective; then risk and protective models were analyzed using binary logistic regression. RESULTS: Weighted analyses revealed that 19.3% self-reported perpetrating SV before college. Before starting college, young men who reported more sexual media consumption, heavy episodic drinking, hypermasculine beliefs, and peers who endorsed SV were more likely to have a history of SV perpetration at college matriculation. Alternatively, men with more knowledge of effective sexual consent and stronger family functioning were less likely to arrive to college with an SV perpetration history. CONCLUSIONS: A significant proportion of incoming male freshmen have perpetrated SV previously. Colleges and universities need to assess incoming freshmen for risk behaviors and negative beliefs and to offer both primary and secondary preventions to more effectively reduce further perpetration.

An outbreak of influenza A(H7N2) virus in cats in a shelter in New York, NY, USA, resulted in zoonotic transmission. Virus isolated from the infected human was closely related to virus isolated from a cat; both were related to low pathogenicity avian influenza A(H7N2) viruses detected in the United States during the early 2000s.

Background: A single subtype of canine influenza virus (CIV), A(H3N8), was circulating in the United States until a new subtype, A(H3N2), was detected in Illinois in spring 2015. Since then, this CIV has caused thousands of infections in dogs in multiple states. Methods: In this study, genetic and antigenic properties of the new CIV were evaluated. In addition, structural and glycan array binding features of the recombinant hemagglutinin were determined. Replication kinetics in human airway cells and pathogenesis and transmissibility in animal models were also assessed. Results: A(H3N2) CIVs maintained molecular and antigenic features related to low pathogenicity avian influenza A(H3N2) viruses and were distinct from A(H3N8) CIVs. The structural and glycan array binding profile confirmed these findings and revealed avian-like receptor-binding specificity. While replication kinetics in human airway epithelial cells was on par with that of seasonal influenza viruses, mild-to-moderate disease was observed in infected mice and ferrets, and the virus was inefficiently transmitted among cohoused ferrets. Conclusions: Further adaptation is needed for A(H3N2) CIVs to present a likely threat to humans. However, the potential for coinfection of dogs and possible reassortment of human and other animal influenza A viruses presents an ongoing risk to public health.

The emergence of A(H7N9) virus strains with resistance to neuraminidase (NA) inhibitors highlights a critical need to discover new countermeasures for treatment of A(H7N9) virus-infected patients. We previously described an anti-NA mAb (3c10-3) that has prophylactic and therapeutic efficacy in mice lethally challenged with A(H7N9) virus when delivered intraperitoneally (i.p.). Here we show that intrananasal (i.n.) administration of 3c10-3 protects 100% of mice from mortality when treated 24h post-challenge and further characterize the protective efficacy of 3c10-3 using a nonlethal A(H7N9) challenge model. Administration of 3c10-3 i.p. 24h prior to challenge resulted in a significant decrease in viral lung titers and deep sequencing analysis indicated that treatment did not consistently select for viral variants in NA. Furthermore, prophylactic administration of 3c10-3 did not inhibit the development of protective immunity to subsequent homologous virus re-challenge. Taken together, 3c10-3 highlights the potential use of anti-NA mAb to mitigate influenza virus infection.

On September 8, 2015, the District of Columbia Department of Health (DCDOH) received a call from a person who reported experiencing gastrointestinal illness after eating at a District of Columbia (DC) restaurant with multiple locations throughout the United States (restaurant A). Later the same day, a local emergency department notified DCDOH to report four persons with gastrointestinal illness, all of whom had eaten at restaurant A during August 30-September 5. Two patients had laboratory-confirmed Salmonella group D by stool culture. On the evening of September 9, a local newspaper article highlighted a possible outbreak associated with restaurant A. Investigation of the outbreak by DCDOH identified 159 patrons who were residents of 11 states and DC with gastrointestinal illness after eating at restaurant A during July 1-September 10. A case-control study was conducted, which suggested truffle oil-containing food items as a possible source of Salmonella enterica serotype Enteritidis infection. Although several violations were noted during the restaurant inspections, the environmental, laboratory, and traceback investigations did not confirm the contamination source. Because of concern about the outbreak, the restaurant's license was suspended during September 10-15. The collaboration and cooperation of the public, media, health care providers, and local, state, and federal public health officials facilitated recognition of this outbreak involving a pathogen commonly implicated in foodborne illness.

Zoonotic A(H7N9) avian influenza viruses emerged in China in 2013 and continue to be a threat to human public health, having infected over 800 individuals with a mortality rate approaching 40%. Treatment options for people infected with A(H7N9) include the use of neuraminidase (NA) inhibitors. However, like other influenza viruses, A(H7N9) can become resistant to these drugs. The use of monoclonal antibodies is a rapidly developing strategy for controlling influenza virus infection. Here we generated a murine monoclonal antibody (3c10-3) directed against the NA of A(H7N9) and show that prophylactic systemic administration of 3c10-3 fully protected mice from lethal challenge with wild-type A/Anhui/1/2013 (H7N9). Further, post-infection treatment with a single systemic dose of 3c10-3 at either 24, 48 or 72 h post A(H7N9) challenge resulted in both dose- and time-dependent protection of up to 100% of mice, demonstrating therapeutic potential for 3c10-3. Epitope mapping revealed that 3c10-3 binds near the enzyme active site of NA, and functional characterization showed that 3c10-3 inhibits the enzyme activity of NA and restricts the cell-to-cell spread of the virus in cultured cells. Affinity analysis also revealed that 3c10-3 binds equally well to recombinant NA of wild-type A/Anhui/1/2013 and to a variant NA carrying a R289K mutation known to infer NAI resistance. These results suggest that 3c10-3 has the potential to be used as a therapeutic to treat A(H7N9) infections either as an alternative to, or in combination with, current NA antiviral inhibitors.

During 2014, a subclade 2.3.4.4 HPAI A(H5N8) virus caused poultry outbreaks around the world. In late 2014/early 2015 the virus was detected in wild birds in Canada and the U.S. and these viruses also gave rise to reassortant progeny, composed of viral RNA segments (vRNAs) from both Eurasian and North America lineages. In particular, viruses were found with N1, N2 and N8 neuraminidase vRNAs, and are collectively referred to as H5Nx viruses. In the U. S., more than 48 million domestic birds have been affected. Here, we present a detailed structural and biochemical analysis of the surface antigens from H5N1, H5N2 and H5N8 in addition to a recent human H5N6 virus. Our results with recombinant hemagglutinin reveal that these viruses have a strict avian receptor binding preference, while recombinantly expressed neuraminidases are sensitive to FDA approved and investigational antivirals. Although H5Nx viruses currently pose a low risk to humans, it is important to maintain surveillance of these circulating viruses, and to continually assess future changes that may increase their pandemic potential. IMPORTANCE: The H5Nx viruses emerging in North America, Europe, and Asia are of great public health concern. Herein, we report a molecular and structural study of the major surface proteins from several H5Nx influenza viruses. Our results improve the understanding of these new viruses and provide important information on their receptor preference and susceptibility to antivirals, which is central to pandemic risk assessment.

During 2013, three new avian influenza A virus subtypes, A(H7N9), A(H6N1), and A(H10N8), resulted in human infections. While the A(H7N9) virus resulted in a significant epidemic in China across 19 provinces and municipalities, both A(H6N1) and A(H10N8) viruses resulted in only a few human infections. This study focuses on the major surface glycoprotein hemagglutinins from both of these novel human viruses. The detailed structural and glycan microarray analyses presented here highlight the idea that both A(H6N1) and A(H10N8) virus hemagglutinins retain a strong avian receptor binding preference and thus currently pose a low risk for sustained human infections. IMPORTANCE: Human infections with zoonotic influenza virus subtypes continue to be a great public health concern. We report detailed structural analysis and glycan microarray data for recombinant hemagglutinins from A(H6N1) and A(H10N8) viruses, isolated from human infections in 2013, and compare them with hemagglutinins of avian origin. This is the first structural report of an H6 hemagglutinin, and our results should further the understanding of these viruses and provide useful information to aid in the continuous surveillance of these zoonotic influenza viruses.

After their disappearance from the human population in 1968, influenza H2 viruses have continued to circulate in the natural avian reservoir. The isolation of this virus subtype from multiple bird species as well as swine highlights the need to better understand the potential of these viruses to spread and cause disease in humans. Here we analyzed the virulence, transmissibility and receptor-binding preference of two avian influenza H2 viruses (H2N2 and H2N3) and compared them to a swine H2N3 (A/swine/Missouri/2124514/2006 [swMO]), and a human H2N2 (A/England/10/1967 [Eng/67]) virus using the ferret model as a mammalian host. Both avian H2 viruses possessed the capacity to spread efficiently between cohoused ferrets, and the swine (swMO) and human (Eng/67) viruses transmitted to naive ferrets by respiratory droplets. Further characterization of the swMO hemagglutinin (HA) by x-ray crystallography and glycan microarray array identified receptor-specific adaptive mutations. As influenza virus quasispecies dynamics during transmission have not been well characterized, we sequenced nasal washes collected during transmission studies to better understand experimental adaptation of H2 HA. The avian H2 viruses isolated from ferret nasal washes contained mutations in the HA1, including a Gln226Leu substitution, which is a mutation associated with alpha2,6 sialic acid (human-like) binding preference. These results suggest that the molecular structure of HA in viruses of the H2 subtype continue to have the potential to adapt to a mammalian host and become transmissible, after acquiring additional genetic markers.

A(H1N1)pdm09 influenza A viruses predominated in the 2013-2014 USA influenza season, and although most of these viruses remain sensitive to Food and Drug Administration-approved neuraminidase (NA) inhibitors, alternative therapies are needed. Here we show that monoclonal antibody CD6, selected for binding to the NA of the prototypic A(H1N1)pdm09 virus, A/California/07/2009, protects mice against lethal virus challenge. The crystal structure of NA in complex with CD6 Fab reveals a unique epitope, where the heavy-chain complementarity determining regions (HCDRs) 1 and 2 bind one NA monomer, the light-chain CDR2 binds the neighbouring monomer, whereas HCDR3 interacts with both monomers. This 30-amino-acid epitope spans the lateral face of an NA dimer and is conserved among circulating A(H1N1)pdm09 viruses. These results suggest that the large, lateral CD6 epitope may be an effective target of antibodies selected for development as therapeutic agents against circulating H1N1 influenza viruses.