congo fever

8/7/2019 congo fever

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Crimean-Congo Haemorrhagic Fever

DIAGNOSIS

Herv ZellerNational Reference Center - WHO Collaborating Centre for

Arboviruses and Viral Haemorrhagic Fevers, Institut Pasteur, Lyon


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Viral Haemorrhagic Fevers

Flaviviridae

(dengue, yellow fever,

Groupe TBE)


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Arenaviridae

(Lassa, Junin, Machupo, Guanarito)


Flaviviridae


Groupe TBE)


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Filoviridae

(Ebola, Marburg)

Arenaviridae



Flaviviridae


Groupe TBE)


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Filoviridae

(Ebola, Marburg)

Arenaviridae


Bunyaviridae

(CCHF, RVF,

Hantaviruses)


Flaviviridae


Groupe TBE)

Envelopped

RNA viruses


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Family Genus VIRUS DISTRIBUTION

Flaviviridae Flavivirus Yellow Fever Africa South AmericaDengue 1,2,3,4. Tropical areas

Omsk HF Russia

Alkhurma Saudi Arabia

Kyasanur Forest HF India

Bunyaviridae Phlebovirus Rift Valley Fever Africa, Saudi Arabia

Nairovirus Crimean-Congo HF Africa, Eurasia

Hantavirus Hantan Dobrava Puumala Eurasia

Sin Nombre, Andes Americas


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Hemorrhages are

inconstant :Emerging part of the

iceberg

Most frequentlyasymptomatic infections

+++


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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


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CCHF VIABILITY

SENSITIVITY TO DESINFECTANTS:

sodium hypochlorite 2%, glutaraldehyde 2%,

formaldehyde

SENSITIVE TO DESSICATION

INACTIVATION :

IRRADIATION

UV

TEMPERATURE : 1 hour 60C

not complete inactivation

beta propiolactone 4C

not complete

inactivation


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S M

L

N

G1

G2

10 nm

Nairovirus structure

L


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CCHF Nairovirus genome

_____________________________________________

Segment Nucleotides Amino acids Protein

_____________________________________________

S 1659-1712 442-482 N

M 4888 1551 G1G2 NSm?

L 12255 4036 L?

_____________________________________________


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VHF SUSPECT CASE


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VHF SUSPECT CASE

Malaria Hepatitis Typhoidis Toxicosis

Septicemia Leptospirosis

Rickettsiosis


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VHF SUSPECT CASE

Epidemiological data, risk evaluation

biological analysis, differential diagnostic



Rickettsiosis


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VHF SUSPECT CASE

Contact between clinicians and biologists

Epidemiological data, risk evaluation

biological analysis, differential diagnostic



Rickettsiosis


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CCHF : laboratory dataCCHF : laboratory dataLEUCOPENIA, particularly neutropenia

THROMBOCYTOPENIA

Hematocrite increases early then falls down

ASL, AST levels increases

Proteinuria and hematuria

Mild azotemia, bilirubine increase


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Viral detection: blood specimen

RT-PCR (nested)

Cell culture (Vero E6 cells)

CCHF : laboratory diagnosisCCHF : laboratory diagnosis


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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 diagnosisCCHF : laboratory diagnosis


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CCHF : laboratory diagnosisViremia 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


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viremia

5

IgM

RT-PCR

ELISA IgM IgG IFA

CCHF : viral/antibody kinetics

Viral isolation

0 10

IgG

16

IgM duration: 2-3 months up to 6 months


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Primers for RT-PCR on S segment

100 300 500 700

3'5'

F2 R3

Fragments amplifis: F2 / R3 (536 bp) , F3/R2 (259 bp)

Squences des amorces:

F2 5' TGG ACA CCT TCA CCA AAC TC 3' R3 5' GAC AAA TTC CCT GCA CCA 3'

F3 5' GAA TGT GCA TGG GTT AGC TC 3' R2 5' GAC ATC TTC CCT GCA CCA 3'

670135

Segment S ARN CCHF

F3 R2

290 550

From J. Smith, 1990


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

From Burt et al,

J Virol Methods

1998, 70:129-

137

Day of illness

PCR +

Virus +

PCR +

Virus -

PCR -

Virus +

PCR -

virus - Ab +

Total

tested

3 1 1 1 34 1 1 1 2

5 3 2 3 5

6 5 3 4 3 8 15

7 5 4 8 9

8 1 2 4 7 79 5 1 5 6

10 3 3 3

11 1 2 1 1 5 5

12 1 1 2 2

13 1 4 3 8 814 4 1 3 8 8

15 2 1 3 3

16 2 1 3 3

18 1 1 1

Total 18 34 6 22 65 80


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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

day ofillness

PCR +Virus +

PCR +Virus -

PCR -Virus +

PCR -virus -

IFAAb +

Totaltested

3 1 1

4 1 1

5 1 1

6 4 1 2 5

7 3 1 1 5 5

8 6 1 2 8 9

9 2 2 2

10

11 1 1 1

12 1 1 1

Total 14 1 3 8 19 26


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Hyalomma sp.ticks

RT-PCR

Viral isolation


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DUGBE

AP 92

AnD 15786

ArD 8194

ArTeh 193-3

HD 49199

ArMg 951

C 68031

ArB 604

HD 38562

ArD 39554

ArD 97264

ArD 97268

HAZARA

3 2 1

1 1 1

1 3 1

1 2 3

2 2 4

2 2 3

2 2 3

2 2 2

2 2 2

2 2 2

1 3 1

1 3 1

Grce

Sngal

Sngal

Sngal

Sngal

Mauritanie

Mauritanie

Iran

Madagascar

Chine

Rp. Centrafr.

Burkina Faso

ORIGINEPROFIL

RFLP

(100)(100)

(96)

(100)

(100)

(99)

(100)

(84)(57)

536 pb amplicons of the S fragment of CCHF genome using primers

CSDR3/CSDF2. RFLP with Hinf I, Hae III, and Alu I endonucleases

Rapport IP Dakar1993


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Phylogenetic analysis of 46 partial sequences

(219 bp) of the S segment of CCHF virus

Turkey 2003


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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


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National Reference Center - WHO Collaborating Centre for

Arboviruses and Viral Haemorrhagic Fevers, Institut Pasteur, Lyon

Marie-Claude Georges

Isabelle Schuffenecker

Ingrid Marendat

Sverine Murri

Herv Zeller

BSL 4

BSL 3

congo fever

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