Studies of influenza virus tropism 公开
Hannah Creager (Fall 2017)
Published
Abstract
Influenza A viruses are capable of causing a wide spectrum of disease across a variety of avian and mammalian hosts. Pathogenicity and transmissibility are thought to be influenced by the cellular and tissue tropism of the virus. Animal models allow for study of tropism in the context of complex hosts, but imperfectly replicate human physiology. In contrast, in vitro studies allow for the examination of human tissue, and the isolation of specific cell types, but are typically of limited complexity. This work bridges the gap between these disparate types of studies by adding additional nuances to in vitro models. For example, infection of adherent cell monolayers using a liquid inoculum represents an established method to reliably and quantitatively study virus infection, but poorly recapitulates the exposure and infection of cells in the respiratory tract that occurs during infection with aerosolized pathogens. We therefore began by developing methodology to expose adherent mammalian cell monolayers to defined quantities of aerosolized influenza virus. We found that certain viruses were able to replicate productively in human primary alveolar epithelial cells following liquid, but not aerosol inoculation. Additionally, aerosolized virus was able to infect a human bronchial epithelial cell line in spite of a thick apical mucus layer resulting from three weeks of culture at air-liquid interface, but viral infectivity was reduced up to 25-fold. By facilitating study of viral infectivity under conditions similar to those of natural infection and transmission, this novel method has the potential to enhance our understanding of influenza and respiratory viruses.
Next, we applied aerosol inoculation to the study of ocular tropism. Though influenza viruses typically cause respiratory tract disease, some viruses, particularly those with an H7 hemagglutinin, have been isolated from the eyes of conjunctivitis cases. We asked whether conjunctivitis-associated viruses were unique in the ability to infect ocular cells by the aerosol route or to replicate efficiently at 33°C and found that they were not. Similarly, the membrane associated mucins expressed on differentiated corneal tissue constructs did not restrict the infection or replication of respiratory isolates. However, human tears significantly inhibited hemagglutination of both ocular and non-ocular isolates but the effect on viral infectivity was more variable, with tears reducing the infectivity of non-ocular isolates more than ocular isolates. These data suggest that most influenza viruses may be capable of establishing infection if they reach the ocular surface, but that this is more likely for ocular isolates as they are better able to maintain their infectivity in the presence of tears.
Table of Contents
Abstract
Table of Contents
List of Tables and Figures
Chapter 1: Introduction 1
Virus Structure and Replication 1
Avian influenza 3
Influenza in humans 4
Influenza viral tropism 8
Chapter 2: In vitro exposure system for study of aerosolized influenza virus 13
Abstract 14
Introduction 15
Materials and Methods 17
Results 21
Discussion 27
Acknowledgements 30
References 32
Tables 43
Figures 44
Supplementary Materials 51
Chapter 3: Infection and replication of influenza virus at the ocular surface 55
Abstract 56
Introduction 57
Results 58
Discussion 67
Materials and Methods 73
Acknowledgements 76
References 77
Tables 88
Figures 89
Supplemental Information 100
Chapter 4: Discussion 104
References 112
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Studies of influenza virus tropism () | 2017-09-29 15:21:28 -0400 |
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