Crimean-Congo Haemorrhagic Fever
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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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Crimean-Congo Haemorrhagic Fever DIAGNOSIS

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Crimean congo haemorrhagic fever diagnosis

Crimean-Congo Haemorrhagic Fever

DIAGNOSIS

Hervé Zeller

National Reference Center - WHO Collaborating Centre for

Arboviruses and Viral Haemorrhagic Fevers, Institut Pasteur, Lyon


Crimean congo haemorrhagic fever diagnosis

Flaviviridae

(dengue, yellow fever,

Groupe TBE)

Viral Haemorrhagic Fevers


Crimean congo haemorrhagic fever diagnosis

Flaviviridae

(dengue, yellow fever,

Groupe TBE)

Viral Haemorrhagic Fevers

Arenaviridae

(Lassa, Junin, Machupo, Guanarito)


Crimean congo haemorrhagic fever diagnosis

Flaviviridae

(dengue, yellow fever,

Groupe TBE)

Viral Haemorrhagic Fevers

Arenaviridae

(Lassa, Junin, Machupo, Guanarito)

Filoviridae

(Ebola, Marburg)


Crimean congo haemorrhagic fever diagnosis

Flaviviridae

(dengue, yellow fever,

Groupe TBE)

Viral Haemorrhagic Fevers

Arenaviridae

(Lassa, Junin, Machupo, Guanarito)

Envelopped

RNA viruses

Bunyaviridae

(CCHF, RVF,

Hantaviruses)

Filoviridae

(Ebola, Marburg)


Crimean congo haemorrhagic fever diagnosis

FamilyGenus VIRUS DISTRIBUTION

FlaviviridaeFlavivirusYellow FeverAfrica South America

Dengue 1,2,3,4. Tropical areas

Omsk HFRussia

AlkhurmaSaudi Arabia

Kyasanur Forest HFIndia

BunyaviridaePhlebovirusRift Valley FeverAfrica, Saudi Arabia

NairovirusCrimean-Congo HF Africa, Eurasia

HantavirusHantan Dobrava Puumala Eurasia

Sin Nombre, AndesAmericas


Crimean congo haemorrhagic fever diagnosis

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


Crimean congo haemorrhagic fever diagnosis

  • 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


  • Crimean congo haemorrhagic fever diagnosis

    Nairovirus structure

    G1

    M

    S

    N

    G2

    L

    L

    10 nm


    Crimean congo haemorrhagic fever diagnosis

    CCHFNairovirus genome

    _____________________________________________

    SegmentNucleotidesAmino acids Protein

    _____________________________________________

    S1659-1712442-482 N

    M48881551 G1 G2 NSm?

    L122554036 L?

    _____________________________________________


    Crimean congo haemorrhagic fever diagnosis

    VHF SUSPECT CASE


    Crimean congo haemorrhagic fever diagnosis

    VHF SUSPECT CASE

    Malaria – Hepatitis – Typhoidis – Toxicosis

    Septicemia – Leptospirosis

    Rickettsiosis…


    Crimean congo haemorrhagic fever diagnosis

    VHF SUSPECT CASE

    Malaria – Hepatitis – Typhoidis – Toxicosis

    Septicemia – Leptospirosis

    Rickettsiosis…

    Epidemiological data, risk evaluation

    biological analysis, differential diagnostic


    Crimean congo haemorrhagic fever diagnosis

    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


    Crimean congo haemorrhagic fever diagnosis

    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…


    Crimean congo haemorrhagic fever diagnosis

    Primers for RT-PCR on S segment

    From J. Smith, 1990


    Crimean congo haemorrhagic fever diagnosis

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

    From Burt et al,

    J Virol Methods 1998, 70:129-137


    Crimean congo haemorrhagic fever diagnosis

    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


    Crimean congo haemorrhagic fever diagnosis

    Hyalomma sp.

    ticks

    RT-PCR

    Viral isolation


    Crimean congo haemorrhagic fever diagnosis

    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


    Crimean congo haemorrhagic fever diagnosis

    Turkey 2003

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


    Crimean congo haemorrhagic fever diagnosis

    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


    Crimean congo haemorrhagic fever diagnosis

    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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