LevelLevel of viraemia of viraemia and genotype characteristics and genotype characteristics

of Parvo B19 of Parvo B19 in blood donors and patientsin blood donors and patients

Brojer E, Grabarczyk P

Depatment of ImmuhematologyInstitute of Hematology and Blood Transfusion, Warsaw,

Poland

Parvo B19 testing in IHBTParvo B19 testing in IHBT

MATERIALSMATERIALS plasma pools of 24 donations for anti-D and anti-HBsplasma pools of 24 donations for anti-D and anti-HBs plasma from patients with clinical symptomsplasma from patients with clinical symptoms

METHODSMETHODS Quantitative real-time PCRQuantitative real-time PCR

Qiagen/Nuclisens Extractor + home made (Aberham C, 2001).Qiagen/Nuclisens Extractor + home made (Aberham C, 2001). Qiagen + RealArt Parvo (B19 PE PCR Kit, Artus Biotech)Qiagen + RealArt Parvo (B19 PE PCR Kit, Artus Biotech)

quantitative anti-B19 IgG and IgMquantitative anti-B19 IgG and IgM recomWell Parvowirus B19 IgG and IgM (MIKROGEN GmbH)recomWell Parvowirus B19 IgG and IgM (MIKROGEN GmbH)

genotypinggenotyping sequencing of NS1/VP1 (Servant A, 2002) sequencing of NS1/VP1 (Servant A, 2002)

Identification of B19 positive donations Identification of B19 positive donations 2004 -2005 (IHiT)2004 -2005 (IHiT)

Positives Donations tested (n) (N) (%)

3432 6 0,17

Parvo B19 infection characteristics

Clinical features Characteristics of B19 infection molecular

markers

DNA B19 (IU/ml)

Reason for B19 testing/Clinical manifestation

Imm

un

oco

mp

eten

ce

genotype

Max. Min.

Period of DNA B19

observation

Healthy plasma donor for anti-HBs Yes 1 2 x 105 1 x102 3 yrs and 2 mth

Plasma donor for anti-D Yes 1 1x103 Ind

Plasma donor for anti-D Yes 1 2x103 Ind

Healthy HSC donor for nr3 (sister) Yes 1 2x104 104 3 months

pure red cell aplasia after kidney transplantation No 2 6,5x109 7x103 1 yr and 6 mth

pure red cell aplasia after allo HSCT No 1 1,2x109 8,5x103 1 yr and 1 mth

Pancytopenic stage during chemotherapy of ALL No 1 5,2x107 3,2x103 3 months

Pancytopenic stage during cancer treatment No 5x102 Ind

BM aplasia, anti-B19 IgG(+) i IgM(+) Yes Nd 8,8x104 5x102 8 months

Rush and subsequently pure red cell aplasia Yes Nd 5,6x107 9,8x104 2 weeks

Mother of nr 7, rush Yes 1 2,1x105 Ind

Rush during pregnancy, fifth disease in 3 years old daughter Yes Nd 3,1x107 Ind

Mother of newborn-HGB 1g/dl, ASPAT, ALAT 900 IU/ml Yes nd 3 x103 Ind

Mother after fetus loss, enlarged nodes Yes 1,6x105 8x102 3 months

After liver transplantation No 1,2x106 Ind

Parvo B19 genotype 2 infection Parvo B19 genotype 2 infection after kidney transplantationafter kidney transplantation

BD, (dr A.Wieczorek - Transplantation Ward, Poznań)BD, (dr A.Wieczorek - Transplantation Ward, Poznań)•11.2003 kidney transplantation11.2003 kidney transplantation•02.2004 red cell aplasia02.2004 red cell aplasia•18.03.2004 DNA B19 (+)18.03.2004 DNA B19 (+)•the and of the and of March 2 March 2004 immunosupresion treatment switch004 immunosupresion treatment switch•April April 2004 clinical parameters in the normal range2004 clinical parameters in the normal range•May May 2004 – 2004 – January January 2005 - pregnancy (no clinical sympthoms)2005 - pregnancy (no clinical sympthoms)

29 lipca6 lipca6 maja14 kw ietnia18 marca 3 w rześnia08-paź 21-paź 17-lis 08 grudź05-sty 22 luty 28-kw i 28-cze1.0E+00

1.0E+01

1.0E+02

1.0E+03

1.0E+04

1.0E+05

1.0E+06

1.0E+07

1.0E+08

1.0E+09

1.0E+10

1 2 3 4 5 6 7 8 9 10 11 12 13 14

DN

A B

19 (

IU/m

l)

0

100

200

300

400

500

600

Ig a

nty

B19

(IU

/ml)

DNA B19 B19 IgM B19 IgG

immunosupresion treatmentswitch

pregnancy

B19 genotype 1 infection in symptomless blood donorB19 genotype 1 infection in symptomless blood donor

• blood donor (PK), donor of plasma for anti-HBs production (RCKiK Poznań)blood donor (PK), donor of plasma for anti-HBs production (RCKiK Poznań)•38 years old, 3 children,38 years old, 3 children,• throughout observation period (3 years and 3 months) no clinical symptoms: Hb, Ht, PLT, RBC, throughout observation period (3 years and 3 months) no clinical symptoms: Hb, Ht, PLT, RBC, WBC, ALT in normal range.WBC, ALT in normal range.

1

10

100

1000

10000

100000

1000000

days

DN

A B

19 (

IU/m

l)

0

50

100

150

200

250

anty

B19

(IU

/ml)

DNA B19

B19 IgM

B19 IgG

20.09.28-cze28-kw i22 luty05-sty08 grudź17-lis21-paź08-paź3 w rześnia18 marca 14 kw ietnia6 maja 6 lipca 29 lipca

1,0E+00

1,0E+01

1,0E+02

1,0E+03

1,0E+04

1,0E+05

1,0E+06

1,0E+07

1,0E+08

1,0E+09

1,0E+10

DN

A B

19 (

IU/m

l)

0

100

200

300

400

500

600

Ig a

nty

B19 (

IU/m

l)

DNA B19 B19 IgM B19 IgG

change of immunosupression

Patient 2red cell aplasiaAnaemia

after allo BMT

due to the parvo B19 genotype 1 infection

transmitted by BM donor

•B19 DNA detected in pretransplant DNA sample of the donor was 100% homologous to B19 DNA of the patient

Comparison of genotyp 1 and 2 sequences Comparison of genotyp 1 and 2 sequences from patients with aplastic anaemiafrom patients with aplastic anaemia

Observations and discussionObservations and discussion

GGenotype 1 and genotype 2 were identified in enotype 1 and genotype 2 were identified in clinical speciments from patients with similar clinical speciments from patients with similar symptoms symptoms

Should NAT for plasma screenig identify not only genotype Should NAT for plasma screenig identify not only genotype 1 but also genotype 2? 1 but also genotype 2? ( and genotype 3, PARV 4( and genotype 3, PARV 4, PARV5, PARV5))

B19 transmission by bone marrB19 transmission by bone marroow cellsw cells Should BM/SC or other tissue or organ donors (especially Should BM/SC or other tissue or organ donors (especially

young) be tested for B19 markers?young) be tested for B19 markers?

Correlation of B19 viral load with clinical Correlation of B19 viral load with clinical manifestations – manifestations –

high viremia during syptoms in most but not all patientshigh viremia during syptoms in most but not all patients