Silent Infection of B and CD8 + T Lymphocytes by Influenza A Virus in Children with Tonsillar Hypertrophy
Influenza A viruses (IAVs) cause more than 2 million annual episodes of seasonal acute respiratory infections (ARI) and approximately 500,000 deaths worldwide. Depending on virus strain and host immune status, acute infections by IAV may reach sites other than the respiratory tract. In the present s...
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| Vydáno v: | Journal of virology Ročník 94; číslo 9 |
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| Hlavní autoři: | , , , , , , , , , , , , , |
| Médium: | Journal Article |
| Jazyk: | angličtina |
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16.04.2020
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| ISSN: | 1098-5514, 1098-5514 |
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| Abstract | Influenza A viruses (IAVs) cause more than 2 million annual episodes of seasonal acute respiratory infections (ARI) and approximately 500,000 deaths worldwide. Depending on virus strain and host immune status, acute infections by IAV may reach sites other than the respiratory tract. In the present study, IAV RNA and antigens were searched for in tissues of palatine tonsils and adenoids removed from patients without ARI symptoms. A real-time reverse transcriptase PCR (RT-PCR) screening revealed that 8 tissue samples from 7 patients out of 103 were positive for IAV. Positive samples were subjected to next-generation sequencing (NGS) and 3 of 8 tissues yielded complete IAV pH1N1 genomes, whereas in 5 samples, the PB1 gene was not fully assembled. Phylogenetic analysis placed tonsil-derived IAV in clusters clearly segregated from contemporaneous Brazilian viruses. Flow cytometry of dispersed tissue fragments and serial immunohistochemistry of paraffin-embedded sections of naturally infected biopsies indicated that CD20
B lymphocytes, CD8
T lymphocytes, and CD11c
cells are susceptible to IAV infection. We sought to investigate whether these lymphoid tissues could be sites of viral replication and sources of viable virus particles. MDCK cells were inoculated with tissue lysates, enabling recovery of one IAV isolate confirmed by immunofluorescence, reverse transcriptase quantitative PCR (RT-qPCR), and NGS. The data indicate that lymphoid tissues not only harbor expression of IAV proteins but also contain infectious virus. Asymptomatic long-term infection raises the possibility of IAV shedding from tonsils, which may have an impact on host-to-host transmission.
Influenza A virus (IAV) infections are important threats to human health worldwide. Although extensively studied, some aspects of virus pathogenesis and tissue tropism remain unclear. Here, by different strategies, we describe the asymptomatic infection of human lymphoid organs by IAV in children. Our results indicate that IAV was not only detected and isolated from human tonsils but displayed unique genetic features in comparison with those of contemporaneous IAVs circulating in Brazil and detected in swabs and nasal washes. Inside the tissue microenvironment, immune cells were shown to be carrying IAV antigens, especially B and T CD8
lymphocytes. Taken together, these results suggest that human lymphoid tissues can be sites of silent IAV infections with possible impact on virus shedding to the population. |
|---|---|
| AbstractList | Influenza A viruses (IAVs) cause more than 2 million annual episodes of seasonal acute respiratory infections (ARI) and approximately 500,000 deaths worldwide. Depending on virus strain and host immune status, acute infections by IAV may reach sites other than the respiratory tract. In the present study, IAV RNA and antigens were searched for in tissues of palatine tonsils and adenoids removed from patients without ARI symptoms. A real-time reverse transcriptase PCR (RT-PCR) screening revealed that 8 tissue samples from 7 patients out of 103 were positive for IAV. Positive samples were subjected to next-generation sequencing (NGS) and 3 of 8 tissues yielded complete IAV pH1N1 genomes, whereas in 5 samples, the PB1 gene was not fully assembled. Phylogenetic analysis placed tonsil-derived IAV in clusters clearly segregated from contemporaneous Brazilian viruses. Flow cytometry of dispersed tissue fragments and serial immunohistochemistry of paraffin-embedded sections of naturally infected biopsies indicated that CD20+ B lymphocytes, CD8+ T lymphocytes, and CD11c+ cells are susceptible to IAV infection. We sought to investigate whether these lymphoid tissues could be sites of viral replication and sources of viable virus particles. MDCK cells were inoculated with tissue lysates, enabling recovery of one IAV isolate confirmed by immunofluorescence, reverse transcriptase quantitative PCR (RT-qPCR), and NGS. The data indicate that lymphoid tissues not only harbor expression of IAV proteins but also contain infectious virus. Asymptomatic long-term infection raises the possibility of IAV shedding from tonsils, which may have an impact on host-to-host transmission.IMPORTANCE Influenza A virus (IAV) infections are important threats to human health worldwide. Although extensively studied, some aspects of virus pathogenesis and tissue tropism remain unclear. Here, by different strategies, we describe the asymptomatic infection of human lymphoid organs by IAV in children. Our results indicate that IAV was not only detected and isolated from human tonsils but displayed unique genetic features in comparison with those of contemporaneous IAVs circulating in Brazil and detected in swabs and nasal washes. Inside the tissue microenvironment, immune cells were shown to be carrying IAV antigens, especially B and T CD8+ lymphocytes. Taken together, these results suggest that human lymphoid tissues can be sites of silent IAV infections with possible impact on virus shedding to the population.Influenza A viruses (IAVs) cause more than 2 million annual episodes of seasonal acute respiratory infections (ARI) and approximately 500,000 deaths worldwide. Depending on virus strain and host immune status, acute infections by IAV may reach sites other than the respiratory tract. In the present study, IAV RNA and antigens were searched for in tissues of palatine tonsils and adenoids removed from patients without ARI symptoms. A real-time reverse transcriptase PCR (RT-PCR) screening revealed that 8 tissue samples from 7 patients out of 103 were positive for IAV. Positive samples were subjected to next-generation sequencing (NGS) and 3 of 8 tissues yielded complete IAV pH1N1 genomes, whereas in 5 samples, the PB1 gene was not fully assembled. Phylogenetic analysis placed tonsil-derived IAV in clusters clearly segregated from contemporaneous Brazilian viruses. Flow cytometry of dispersed tissue fragments and serial immunohistochemistry of paraffin-embedded sections of naturally infected biopsies indicated that CD20+ B lymphocytes, CD8+ T lymphocytes, and CD11c+ cells are susceptible to IAV infection. We sought to investigate whether these lymphoid tissues could be sites of viral replication and sources of viable virus particles. MDCK cells were inoculated with tissue lysates, enabling recovery of one IAV isolate confirmed by immunofluorescence, reverse transcriptase quantitative PCR (RT-qPCR), and NGS. The data indicate that lymphoid tissues not only harbor expression of IAV proteins but also contain infectious virus. Asymptomatic long-term infection raises the possibility of IAV shedding from tonsils, which may have an impact on host-to-host transmission.IMPORTANCE Influenza A virus (IAV) infections are important threats to human health worldwide. Although extensively studied, some aspects of virus pathogenesis and tissue tropism remain unclear. Here, by different strategies, we describe the asymptomatic infection of human lymphoid organs by IAV in children. Our results indicate that IAV was not only detected and isolated from human tonsils but displayed unique genetic features in comparison with those of contemporaneous IAVs circulating in Brazil and detected in swabs and nasal washes. Inside the tissue microenvironment, immune cells were shown to be carrying IAV antigens, especially B and T CD8+ lymphocytes. Taken together, these results suggest that human lymphoid tissues can be sites of silent IAV infections with possible impact on virus shedding to the population. Influenza A viruses (IAVs) cause more than 2 million annual episodes of seasonal acute respiratory infections (ARI) and approximately 500,000 deaths worldwide. Depending on virus strain and host immune status, acute infections by IAV may reach sites other than the respiratory tract. In the present study, IAV RNA and antigens were searched for in tissues of palatine tonsils and adenoids removed from patients without ARI symptoms. A real-time reverse transcriptase PCR (RT-PCR) screening revealed that 8 tissue samples from 7 patients out of 103 were positive for IAV. Positive samples were subjected to next-generation sequencing (NGS) and 3 of 8 tissues yielded complete IAV pH1N1 genomes, whereas in 5 samples, the PB1 gene was not fully assembled. Phylogenetic analysis placed tonsil-derived IAV in clusters clearly segregated from contemporaneous Brazilian viruses. Flow cytometry of dispersed tissue fragments and serial immunohistochemistry of paraffin-embedded sections of naturally infected biopsies indicated that CD20 B lymphocytes, CD8 T lymphocytes, and CD11c cells are susceptible to IAV infection. We sought to investigate whether these lymphoid tissues could be sites of viral replication and sources of viable virus particles. MDCK cells were inoculated with tissue lysates, enabling recovery of one IAV isolate confirmed by immunofluorescence, reverse transcriptase quantitative PCR (RT-qPCR), and NGS. The data indicate that lymphoid tissues not only harbor expression of IAV proteins but also contain infectious virus. Asymptomatic long-term infection raises the possibility of IAV shedding from tonsils, which may have an impact on host-to-host transmission. Influenza A virus (IAV) infections are important threats to human health worldwide. Although extensively studied, some aspects of virus pathogenesis and tissue tropism remain unclear. Here, by different strategies, we describe the asymptomatic infection of human lymphoid organs by IAV in children. Our results indicate that IAV was not only detected and isolated from human tonsils but displayed unique genetic features in comparison with those of contemporaneous IAVs circulating in Brazil and detected in swabs and nasal washes. Inside the tissue microenvironment, immune cells were shown to be carrying IAV antigens, especially B and T CD8 lymphocytes. Taken together, these results suggest that human lymphoid tissues can be sites of silent IAV infections with possible impact on virus shedding to the population. |
| Author | Jesus, Bruna L S Castro, Italo A Cardoso, Ricardo S Valera, Fabiana C P Pontelli, Marjorie C Jorge, Daniel M M Martins, Ronaldo B Carenzi, Lucas Ferreri, Lucas M Arruda, Eurico Anselmo-Lima, Wilma T Perez, Daniel R Criado, Miria F Tamashiro, Edwin |
| Author_xml | – sequence: 1 givenname: Italo A orcidid: 0000-0001-7885-7547 surname: Castro fullname: Castro, Italo A email: italo.a.castro@usp.br, eaneto@fmrp.usp.br organization: Department of Cell and Molecular Biology, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil italo.a.castro@usp.br eaneto@fmrp.usp.br – sequence: 2 givenname: Daniel M M surname: Jorge fullname: Jorge, Daniel M M organization: Department of Cell and Molecular Biology, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 3 givenname: Lucas M surname: Ferreri fullname: Ferreri, Lucas M organization: Poultry Diagnostic and Research Center, Department of Population Health, University of Georgia, Athens, Georgia, USA – sequence: 4 givenname: Ronaldo B orcidid: 0000-0002-8902-5962 surname: Martins fullname: Martins, Ronaldo B organization: Department of Cell and Molecular Biology, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 5 givenname: Marjorie C orcidid: 0000-0003-3173-5977 surname: Pontelli fullname: Pontelli, Marjorie C organization: Department of Cell and Molecular Biology, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 6 givenname: Bruna L S surname: Jesus fullname: Jesus, Bruna L S organization: Department of Cell and Molecular Biology, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 7 givenname: Ricardo S orcidid: 0000-0003-2858-2076 surname: Cardoso fullname: Cardoso, Ricardo S organization: Department of Cell and Molecular Biology, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 8 givenname: Miria F surname: Criado fullname: Criado, Miria F organization: Department of Cell and Molecular Biology, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 9 givenname: Lucas surname: Carenzi fullname: Carenzi, Lucas organization: Department of Ophthalmology, Otorhinolaryngology and Head and Neck Surgery, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 10 givenname: Fabiana C P surname: Valera fullname: Valera, Fabiana C P organization: Department of Ophthalmology, Otorhinolaryngology and Head and Neck Surgery, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 11 givenname: Edwin surname: Tamashiro fullname: Tamashiro, Edwin organization: Department of Ophthalmology, Otorhinolaryngology and Head and Neck Surgery, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 12 givenname: Wilma T surname: Anselmo-Lima fullname: Anselmo-Lima, Wilma T organization: Department of Ophthalmology, Otorhinolaryngology and Head and Neck Surgery, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil – sequence: 13 givenname: Daniel R surname: Perez fullname: Perez, Daniel R organization: Poultry Diagnostic and Research Center, Department of Population Health, University of Georgia, Athens, Georgia, USA – sequence: 14 givenname: Eurico orcidid: 0000-0002-0978-410X surname: Arruda fullname: Arruda, Eurico email: italo.a.castro@usp.br, eaneto@fmrp.usp.br organization: Department of Cell and Molecular Biology, Ribeirao Preto School of Medicine, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil italo.a.castro@usp.br eaneto@fmrp.usp.br |
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