Ebola virus crisis Prof. Viktor Volchkov Molecular basis of viral - - PowerPoint PPT Presentation

Ebola virus crisis

Molecular basis of viral pathogenicity

• Prof. Viktor Volchkov

International Centre for Research in Infectiology (CIRI), INSERM Claude Bernard University Lyon-1, Lyon, France viktor.volchkov@inserm.fr Leem Press Workshop March 10, 2015

Jean Mérieux INSERM BSL 4 Laboratory

 The molecular basis of high pathogenicity  Emergency of highly pathogenic viruses  Lessons from current Ebola crisis

BSL P4 viral agents

Nipah virus / Henipaviridae Ebola and Marburg viruses / Filoviridae Crimean-Congo hemorrhagic fever virus / Bunyaviridae

BSL P4 viral agents

Infections kill up to 60-90% of patients Spread through human-to-human or animal-to-human contacts Intercontinental virus transfer / Potential bio-terrorism agent Currently no vaccines or treatments are available for human use Molecular bases of high pathogenicity are not well understood

Genomic RNA membrane Surface glycoprotein: GP

. Nucleoprotein(NP) . VP30 . VP35

. Polymerase (L) Matrix protein VP40 VP24

• Enveloped, non-segmented negative-stranded RNA containing virus
• 7 structural proteins and 3 non-structural soluble proteins

5’

NP L 24 30 GP 40 35

3’

Ebola virus

Massive virus replication

Innate Immune responses Oxidative stress Disregulation of host inflammatory responses

Leung et al., JID 2011; Page et al., Cell Reports 2014

High pathogenicity

“Masking” phenomenon/ GP gene editing and virus adaptation

Reynard et al., J Virol 2009; Volchkova et al., JID 2011

GP/shedGP VP35/VP24 EBOV and MARV VP24

Inflammation Molecular determinants of virulence

Mateo et al., JID 2011

VP35 and VP24 are IFN antagonists

Prins et al., J Virol 2010; Mateo et al., J Virol 2010.

VP30/VP24

RNA Synthesis/Virus assembly Molecular switch: RNA replication versus transcription

Martinez et al., J Virol 2010, JID 2011

Role of VP24 in viral nucleocapsids assembly

Mateo et al., JID 2011

Role of EBOV GP shedding

Esqudero-Perez et al., Plos Pathogenes 2014

EBOV-infected cell EBOV virion Shed GP Shed GP Membrane GP

Introduction

Shedding of EBOV surface glycoprotein GP

Block virus neutralizing antibodies sGP

PLOS Pathogens, 2014

Results

Increase of permeability Activati

• n

Imballance d inflammation DC/MØ B/T lymphocytes Hepatocyte, renal cells, etc Endothelial cells Organ failure Blood cells / fluid

Non-infected cells

TLR-4

Transcriptional activation and release of cytokines EBOV-infected cell EBOV Shed GP

Shed GP of Ebola virus Triggers Systemic Immune Activation and Increased Vascular Permeability

TNF-α IL-6 IL-12 IL-10 etc.

 Multiple antagonists of innate immunity  Excessive and dysregulated host responses to viral replication  High efficiency of replication and viral pantropism

Factors that contribute to high pathogenicity

« Loaded gun »

Natural reservoir  Asymptomatic infection High level of virus adaptation  Low virus replication

Virus –host interaction (I)

 High pathogenicity Low level of virus adaptation  High level of virus replication Accidental hosts

Virus –host interaction (II)

Virus emergence (I)

Natural reservoir Accidental hosts

Local infections End of outbreak Hospital Strict Quarantine Low Level Health Care Virus spread Difficult to apply quarantine Nosocomial infection

Virus emergence (II)

• Always keep watching for known highly pathogenic viruses.
• Keep moving forward. Learn more on molecular basis of high

pathogenicity.

Lessons from current Ebola crisis:

• Preparedness:

 Diagnosis  Vaccine  Therapeutics For all known highly pathogenic viruses.

• Search for new, related viruses (African henipa- and filoviruses)
• Exploring the link between Academic Science and Industrials.

BACK UP

Ebola and Marburg viruses, Nipah virus and CCHF virus

Research tools

NP 35 L 30 NP 35 L 30

30 NC genes (T7 promoter)

Recombinant Ebola virus

(cross section)

pFL-EBOV T7 promoter Ribozyme

L 24 GP 30 40 35 NP

pFL-EBOV

L 35 NP BSR T7/5 cells

3 4 1 5 6

mRNA`s

1 2

T7 T7 T7 T7

Volchkov et al Science 2001

Reverse genetics approach

200 250 300 350 400 450 500 550 600

• 5
• 3
• 1

1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 days post infection

weight(g)

200 250 300 350 400 450 500

• 5
• 3
• 1

1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 days post infection

weight(g)

NE WT

D7 D27 D16 αVP40

200 250 300 350 400 450 500

• 5
• 3
• 1

1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 days post infection

weight(g)

JID, 2015

Results

EBOV mutant is completely attenuated