Diferenciace Diferenciace vakcinačních a vakcinačních a divokých kmenů divokých kmenů varicella-zoster viru varicella-zoster viru (VZV) pomocí (VZV) pomocí molekulárně molekulárně biologických metod biologických metod RNDr. RNDr. Vanda Bo Vanda Bo štíková, Ph.D. štíková, Ph.D.
Diferenciace vakcinačních a divokých kmenů varicella - zoster viru (VZV) pomocí molekulárně biologických metod. RNDr. Vanda Bo štíková , Ph.D. Fylogenetick á analýza založená na genotypizaci celých sekvencí VZV. Loparev et al., J. Virol., Vol. 81, 2007. (rubella). - PowerPoint PPT Presentation
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Diferenciace Diferenciace vakcinačních a vakcinačních a
divokých kmenů divokých kmenů varicella-zoster viru varicella-zoster viru
Fylogenetická analýza založená na genotypizaci celých sekvencí VZV
Loparev et al., J. Virol., Vol. 81, 2007
(rubella)
Loparev et al., J. Virol., Vol. 78, 2004
Globální distribuce VZV - prevalence 70 % a více ve vyznačených oblastech
(2011 - E1, E2, M1 – M4, J)
VZV Oka live-attenuated vaccine developed 1974 in Japan by Takahashi and others by extensive passage of a clinical isolate named “Oka” through hu and non-hu cells in vitro
A series of clinical trials (11 000 healthy adults and children) demonstrated the safety and efficacy of the vaccine in the USA in 1995: approx. 85% of individuals had complete protection
Immunity persists for at least 20 yearsHealthy People 2020 goals 2010 90.51% VZV USA
Varicella in Vaccinated Varicella in Vaccinated PersonsPersons
Should we worry about it?Should we worry about it?
Gomi et al., J. Virol., Vol. 76, 2002
VZV
Genotyping
Pst -
Bgl +
Pst -
Bgl +
Pst +
Bgl -
Msp +
Sma +
Msp -
Sma -
Msp -
Sma -
Pst +
Bgl +
Msp -
Sma -
VaccineWildtype
Japanese
Wildtype
USWildtype
VZVGenotyping
Pst -
Bgl +
Pst +/-
Bgl +
Pst +
Bgl +
Msp +
Sma +
Msp -
Sma -
Msp -
Sma -
Pst +
Bgl -
Msp -
Sma -
OKAvWildtypeJapan
Wildtype20% USAWAustraliaAfrica,Asia
WildtypeEurope,EAustralia,80% US
Pst-,Bgl- reported from Australia
Preliminary genotyping:
The implementation of routine universal varicella
vaccination in the US created a need to distinguish
wild-type infections from vaccine injuries.
Our lab developed LightCycler real-time PCR hybridization probe-based assays to detect VZV(PCR) and genotype VZV strains based on the presence or absence of a BglI, PstI, or MspI restriction sites within open reading frames (ORFs) 54, 38, and 62, respectively.
Pst+ Bgl+Pst-Bgl+=OKAv
Pst+Bgl-
Pst+Bgl-
Pst+Bgl-
Pst+Bgl+
Pst+Bgl+
Pst+Bgl+
Pst+Bgl+
20% Pst+Bgl+
Pst-Bgl+
Pst+Bgl-
Pst+Bgl-
37%
The differences in the melting temperature of the Sma I ORF62 PCR product discriminate Oka vaccine and wt VZV strains.
VZVgenotyp ORF 22 :
4 SNPs #37902,38055,38081,38177
European Clade A or D
Mosaic Clade CM1M2M3M4
JapaneseClade B
+
ORF 50/ORF 21#87841/#3725
Clade A (E1)Genotype like DumasC/T
Clade D (E2)T/C
Loparev et al., Virology, 2008
Virology. 2009 Jan 20;383(2):216-25. Epub 2008 Nov 20.
Distribution of varicella-zoster virus (VZV) wild-type genotypes in northern and southern Europe: evidence for
high conservation of circulating genotypes.
Loparev VN, Rubtcova EN, Bostik V, Tzaneva V, Sauerbrei A, Robo A, Sattler-Dornbacher E, Hanovcova I, Stepanova V, Splino
M, Eremin V, Koskiniemi M, Vankova OE, Schmid DS.
Marlene Deleon-CarnesMarlene Deleon-Carnes Antonio GonzalezAntonio Gonzalez Myron LevinMyron Levin Stephanie LiffickStephanie Liffick Vladimir LoparevVladimir Loparev Stanislav PlíšekStanislav Plíšek Miloslav SalavecMiloslav Salavec Scott D. Schmid Scott D. Schmid