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Crimean-Congo Haemorrhagic Fever DIAGNOSIS. Hervé Zeller National Reference Center - WHO Collaborating Centre for Arboviruses and Viral Haemorrhagic Fevers, Institut Pasteur, Lyon. Flaviviridae (dengue, yellow fever, Groupe TBE). Viral Haemorrhagic Fevers. Flaviviridae

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slide1

Crimean-Congo Haemorrhagic Fever

DIAGNOSIS

Hervé Zeller

National Reference Center - WHO Collaborating Centre for

Arboviruses and Viral Haemorrhagic Fevers, Institut Pasteur, Lyon

slide2

Flaviviridae

(dengue, yellow fever,

Groupe TBE)

Viral Haemorrhagic Fevers

slide3

Flaviviridae

(dengue, yellow fever,

Groupe TBE)

Viral Haemorrhagic Fevers

Arenaviridae

(Lassa, Junin, Machupo, Guanarito)

slide4

Flaviviridae

(dengue, yellow fever,

Groupe TBE)

Viral Haemorrhagic Fevers

Arenaviridae

(Lassa, Junin, Machupo, Guanarito)

Filoviridae

(Ebola, Marburg)

slide5

Flaviviridae

(dengue, yellow fever,

Groupe TBE)

Viral Haemorrhagic Fevers

Arenaviridae

(Lassa, Junin, Machupo, Guanarito)

Envelopped

RNA viruses

Bunyaviridae

(CCHF, RVF,

Hantaviruses)

Filoviridae

(Ebola, Marburg)

slide6

Family Genus VIRUS DISTRIBUTION

Flaviviridae Flavivirus Yellow Fever Africa South America

Dengue 1,2,3,4. Tropical areas

Omsk HF Russia

Alkhurma Saudi Arabia

Kyasanur Forest HF India

Bunyaviridae Phlebovirus Rift Valley Fever Africa, Saudi Arabia

NairovirusCrimean-Congo HF Africa, Eurasia

Hantavirus Hantan Dobrava Puumala Eurasia

Sin Nombre, Andes Americas

slide7

Hemorrhages are inconstant :

Emerging part of the iceberg

…Most frequently

asymptomatic infections +++

biosafety issues related to haemorrhagic fever viruses
Biosafety Issues Related to Haemorrhagic Fever Viruses
  • Criteria for classification
    • Disease severity
    • Transmissibility to laboratory workers
    • Availability of treatment
    • Availability of vaccine
  • Classification BSL 1 to BSL4
slide9

CCHF VIABILITY

  • SENSITIVITY TO DESINFECTANTS:
  • sodium hypochlorite 2%, glutaraldehyde 2%, formaldehyde
  • SENSITIVE TO DESSICATION
  • INACTIVATION :
  • IRRADIATION
  • UV
  • TEMPERATURE : 1 hour 60°C
  • not complete inactivation
        • beta propiolactone 4°C
        • not complete inactivation
  • Triton X100
slide10

Nairovirus structure

G1

M

S

N

G2

L

L

10 nm

slide11

CCHFNairovirus genome

_____________________________________________

Segment Nucleotides Amino acids Protein

_____________________________________________

S 1659-1712 442-482 N

M 4888 1551 G1 G2 NSm?

L 12255 4036 L?

_____________________________________________

slide13

VHF SUSPECT CASE

Malaria – Hepatitis – Typhoidis – Toxicosis

Septicemia – Leptospirosis

Rickettsiosis…

slide14

VHF SUSPECT CASE

Malaria – Hepatitis – Typhoidis – Toxicosis

Septicemia – Leptospirosis

Rickettsiosis…

Epidemiological data, risk evaluation

biological analysis, differential diagnostic

slide15

VHF SUSPECT CASE

Malaria – Hepatitis – Typhoidis – Toxicosis

Septicemia – Leptospirosis

Rickettsiosis…

Epidemiological data, risk evaluation

biological analysis, differential diagnostic

Contact between clinicians and biologists

cchf laboratory data
CCHF : laboratory data

LEUCOPENIA, particularly neutropenia

THROMBOCYTOPENIA

Hematocrite increases early then falls down

ASL, AST levels increases

Proteinuria and hematuria

Mild azotemia, bilirubine increase

cchf laboratory diagnosis
CCHF : laboratory diagnosis

Viral detection: blood specimen

  • RT-PCR (nested)
  • Cell culture (Vero E6 cells)
cchf laboratory diagnosis1
CCHF : laboratory diagnosis

Viral detection: (blood specimen)

  • RT-PCR (nested)
  • Cell culture (Vero E6 cells)

Antibody detection : (serum sample)

  • IFA
  • ELISA IgM (immuno-capture) IgG
  • NT
cchf laboratory diagnosis2
CCHF : laboratory diagnosis

Viremia 10-12 days (although afebrile).

Can be detected by PCR up to day 16

By day 9 all patients will have IgM or IgG antibody

Information needed : DATE OF ONSET OF FEVER

slide20

CCHF : viral/antibody kinetics

IgM

IgG

viremia

0

5

10

RT-PCR

16

Viral isolation

ELISA IgM IgG

IFA

IgM duration: 2-3 months up to 6 months…

slide22

RT-PCR /Southern blot hybridization/antibody : retrospective study

From Burt et al,

J Virol Methods 1998, 70:129-137

slide23

RT-PCR /Southern blot hybridization/IFA antibody :

26 samples from 19 patients from day 3-12 of illness

From :

Burt et al, J Virol Methods 1998, 70:129-137

slide24

Hyalomma sp.

ticks

RT-PCR

Viral isolation

slide25

536 pb amplicons of the S fragment of CCHF genome using primers CSDR3/CSDF2. RFLP with Hinf I, Hae III, and Alu I endonucleases

PROFIL

ORIGINE

DUGBE

RFLP

Grèce

AP 92

3 2 1

AnD 15786

1 1 1

Sénégal

(100)

Sénégal

(100)

ArD 8194

1 3 1

ArTeh 193-3

Iran

1 3 1

(96)

HD 49199

1 3 1

Mauritanie

ArMg 951

1 2 3

(57)

Madagascar

(84)

Chine

C 68031

2 2 4

Rép. Centrafr.

ArB 604

2 2 3

(100)

(99)

Burkina Faso

HD 38562

2 2 3

Mauritanie

ArD 39554

2 2 2

(100)

Sénégal

2 2 2

ArD 97264

(100)

2 2 2

Sénégal

ArD 97268

HAZARA

Rapport IP Dakar 1993

slide26

Turkey 2003

Phylogenetic analysis of 46 partial sequences (219 bp) of the S segment of CCHF virus

slide27

Phylogenetic analysis of 46 partial sequences (219 bp) of the S segment of CCHF virus.

Seven major genetic groups.

Strains from the Middle and Far East and from different African regions cluster in clearly separated groups.

TURKEY 2003

Preliminary data: 96-98% homology with strains from the Balck Sea area and Kosovo

KOSOVO AF404507; STAVROPOL AF481802 ; DROSDOV U88412 ; ROSTOV AY277672

Drostein et al, J Clin Microbiol 2002, 40 1122

slide28

National Reference Center - WHO Collaborating Centre for

Arboviruses and Viral Haemorrhagic Fevers, Institut Pasteur, Lyon

Marie-Claude Georges

Isabelle Schuffenecker

Ingrid Marendat

Séverine Murri

Hervé Zeller

BSL 3

BSL 4

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