History of Swine influenza.

Swine influenza is a continual problem for the Swine industry and can pose a public health threat as evidenced by the 2009 H1N1 influenza virus pandemic. Given its importance, it is not surprising to find papers describing the disease from the early 20th century. In this chapter, we discuss the history of Swine influenza, the important role swine influenza virus has played in our understanding of influenza virus pathogenesis and virology, and its impact on public health worldwide.

Students' perception of case-based continuous assessment and multiple-choice assessment in a small animal surgery course for veterinary medical students.

OBJECTIVE: To determine the relationship between students' perceptions of 2 assessment methods and academic performance. STUDY DESIGN: A 2-year prospective survey study in a 4-year Doctor of Veterinary Medicine (DVM) curriculum. SAMPLE POPULATION: Year 3 DVM students (n = 44). METHODS: An assessment of learning gain questionnaire was used to investigate students' perceptions regarding multiple-choice examination (MCE) versus take-home case-based continuous assessment (CA) in a 3rd year small animal surgery lecture course. Academic performance and student assessment of learning gain in the 2 course components were compared. Relationships between student perceptions and academic performance were examined. A follow-up survey was conducted during clinical rotations in 4th year to determine change in student perceptions over time. RESULTS: Academic performance in 3rd year was significantly enhanced by use of CA, particularly for students with weaker grades. Academic performance in 4th year clinical rotations was not closely related to 3rd year performance. Many students preferred an instructional approach with provision of comprehensive notes and assessment with multiple-choice questions based on the notes. However, students recognized that feedback on work submitted for CA grading significantly facilitated learning. Student assessment of learning gain was correlated with academic performance in the 3rd year course component examined by CA, but not the component assessed using MCE. CONCLUSION: Our data suggest that perceptions of learning gain, academic accomplishment, and clinical performance in 4th year are weakly correlated. Teachers should better explain to veterinary students that learning to be a clinician is more than replication of knowledge.

Glycan analysis and influenza A virus infection of primary swine respiratory epithelial cells: the importance of NeuAc{alpha}2-6 glycans.

To better understand influenza virus infection of pigs, we examined primary swine respiratory epithelial cells (SRECs, the primary target cells of influenza viruses in vivo), as a model system. Glycomic profiling of SRECs by mass spectrometry revealed a diverse range of glycans terminating in sialic acid or GalαGal. In terms of sialylation, α2-6 linkage was more abundant than α2-3, and NeuAc was more abundant than NeuGc. Virus binding and infection experiments were conducted to determine functionally important glycans for influenza virus infection, with a focus on recently emerged swine viruses. Infection of SRECs with swine and human viruses resulted in different infectivity levels. Glycan microarray analysis with a high infectivity "triple reassortant" virus ((A/Swine/MN/593/99 (H3N2)) that spread widely throughout the North American swine population and a lower infectivity human virus isolated from a single pig (A/Swine/ONT/00130/97 (H3N2)) showed that both viruses bound exclusively to glycans containing NeuAcα2-6, with strong binding to sialylated polylactosamine and sialylated N-glycans. Treatment with mannosamine precursors of sialic acid (to alter NeuAc/NeuGc abundances) and linkage-specific sialidases prior to infection indicated that the influenza viruses tested preferentially utilize NeuAcα2-6-sialylated glycans to infect SRECs. Our data indicate that NeuAcα2-6-terminated polylactosamine and sialylated N-glycans are important determinants for influenza viruses to infect SRECs. As NeuAcα2-6 polylactosamine glycans play major roles in human virus infection, the importance of these receptor components in virus infection of swine cells has implications for transmission of viruses between humans and pigs and for pigs as possible adaptation hosts of novel human influenza viruses.

Serologic cross-reactivity with pandemic (H1N1) 2009 virus in pigs, Europe.

We tested serum samples from pigs infected or vaccinated with European swine influenza viruses (SIVs) in hemagglutination-inhibition assays against pandemic (H1N1) 2009 virus and related North American SIVs. We found more serologic cross-reaction than expected. Data suggest pigs in Europe may have partial immunity to pandemic (H1N1) 2009 virus.

Amino acid 226 in the hemagglutinin of H4N6 influenza virus determines binding affinity for alpha2,6-linked sialic acid and infectivity levels in primary swine and human respiratory epithelial cells.

Avian lineage H4N6 influenza viruses previously isolated from pigs differ at hemagglutinin amino acids 226 and 228 from H4 subtype viruses isolated from birds. Using a parental H4N6 swine isolate and hemagglutinin mutant viruses (at residues 226 and/or 228), we determined that viruses which contain L226 had a higher affinity for sialic acid alpha2,6 galactose (SAalpha2,6Gal) and a higher infectivity level for primary swine and human respiratory epithelial cells, whereas viruses which contain Q226 had lower SAalpha2,6Gal affinity and lower infectivity levels for both types of cells. Using specific neuraminidases, we found that irrespective of their relative binding preferences, all of the influenza viruses examined utilized SAalpha2,6Gal to infect swine and human cells.

Gender Analysis for One Health: Theoretical Perspectives and Recommendations for Practice.

One health emphasizes the interdependent health of humans, animals, and their shared environments and shows promise as an integrated, equitable transdisciplinary approach to important ecohealth issues. Notably, research or programming explicitly examining the intersection of gender and one health is limited, although females represent half of the human population and play important roles in human and animal health around the world. Recognizing these gaps, scholars from the University of Wisconsin-Madison in collaboration with United States Department of Agriculture convened a consultative workshop, "Women and One Health," in 2016. This paper outlines the workshop methods and highlights outcomes toward shared terminology and integration of frameworks from one health, gender analysis, and women in agriculture. Further, recommendations for education, policy, and service delivery at the intersection of women's empowerment and one health are offered as important efforts toward the dual goals of gender equality and sustainable health of humans, animals, and their shared ecosystems.

Identification of amino acids in the HA of H3 influenza viruses that determine infectivity levels in primary swine respiratory epithelial cells.

In the late 1990s, triple reassortant H3N2 influenza A viruses emerged and spread widely within the swine population of the United States. We have shown previously that an isolate representative of this lineage of viruses, A/Swine/Minnesota/593/99 (Sw/MN), has higher infectivity and accelerated replication kinetics in pigs, compared to a human-lineage H3N2 virus isolated from a pig during the same time period, A/Swine/Ontario/00130/97 (Sw/ONT [Landolt, G.A., Karasin, A.I., Phillips, L., Olsen, C.W., 2003. Comparison of the pathogenesis of two genetically different H3N2 influenza A viruses in pigs. J. Clin. Microbiol. 41, 1936-1941]). Additional in vivo experiments using reverse genetics-generated reassortant viruses demonstrated that these phenotypes are dependent upon the HA and/or NA genes (Landolt, G.A., Karasin, A.I., Schutten, M.M., Olsen, C.W., 2006. Restricted infectivity of a human-lineage H3N2 influenza A virus in pigs is hemagglutinin and neuraminidase gene dependent. J. Clin. Microbiol. 44, 297-301). To further study the infectivity of influenza viruses for pigs, we developed a primary swine respiratory epithelial cell (SREC) culture model. In SRECs, Sw/MN infects a significantly higher number of cells compared to Sw/ONT. Using reverse genetics-generated Sw/MN x Sw/ONT reassortant viruses we demonstrate that the infectivity phenotypes of these viruses in SRECs are strongly dependent upon the HA gene. Using chimeras and point directed mutations within the HA genes, we have identified amino acids that, either alone or in combination with other amino acids, impact infectivity. In particular, amino acid 138 is the dominant factor in determining infectivity levels in SRECs.

Creating a center for global health at the University of Wisconsin-Madison.

Globalization, migration, and widespread health disparities call for interdisciplinary approaches to improve health care at home and abroad. Health professions students are pursuing study abroad in increasing numbers, and universities are responding with programs to address these needs. The University of Wisconsin (UW)-Madison schools of medicine and public health, nursing, pharmacy, veterinary medicine, and the division of international studies have created an interdisciplinary center for global health (CGH). The CGH provides health professions and graduate students with courses, field experiences, and a new Certificate in Global Health. Educational programs have catalyzed a network of enthusiastic UW global health scholars. Partnerships with colleagues in less economically developed countries provide the foundation for education, research, and service programs. Participants have collaborated to improve the education of health professionals and nutrition in Uganda; explore the interplay between culture, community development, and health in Ecuador; improve animal health and address domestic violence in Mexico; and examine successful public health efforts in Thailand. These programs supply students with opportunities to understand the complex determinants of health and structure of health systems, develop adaptability and cross-cultural communication skills, experience learning and working in interdisciplinary teams, and promote equity and reduce health disparities at home and abroad. Based on the principles of equity, sustainability, and reciprocity, the CGH provides a strong foundation to address global health challenges through networking and collaboration among students, staff, and faculty within the UW and beyond.

The dual DVM/MPH degree at the University of Wisconsin--Madison: a uniquely interdisciplinary collaboration.

The University of Wisconsin-Madison (UWM) launched a new Master of Public Health (MPH) degree program in 2005. This 42-credit MPH degree consists of 18 core and 14 elective course credits, two seminar credits, and eight field project/culminating experience credits. Unique strengths of the program include its strongly interdisciplinary philosophy, encompassing both health science (human medicine, veterinary medicine, pharmacy, nursing) and social science units on campus, and its emphasis on service learning through instructional and field project ties to the public-health community of the state and beyond. To date, the program has admitted 87 students, including full-time students as well as part-time students who continue to work in the health care and/or public-health sectors. The program is currently proceeding with the process for accreditation through the Council for Education in Public Health. In 2007, a formal dual DVM/MPH program was approved to allow students to integrate DVM and MPH training and complete both degrees in a total of five years. Nine MPH students over the first three years of admissions have been individuals affiliated with veterinary medicine (five DVM students and four post-graduate veterinarians).

Cases of swine influenza in humans: a review of the literature.

As the threat of a pandemic looms, improvement in our understanding of interspecies transmission of influenza is necessary. Using the search terms "swine," "influenza," and "human," we searched the PubMed database in April 2006 to identify publications describing symptomatic infections of humans with influenza viruses of swine origin. From these reports, we extracted data regarding demographic characteristics, epidemiological investigations, and laboratory results. We found 50 cases of apparent zoonotic swine influenza virus infection, 37 of which involved civilians and 13 of which involved military personnel, with a case-fatality rate of 14% (7 of 50 persons). Most civilian subjects (61%) reported exposure to swine. Although sporadic clinical cases of swine influenza occur in humans, the true incidence of zoonotic swine influenza virus infection is unknown. Because prior studies have shown that persons who work with swine are at increased risk of zoonotic influenza virus infection, it is prudent to include them in pandemic planning efforts.

Up to new tricks - a review of cross-species transmission of influenza A viruses.

Influenza is a highly contagious disease that has burdened both humans and animals since ancient times. In humans, the most dramatic consequences of influenza are associated with periodically occurring pandemics. Pandemics require the emergence of an antigenically novel virus to which the majority of the population lacks protective immunity. Historically, influenza A viruses from animals have contributed to the generation of human pandemic viruses and they may do so again in the future. It is, therefore, critical to understand the epidemiological and molecular mechanisms that allow influenza A viruses to cross species barriers. This review summarizes the current knowledge of influenza ecology, and the viral factors that are thought to determine influenza A virus species specificity.

Are swine workers in the United States at increased risk of infection with zoonotic influenza virus?

BACKGROUND: Pandemic influenza strains originate in nonhuman species. Pigs have an important role in interspecies transmission of the virus. We examined multiple swine-exposed human populations in the nation's number 1 swine-producing state for evidence of previous swine influenza virus infection. METHODS: We performed controlled, cross-sectional seroprevalence studies among 111 farmers, 97 meat processing workers, 65 veterinarians, and 79 control subjects using serum samples collected during the period of 2002-2004. Serum samples were tested using a hemagglutination inhibition assay against the following 6 influenza A virus isolates collected recently from pigs and humans: A/Swine/WI/238/97 (H1N1), A/Swine/WI/R33F/01 (H1N2), A/Swine/Minnesota/593/99 (H3N2), A/New Caledonia/20/99 (H1N1), A/Panama/2007/99 (H3N2), and A/Nanchang/933/95 (H3N2). RESULTS: Using multivariable proportional odds modeling, all 3 exposed study groups demonstrated markedly elevated titers against the H1N1 and H1N2 swine influenza virus isolates, compared with control subjects. Farmers had the strongest indication of exposure to swine H1N1 virus infection (odds ratio [OR], 35.3; 95% confidence interval [CI], 7.7-161.8), followed by veterinarians (OR, 17.8; 95% CI, 3.8-82.7), and meat processing workers (OR, 6.5; 95% CI, 1.4-29.5). Similarly, farmers had the highest odds for exposure to swine H1N2 virus (OR, 13.8; 95% CI, 5.4-35.4), followed by veterinarians (OR, 9.5; 95% CI, 3.6-24.6) and meat processing workers (OR, 2.7; 95% CI, 1.1-6.7). CONCLUSIONS: Occupational exposure to pigs greatly increases workers' risk of swine influenza virus infection. Swine workers should be included in pandemic surveillance and in antiviral and immunization strategies.

Receptor-binding properties of swine influenza viruses isolated and propagated in MDCK cells.

To study the receptor specificities of H1 and H3 influenza viruses isolated recently from pigs, we employed the analogues of natural receptors, namely sialyloligosaccharides conjugated with polyacrylamide in biotinylated and label free forms. All Madin-Darby canine kidney (MDCK) cell-propagated viruses with human H3 or classical swine H1 hemagglutinins bound only to Neu5Acalpha2-6Galbeta1-bearing polymers, and not to Neu5Acalpha2-3Galbeta1-bearing polymers. This receptor-binding pattern is typical for human influenza viruses and it differs from the previously described receptor-binding specificity of egg-adapted swine influenza viruses. Swine virus isolates with avian-like H1 and H3 hemagglutinins displayed distinct receptor specificity by binding to both Neu5Acalpha2-6Gal- and Neu5Acalpha2-3Gal-containing receptors. These viruses, as well as egg-adapted swine and turkey viruses with a classical swine HA, differed from the related duck viruses by increased affinity to sulfated sialyloligosaccaride, Su-SiaLe(x). Except for avian-like H3 viruses, none of the studied swine viruses bound to Neu5Gc-containing sialoglycopolymers, suggesting that binding to these sialic acid species abundantly expressed in pigs may not be essential for virus replication in this host.

Use of real-time reverse transcriptase polymerase chain reaction assay and cell culture methods for detection of swine influenza A viruses.

OBJECTIVE: To evaluate sensitivity and specificity of a real-time reverse transcriptase polymerase chain reaction (RT-PCR) assay performed on pooled nasal swab specimens, compared with virus isolation performed on individual nasal swab specimens by use of 2 cell culture lines for detection of swine influenza A viruses. SAMPLE POPULATION: 900 nasal swab specimens obtained from pigs at an abattoir and 62 nasal swab specimens submitted for diagnostic testing. PROCEDURES: Primers were chosen to amplify a conserved portion of the influenza virus matrix gene. Assay sensitivity was initially determined by testing serial dilutions of various subtypes of swine influenza viruses. Sensitivity and specificity were confirmed by use of nasal swab specimens with or without addition of known concentrations of influenza virus and further validated by testing nasal swab specimens obtained through an abattoir surveillance program or submitted for diagnostic testing. Aliquots of specimens were pooled in sets of 10, and results of real-time RT-PCR assays were compared with results of virus isolation of individual specimens in Madin Darby canine kidney (MDCK) and mink lung (Mv1Lu) cells. RESULTS: Real-time RT-PCR assay was highly specific (100%) and sensitive (88% to 100%). Among the 16 viruses isolated, 3 grew only in Mv1Lu cells and 3 grew only in MDCK cells. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that real-time RT-PCR assay is a fast and accurate test for screening numerous nasal swab specimens for swine influenza virus. Some viruses were isolated in only MDCK or Mv1Lu cells, indicating that use of >1 cell line may be required to isolate a broad range of influenza A viruses.

The emergence of novel swine influenza viruses in North America.

Since 1997, novel viruses of three different subtypes and five different genotypes have emerged as agents of influenza among pigs in North America. The appearance of these viruses is remarkable because there were no substantial changes in the overall epidemiology of swine influenza in the United States and Canada for over 60 years prior to this time. Viruses of the classical H1N1 lineage were virtually the exclusive cause of swine influenza from the time of their initial isolation in 1930 through 1998. Antigenic drift variants of these H1N1 viruses were isolated in 1991-1998, but a much more dramatic antigenic shift occurred with the emergence of H3N2 viruses in 1997-1998. In particular, H3N2 viruses with genes derived from human, swine and avian viruses have become a major cause of swine influenza in North America. In addition, H1N2 viruses that resulted from reassortment between the triple reassortant H3N2 viruses and classical H1N1 swine viruses have been isolated subsequently from pigs in at least six states. Finally, avian H4N6 viruses crossed the species barrier to infect pigs in Canada in 1999. Fortunately, these H4N6 viruses have not been isolated beyond their initial farm of origin. If these viruses spread more widely, they will represent another antigenic shift for our swine population, and could pose a threat to the world's human population. Research on these novel viruses may offer important clues to the genetic basis for interspecies transmission of influenza viruses.

Characterization of a swine-like reassortant H1N2 influenza virus isolated from a wild duck in the United States.

An H1N2 influenza virus (A/Duck/North Carolina/91347/01) (Dk/NC) was isolated from a wild duck in the United States in 2001. Genetic analyses showed that this duck virus has the same human/classical swine/avian reassortant genotype as the H1N2 viruses that have been isolated from pigs and turkeys in the US since 1999. Phylogenetic analyses of each gene segment further confirmed that the Dk/NC virus is closely related to the domestic animal H1N2 isolates. In particular, Dk/NC is most closely related to a swine H1N2 virus also isolated in North Carolina. These two viruses and a phylogenetically-defined subset of additional swine H1N2 viruses share a common mutation in the Sb antigenic site on the hemagglutinin protein. The recovery of Dk/NC from a wild bird raises concerns for further widespread distribution of these H1N2 viruses via waterfowl migration.

Effects of DNA dose, route of vaccination, and coadministration of porcine interleukin-6 DNA on results of DNA vaccination against influenza virus infection in pigs.

OBJECTIVE: To examine the effects of DNA dose, site of vaccination, and coadministration of a cytokine DNA adjuvant on efficacy of H1-subtype swine influenza virus hemagglutinin (HA) DNA vaccination of pigs. ANIMALS: 24 eight-week-old mixed-breed pigs. PROCEDURE: 2 doses of DNA were administered 27 days apart by use of a particle-mediated delivery system (gene gun). Different doses of HA DNA and different sites of DNA administration (skin, tongue) were studied, as was coadministration of porcine interleukin-6 (pIL-6) DNA as an adjuvant. Concentrations of virus-specific serum and nasal mucosal antibodies were measured throughout the experiment, and protective immunity was assessed after intranasal challenge with homologous H1N1 swine influenza virus. RESULTS: Increasing the dose of HA DNA, but not coadministration of pIL6 DNA, significantly enhanced virus-specific serum antibody responses. Pigs that received DNA on the ventral surface of the tongue stopped shedding virus 1 day sooner than pigs vaccinated in the skin of the ventral portion of the abdomen, but none of the vaccinated pigs developed detectable virus-specific antibodies in nasal secretions prior to challenge, nor were they protected from challenge exposure. Vaccinated pigs developed high virus-specific antibody concentrations after exposure to the challenge virus. CONCLUSIONS AND CLINICAL RELEVANCE: Co-administration of pIL-6 DNA did not significantly enhance immune responses to HA DNA vaccination or protection from challenge exposure. However, HA DNA vaccination of pigs, with or without coadministration of pIL-6 DNA, induced strong priming of the humoral immune system.

Differentiated swine airway epithelial cell cultures for the investigation of influenza A virus infection and replication.

BACKGROUND: Differentiated human airway epithelial cell cultures have been utilized to investigate cystic fibrosis, wound healing, and characteristics of viral infections. These cultures, grown at an air-liquid interface (ALI) in media with defined hormones and growth factors, recapitulate many aspects of the in vivo respiratory tract and allow for experimental studies at the cellular level. OBJECTIVES: To optimize growth conditions for differentiated swine airway epithelial cultures and to use these cultures to examine influenza virus infection and replication. METHODS: Primary swine respiratory epithelial cells were grown at an air-liquid interface with varying amounts of retinoic acid and epidermal growth factor. Cells grown with optimized concentrations of these factors for 4 weeks differentiated into multilayer epithelial cell cultures resembling the lining of the swine respiratory tract. Influenza virus infection and replication were examined in these cultures. RESULTS/CONCLUSIONS: Retinoic acid promoted ciliogenesis, whereas epidermal growth factor controlled the thickness of the pseudoepithelium. The optimal concentrations for differentiated swine cell cultures were 1·5 ng/ml epidermal growth factor and 100nm retinoic acid. Influenza A viruses infected and productively replicated in these cultures in the absence of exogenous trypsin, suggesting that the cultures express a protease capable of activating influenza virus hemagglutinin. Differences in virus infection and replication characteristics found previously in pigs in vivo were recapitulated in the swine cultures. This system could be a useful tool for a range of applications, including investigating influenza virus species specificity, defining cell tropism of influenza viruses in the swine respiratory epithelium, and studying other swine respiratory diseases.

Infectivity phenotypes of H3N2 influenza A viruses in primary swine respiratory epithelial cells are controlled by sialic acid binding.

BACKGROUND: In the late 1990s, triple reassortant H3N2 influenza A viruses emerged and spread widely in the US swine population. We have shown previously that an isolate representative of this virus-lineage, A/Swine/Minnesota/593/99 (Sw/MN), exhibits phenotypic differences compared to a wholly human-lineage H3N2 virus isolated during the same time period, A/Swine/Ontario/00130/97 (Sw/ONT). Specifically, Sw/MN was more infectious for pigs and infected a significantly higher proportion of cultured primary swine respiratory epithelial cells (SRECs). In addition, reverse genetics-generated Sw/MN × Sw/ONT reassortant and point mutant viruses demonstrated that the infectivity phenotypes in SRECs were strongly dependent on three amino acids within the hemagglutinin (HA) gene. OBJECTIVES: To determine the mechanism by which Sw/MN attains higher infectivity than Sw/ONT in SRECs. METHODS: A/Swine/Minnesota/593/99, Sw/ONT, and mutant (reverse genetics-generated HA reassortant and point mutant) viruses were compared at various HA-mediated stages of infection: initial sialic acid binding, virus entry, and the pH of virus-endosome fusion. RESULTS/CONCLUSIONS: Sialic acid binding was the sole stage where virus differences directly paralleled infectivity phenotypes in SRECs, indicating that binding is the primary mechanism responsible for differences in the infectivity levels of Sw/MN and Sw/ONT.