Publikationsdatenbank

Mimicking passage of avian Influenza virus through the gastrointestinal tract of chickens and characterization of novel chicken intestinal epithelial cell lines (2020)

Art

Hochschulschrift

Autor

Han, Xuejiao (WE 5)

Quelle

Berlin, 2020 — V, 64 Seiten

Sprache

Englisch

Verweise

URL (Volltext): https://refubium.fu-berlin.de/handle/fub188/26578

Kontakt

Institut für Virologie

Robert-von-Ostertag-Str. 7-13
14163 Berlin
+49 30 838 51833
virologie@vetmed.fu-berlin.de

Abstract / Zusammenfassung

Influenza virus is a contagious pathogen of multiple hosts, including humans, other mammals and avian species. In humans and other mammalian hosts viruses usually infect the respiratory epithelium and are airborne transmitted. In contrast, in birds it also infects the intestinal tract and transmits primarily via the fecal-oral route. The major glycoprotein hemagglutinin (HA) of Influenza virus plays a vital role in receptor binding and subsequent fusion of the viral with a host membrane. Proteolytic processing of HA into HA1 and HA2 subunits by trypsin-like proteases and an irreversible conformational change induced by acidic pH are required for membrane fusion. However, acidic pH (as it prevails in the gizzard of birds) inactivates virus particles and high concentrations of trypsin as well as other digestive proteases, such as pepsin and chymotrypsin proteolytically digest HA. In this study, I mimicked the passage of viruses through the gastrointestinal tract by incubating two avian Influenza strains with gizzard and gut fluid from chicken and determined virus titers and integrity of HA by western-blot. The gizzard fluid completely inactivated virions and degrades HA even at a high dilution, but only if the pH was kept acidic. If the fluid is diluted with neutral buffer (mimicking virus uptake with seawater) particles were more resistant. Virions containing an uncleaved HA were even activated suggesting that gastric juice contains a trypsin-like protease. Undiluted intestinal fluid inactivated particles and destroyed HA, but diluted fluid activated virions. Thus, under certain conditions Influenza viruses can withstand the destructive fluids of the avian digestive tract. A remarkable difference exists in this regard between the fowl plaque virus (FPV M1) and a virus isolated from the duck ́s intestine (A/duck/Bavaria/77). While FPV M1 requires a high dilution of 1:1000 to be activated, lower dilutions inactivated the virus and degraded HA. In contrast; the duck-derived virus is already activated at a dilution of 1:10. Thus, the duck-derived virus is more tolerant against intestinal fluid compared to fowl plaque virus suggesting that the former is better adapted to grow in the intestine of birds. Hitherto, no cell lines from the intestine of birds were available. Here I tested a novel chicken intestinal epithelial cell line, which we obtained from the company “MicroMol”. The cell lines show typical epithelial morphology and do not exhibit any sign of senescence during repeated passage. The cell lines are permissive to five Influenza A viruses (two of avian origin and three of human origin), one Influenza B virus, one Influenza C virus and a velogenic Newcastle Disease Virus strain. These cells effectively process HA with a polybasic cleavage site, probably by the cellular protease furin. In contrast, exogenous trypsin is required in multiple step growth experiments, indicating that viruses with a monobasic HA cleavage site are not activated. Thus, these cells have identical properties as any other known cell line that is susceptible to Influenza infection.