First international proficiency study on west nile virus molecular detection
This presentation is the property of its rightful owner.
Sponsored Links
1 / 15

First international proficiency study on West Nile virus molecular detection. PowerPoint PPT Presentation


  • 56 Views
  • Uploaded on
  • Presentation posted in: General

West Nile. ROBERT KOCH INSTITUT. First international proficiency study on West Nile virus molecular detection. Source: H. Gelderblom, RKI. Matthias Niedrig , Sonja Linke Christian Drosten Hervé Zeller . WN. WN. WN. WN. WN. WN. WN.

Download Presentation

First international proficiency study on West Nile virus molecular detection.

An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.


- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -

Presentation Transcript


First international proficiency study on west nile virus molecular detection

West Nile

ROBERT KOCH INSTITUT

First international proficiency study on West Nile virus molecular detection.

Source: H. Gelderblom, RKI

Matthias Niedrig, Sonja Linke

Christian Drosten Hervé Zeller

ENIVD 06/06

15:10


Distribution of disease vectors and wn virus in europe

WN

WN

WN

WN

WN

WN

WN

Distribution of disease vectors and WN virus in Europe

Aedes albopictus

Ochlerotatus atropalpus

Ochlerotatus japonicus

source:

Snow & Ramsdale,

Biologist (2002) 49 (2)

Hubalek & Halauzka;

EID, (1999) 5 (5)

ENIVD 05/06

15:10


I mport of suspected or confirmed wn cases into europe

Import of suspected or confirmed WN cases into Europe

eight cases in total

ENIVD 05/06

15:10


Preparation of pcr samples for the e xternal q uality a ssurance eqa

Preparation of PCR samples for the External Quality Assurance (EQA)

  • cell culture supernatants of infected cells with the respective viruses

  • heat treatment for 1h at 56°C and gamma irradiation of 30 kGy

  • check for infectivity by cell cultivation

  • dilute with human plasma, aliquots of 100 µl are frozen at –70°C before freeze-drying for 12 hours

  • samples were distributed by normal mail service

ENIVD 05/06

15:10


Phylogenetic tree of some tested wnv samples and reference strains

Phylogenetic tree of some tested WNV samples and reference strains

Source: Alex Platonov

ENIVD 05/06

15:10

based on the UPGMA method by using 216 nt sequence of 5'-UTR/C gene fragment


Evaluation of the wn pcr eqa

Evaluation of the WN-PCR EQA

ENIVD 05/06

15:10


Evaluation of wn pcr diagnostic by eqa 1

Evaluation of WN PCR diagnostic by EQA (1)

ENIVD 05/06

15:10


Evaluation of the wnv pcr diagnostic by eqa 2

Evaluation of the WNV-PCR diagnostic by EQA (2)

ENIVD 05/06

15:10


Evaluation of wnv pcr diagnostic by eqa 3

Probit analysis of the fractions of laboratories achieving positive results (y-axis) in relation to the virus RNA concentration in a given positive sample (x-axis). Data points represent individual samples in the proficiency test panel. The thick line is the regression line calculated on the basis of a probit model (dose-response curve), the thin lines are 95% confidence

lineage 2

Fraction of laboratories

Log RNA copies / ml

Evaluation of WNV PCR diagnostic by EQA (3)

lineage 1

ENIVD 05/06

15:10


Results wnv pcr eqa

Results WNV PCR EQA

  • Does the PCR method used has an influence on the quality of the results?n= 28; 15 real time PCR, 11 RT-PCR, 2 commercial

  • Do other factors influence the PCR results ?genome preparation: n= 28, 19 Qiagen, 5 Roche, 4 other

There is no significant correlation between the PCR method and the quality of the results.

There is no significant correlation between the genome preparation method and the quality of the results.

ENIVD 05/06

15:10


First international proficiency study on west nile virus molecular detection

- Conclusions - WNV PCR EQA

  • using cell culture supernatants, diluting samples with human plasma and freeze-drying before shipping is useful for sample preparation- gamma-irradiation is a suitable method for inactivation of samples

  • 7 laboratories showed excellent results (sensitivity of 100%), whereas 10 laboratories that were less sensitive need to improve their methods

  • PCR methods for detection of WNV should also detect lineage 2 which is now present in Europe.

  • 4 laboratories create false positive results

ENIVD 05/06

15:10


Eqa studies performed by the enivd between 1999 and 2005

EQA studies performed by the ENIVD between 1999 and 2005

ENIVD 05/06

15:10


What s coming next

What‘s coming next

ENIVD 05/06

15:10


Acknowledgement

Acknowledgement:

H. Bin, Central Virol. Laboratory, Sheba Medical Center, Tel Hashomer, Israel

D. J. Gubler, CDC, Fort Collins, USA

A. Platonov, Central Research Institute of Epidemiology, Moscow, Russia

H. Zeller, Jean Mérieux-INSERM P4 laboratory, Institut Pasteur, Lyon, France

C. Drosten, Bernhard Nocht Institut, Hamburg, Germany

Stephan W. Aberle, Franz Heinz, Medical University of Vienna, Austria

Parts of this project were supported by theEuropean Network for diagnostics of "Imported" viral Diseases (ENIVD) presently funded by DG-SANCO of the European Community under the program „AIDS and other communicable diseases“

thanks for your attention

ENIVD 05/06

15:10


E uropean n etwork for diagnostics of i mported viral d iseases enivd

European Network for diagnostics of "Imported" viral Diseases (ENIVD)

for further information:

www.enivd.de

www.enivd.net

www.enivd.org

ENIVD 06/06

15:10


  • Login