Porcine CD3(+)NKp46(+) Lymphocytes Have NK-Cell Characteristics and Are Present in Increased Frequencies in the Lungs of Influenza-Infected Animals.

The CD3(-)NKp46(+) phenotype is frequently used for the identification of natural killer (NK) cells in various mammalian species. Recently, NKp46 expression was analyzed in more detail in swine. It could be shown that besides CD3(-)NKp46(+) lymphocytes, a small but distinct population of CD3(+)NKp46(+) cells exists. In this study, we report low frequencies of CD3(+)NKp46(+) lymphocytes in blood, lymph nodes, and spleen, but increased frequencies in non-lymphatic organs, like liver and lung. Phenotypic analyses showed that the majority of CD3(+)NKp46(+) cells coexpressed the CD8αß heterodimer, while a minor subset expressed the TCR-γδ, which was associated with a CD8αα(+) phenotype. Despite these T-cell associated receptors, the majority of CD3(+)NKp46(+) lymphocytes displayed a NK-related phenotype (CD2(+)CD5(-)CD6(-)CD16(+)perforin(+)) and expressed mRNA of NKp30, NKp44, and NKG2D at similar levels as NK cells. Functional tests showed that CD3(+)NKp46(+) lymphocytes produced IFN-γ and proliferated upon cytokine stimulation to a similar extent as NK cells, but did not respond to the T-cell mitogen, ConA. Likewise, CD3(+)NKp46(+) cells killed K562 cells with an efficiency comparable to NK cells. Cross-linking of NKp46 and CD3 led to degranulation of CD3(+)NKp46(+) cells, indicating functional signaling pathways for both receptors. Additionally, influenza A(H1N1)pdm09-infected pigs had reduced frequencies of CD3(+)NKp46(+) lymphocytes in blood, but increased frequencies in the lung in the early phase of infection. Thus, CD3(+)NKp46(+) cells appear to be involved in the early phase of influenza infections. In summary, we describe a lymphocyte population in swine with a mixed phenotype of NK and T cells, with results so far indicating that this cell population functionally resembles NK cells.

Adverse effects of Influenza A(H1N1)pdm09 virus infection on growth performance of Norwegian pigs - a longitudinal study at a boar testing station.

BACKGROUND: Influenza A(H1N1)pdm09 virus infection in Norwegian pigs was largely subclinical. This study tested the hypothesis that the infection causes negligible impact on pigs' growth performance in terms of feed conversion efficiency, daily feed intake, daily growth, age on reaching 100 kg bodyweight and overall feed intake. A sample of 1955 pigs originating from 43 breeding herds was classified into five infection status groups; seronegative pigs (n = 887); seropositive pigs (n = 874); pigs positive for virus at bodyweight between 33 kg and 60 kg (n = 123); pigs positive for virus at bodyweight between 61 kg and 80 kg (n = 34) and pigs positive for virus at bodyweight between 81 kg and 100 kg (n = 37). Each pig had daily recordings of feed intake and bodyweight from 33 kg to 100 kg. Marginal effects of the virus infection on the outcomes were estimated by multi-level linear regression, which accounted for known fixed effects (breed, birthdate, average daily feed intake and growth phase) and random effects (cluster effects of pig and herd). RESULTS: The seropositive and virus positive pigs had decreased (P value<0.05) growth performance compared to seronegative pigs even though feed intake was not decreased. Reduced feed conversion efficiency led to lower average daily growth, additional feed requirement and longer time needed to reach the 100 kg bodyweight. The effects were more marked (P value<0.03) in pigs infected at a younger age and lasted a longer period. Despite increased feed intake observed, their growth rates were lower and they took more time to reach 100 kg bodyweight compared to the seronegative pigs. CONCLUSION: Our study rejected the null hypothesis that the virus infection had negligible adverse effects on growth performance of Norwegian pigs.

Early responses of natural killer cells in pigs experimentally infected with 2009 pandemic H1N1 influenza A virus.

Natural killer (NK) cells are important players in the innate immune response against influenza A virus and the activating receptor NKp46, which binds hemagglutinin on the surface of infected cells, has been assigned a role in this context. As pigs are natural hosts for influenza A viruses and pigs possess both NKp46- and NKp46+ NK cells, they represent a good animal model for studying the role of the NKp46 receptor during influenza. We explored the role of NK cells in piglets experimentally infected with 2009 pandemic H1N1 influenza virus by flow cytometric analyses of cells isolated from blood and lung tissue and by immunostaining of lung tissue sections. The number of NKp46+ NK cells was reduced while NKp46- NK cells remained unaltered in the blood 1-3 days after infection. In the lungs, the intensity of NKp46 expression on NK cells was increased during the first 3 days, and areas where influenza virus nucleoprotein was detected were associated with increased numbers of NKp46+ NK cells when compared to uninfected areas. NKp46+ NK cells in the lung were neither found to be infected with influenza virus nor to be undergoing apoptosis. The binding of porcine NKp46 to influenza virus infected cells was verified in an in vitro assay. These data support the involvement of porcine NKp46+ NK cells in the local immune response against influenza virus.

Swine influenza in Norway: a distinct lineage of influenza A(H1N1)pdm09 virus.

BACKGROUND: Since the influenza A(H1N1)pdm09 virus was first introduced to the Norwegian pig population in September 2009, it has repeatedly been detected in pigs in Norway. No other subtypes of influenza virus are circulating in Norwegian pigs. OBJECTIVE: To follow the diversity of A(H1N1)pdm09 viruses circulating in pigs in Norway and to investigate the relationship between viruses circulating in Norwegian pigs and in humans. METHODS: Between January 2011 and January 2013, nasal swabs from 507 pigs were tested for A(H1N1)pdm09 virus by real-time RT-PCR. The hemagglutinin (HA) gene of virus-positive samples was sequenced and compared with publically available sequences from viruses circulating in humans at the time. RESULTS: Sequencing and phylogenetic analysis of the HA gene showed that the A(H1N1)pdm09 virus circulating in Norwegian pigs early in 2011 resembled the A(H1N1)pdm09 virus circulating in humans during this time. Viruses detected in pigs by the end of 2011 had acquired four characteristic amino acid substitutions (N31D, S84I S164F, and N473D) and formed a distinct phylogenetic group. CONCLUSIONS: A(H1N1)pdm09 virus detected in Norwegian pigs by the end of 2011 formed a distinct genetic lineage. Also, our findings indicate that reverse-zoonotic transmission from humans to pigs of the A(H1N1)pdm09 virus is still important.