Wednesday, 26 February 2014

Using human volunteers to understand influenza virus behaviour and its impact....

This will be the first in a long term project to catalogue the literature's record of volunteer infection studies using influenza virus (IFV). Much like the one I started on rhinovirus transmission back in Nov-2013...see those posts here and here, so far.

I don't guarantee it will be done before I'm too old to type, but I'll be adding to it as time allows. ;)


So first up is a paper.

Author: Frederick Hayden et al
Journal: J Clin Invest 101(3):643-9
Year: 1998
IFV strain used: A/Texas/36/91 (H1N1)
Inoculation route/amount/viral titre: intranasal/0.25ml/105 TCID50

Volunteers included: 19 healthy susceptible males (26%) and females, median age 21-year
Antibody levels: ≤1:8, haemagglutinin-inhibition (IH)

Some key results...


  • Volunteers were kept in isolation for 1-day prior to inoculation.
  • Most (74%) volunteers shed virus on day 1 (1 day after inoculation) as determined by viral growth from recovered nasal washings
  • All volunteers developed symptoms with scores peaking at day-2 and returned to normal by day-8
  • 12/19 (63%) developed fever, peaking at day-2
  • Upper respiratory tract signs of illness (runny nose, sore throat occurred in 15/19 (79%), peaking at day-2
  • Lower respiratory tract symptoms (cough, hoarseness) peaked on day-5
  • Systemic symptoms (muscle aches, fatigue) also peaked on day-2
  • Proinflammatory cytokines in nasal washes revealed several patterns:
    1. Interleukin (IL)-6 and interferon (IFN)-α increased (day-2), decreased, then increased again (day-5) - a biphasic pattern
    2.  IL-8 tumour necrosis factor (TNF) began rising at day-2, peaking at day-4; IL-8 (peaking when nasal mucous turbidity increased) not decreasing as rapidly as TNF
    3. IL-1β, IL-2 and tissue growth factor (TGF)-β did not rise
  • Cytokines in plasma and serum
    • IL-6 and TNFpeaked weakly but at the same times as in the nose 
    • IFN-α and IL-8 were not detectable
    • IL-1βIL-2 and TGF-β did not rise
  • Nasal lavage virus levels (titres) correlated with fever, total symptoms, systemic symptoms and upper respiratory symptoms, TNF, IFN-α and IL-6 on day-2. Total symptoms, viral titre, IL-6 and TNF all correlated on day-5
  • Lower respiratory tract symptom scores peaked in correlation with IFN-α and IL-6 on day-5 & day-6 (also when IL-8 peaked)
As you'd expect given the inoculation route, signs and symptoms of disease occurred first in the upper respiratory tract and then progressed to the lower respiratory tract over time. This is obviously not a "natural" infection as we do not get 0.25mL of flu-containing liquid flying up our noses when someone sneezes on/near us. So the value of this study lies other than in studying natural transmission.

The authors note a few things in the discussion...


  • The response to infection at the site likely leads to the local (runny nose,cough) and the systemic (fever, myalgia) signs and symptoms. 
  • Cytokines are produced by a variety of cells; monocytes/macrophages, bronchial epithelial cells, CD4+ and CD8+ T cells and the infected epithelial cells themselves.
  • Mouse models seem to have more severe lower respiratory tract disease after influenza virus infection than do human volunteers
  • Other cytokines may be involved during more severe natural influenza infections of humans
  • TNF-α, although rising earlier, peaked later perhaps acting to control inflammation after virus has been contained (in the upper respiratory tract at least; what's happening in the lower respiratory tract?)
  • TNF-α and IL-8 were part of the 2nd wave of cytokines and may be better markers of more severe lower respiratory tract influenza disease
Cytokine background information...
  • IL-6
    • A proinflammatory cytokine
    • Secreted by T cells and macrophages
  • IL-8
    • Also known as neutrophil chemotactic factor
    • A chemokine secreted by macrophages and epithelial cells and anything with tool-like receptors
    • Attracts neutrophils and other granulocytes and indices phagocytosis in them
  • TNF-α
    • Also called cachexin or cachectin
    • Mostly secreted by macrophages,but also CD4+ lymphocytes and natural killer (NK) cells
    • It is a pyrogen, inducing fever, apoptotic cell death, cachexia (weight loss) and inflammation
  • IFN-α
    • A Type I interferon secreted by leukocytes, macrophages, epithelial cells, endothelial cells etc
    • Involved in the innate immune response against viral infection by triggering many other antiviral proteins and processes