No abstract

Pigs are natural hosts for influenza A viruses and play a critical role in influenza epidemiology. However, little is known about their influenza-evoked T-cell response. We performed a thorough analysis of both the local and systemic T-cell response in influenza virus-infected pigs, addressing kinetics and phenotype as well as multifunctionality (gamma interferon [IFN-γ], tumor necrosis factor alpha [TNF-α], and interleukin-2 [IL-2]) and cross-reactivity. A total of 31 pigs were intratracheally infected with an H1N2 swine influenza A virus (FLUAVsw) and consecutively euthanized. Lungs, tracheobronchial lymph nodes, and blood were sampled during the first 15 days postinfection (p.i.) and at 6 weeks p.i. Ex vivo flow cytometry of lung lymphocytes revealed an increase in proliferating (Ki-67+) CD8+ T cells with an early effector phenotype (perforin+ CD27+) at day 6 p.i. Low frequencies of influenza virus-specific IFN-γ-producing CD4+ and CD8+ T cells could be detected in the lung as early as 4 days p.i. On consecutive days, influenza virus-specific CD4+ and CD8+ T cells produced mainly IFN-γ and/or TNF-α, reaching peak frequencies around day 9 p.i., which were up to 30-fold higher in the lung than in tracheobronchial lymph nodes or blood. At 6 weeks p.i., CD4+ and CD8+ memory T cells had accumulated in lung tissue. These cells showed diverse cytokine profiles and in vitro reactivity against heterologous influenza virus strains, all of which supports their potential to combat heterologous influenza virus infections in pigs.IMPORTANCE Pigs not only are a suitable large-animal model for human influenza virus infection and vaccine development but also play a central role in the emergence of new pandemic strains. Although promising candidate universal vaccines are tested in pigs and local T cells are the major correlate of heterologous control, detailed and targeted analyses of T-cell responses at the site of infection are scarce. With the present study, we provide the first detailed characterization of magnitude, kinetics, and phenotype of specific T cells recruited to the lungs of influenza virus-infected pigs, and we could demonstrate multifunctionality, cross-reactivity, and memory formation of these cells. This, and ensuing work in the pig, will strengthen the position of this species as a large-animal model for human influenza virus infection and will immediately benefit vaccine development for improved control of influenza virus infections in pigs.

Although swine are natural hosts for influenza A viruses, the porcine T-cell response to swine influenza A virus (FLUAVsw) infection has been poorly characterized so far. We have studied Ki-67 expression and FLUAVsw-specific production of IFN-γ, TNF-α and IL-2 in CD4+ and CD8β+ T cells isolated from piglets that had been intratracheally infected with a H1N2 FLUAVsw isolate. IFN-γ+TNF-α+IL-2+ multifunctional CD4+ T cells were present in the blood of all infected animals at one or two weeks after primary infection and their frequency increased in four out of six animals after homologous secondary infection. These cells produced higher amounts of IFN-γ, TNF-α and IL-2 than did CD4+ T cells that only produced a single cytokine. The vast majority of cytokine-producing CD4+ T cells expressed CD8α, a marker associated with activation and memory formation in porcine CD4+ T cells. Analysis of CD27 expression suggested that FLUAVsw-specific CD4+ T cells included both central memory and effector memory populations. Three out of six animals showed a strong increase of Ki-67+perforin+ CD8β+ T cells in blood one week post infection. Blood-derived FLUAVsw-specific CD8β+ T cells could be identified after an in vitro expansion phase and were multifunctional in terms of CD107a expression and co-production of IFN-γ and TNF-α. These data show that multifunctional T cells are generated in response to FLUAVsw infection of pigs, supporting the idea that T cells contribute to the efficient control of infection.Electronic supplementary materialThe online version of this article (doi:10.1186/s13567-015-0182-3) contains supplementary material, which is available to authorized users.

In order to estimate the diversity, clinical involvement and zoonotic potential of parasites in pigs submitted for diagnosis to the PathoPig project of the Swiss Federal Food Safety and Veterinary Office, faeces (n=125) from suckling piglets (n=39), weaners (n=60) and piglets beginning fattening (n=26) from 74 Swiss farms were examined by 3 coproscopical methods (i.e. sedimentation/zinc chloride-flotation; SAFC and Ziehl-Neelsen staining). Samples microscopically positive for Cryptosporidium were further tested by PCR/sequencing for species assessment. The most frequently detected parasite was Balantidium coli, a facultative pathogenic ciliate with zoonotic potential, in 5.1, 36.7 and 50.0% of suckling, weaners and fatteners and 43.2% of farms; however, no association with disease was observed. Isospora (syn. Cystoisospora) suis infections were detected in 13.3 and 11.1% of suckling piglets with and without diarrhoea, and in 10.0 and 13.3% of weaners and fatteners with diarrhoea, respectively, and were significant associated with emaciation. Cryptosporidium infections were detected in 10.3, 15.0 and 19.2% of sucklings, weaners and fatteners, respectively, and in 18.9% of the farms. Interestingly, two age-related species were identified: C. suis in younger piglets (2 to 6weeks) and C. scrofarum in older ones (6 to 17weeks). None of the pigs infected with C. scrofarum (n=8), but 3 of 4 piglets infected with C. suis (co-infection with I. suis in 2 cases) had diarrhoea. The zoonotic species C. parvum was not detected, nevertheless, sporadic cases of human infection with the porcine-adapted species have been reported. Ascaris suum, Trichuris suis and Strongylida were rarely detected (<4%) in all age categories.