Are plasma derivatives Are plasma derivatives sources for parvovirus B19 sources for parvovirus B19

transmission?transmission?

Manal M. BaddourManal M. BaddourAssociate Professor of Microbiology Associate Professor of Microbiology

King Saud UniversityKing Saud University

Heegaard and Brown, 2002

Parvovirus B19 is an erythrovirus and Parvovirus B19 is an erythrovirus and was until recently the only known was until recently the only known member of the member of the Parvoviridae Parvoviridae family of family of viruses causing disease in humans. viruses causing disease in humans.

D e n so v irus

Ite ra v irus

B re v id e n so v irus

P e fun d en sov irus

D e n sov irin ae(S u b fa m ily)

M V M

C a n ine p a rvo v irusC P V

P o rc ine p a rvo v irusP P V

Parvovirus

A A V s-1 -?

G o ose pa rvov irusG P V

A v ia n A A VA A A V

Dependovirus

P a rvov irus B 19B 1 9V

S im ian p a rvo v irusS P V

P m P V

Erythrovirus

B o v in e pa rvov irusB P V

M inute v iru s of canine sM V C

H u m an b o ca v irus

Bocavirus

A le u tio n d ise a se(m in ks )

P a rv4 /P a rv5

Am dovirus

Parvovirinae(S u b fa m ily)

Parvoviridae(F a m ily )

Parvovirus B19 (B19) was discovered Parvovirus B19 (B19) was discovered fortuitously in 1973 in the United Kingdom fortuitously in 1973 in the United Kingdom by Cossart, who was looking for the by Cossart, who was looking for the hepatitis B virus in blood from “healthy” hepatitis B virus in blood from “healthy”

donors.donors. Cossart et al, 1975Cossart et al, 1975

Slapped CheeksSlapped Cheeks

Slapped CheeksSlapped Cheeks

Lacey Rash of the Dorsum of the HandsLacey Rash of the Dorsum of the Hands

Gloves and socks rashGloves and socks rash

Gloves and socks rashGloves and socks rash

hydropshydrops

Ascites, hepatomegaly, cardiomegaly Ascites, hepatomegaly, cardiomegaly

ThalassemiaThalassemia

ElliptocytosisElliptocytosis

The virus selectively targets early erythroid The virus selectively targets early erythroid precursors, causing anemia. precursors, causing anemia.

The cellular receptor is the blood group P The cellular receptor is the blood group P antigen antigen

Brown et al., 1993Brown et al., 1993

After infection, the incubation period ranges from After infection, the incubation period ranges from 6 to 18 days. 6 to 18 days.

Viremia in early infection occurs with extremely Viremia in early infection occurs with extremely high titers (up to 10high titers (up to 101111 to 10 to 101313 genome genome equivalents/ml). equivalents/ml).

In immunocompetent persons, viraemia is In immunocompetent persons, viraemia is usually cleared a few weeks after infection. usually cleared a few weeks after infection.

In immunocompromised patients, the virus can In immunocompromised patients, the virus can cause a persistent infection for several months cause a persistent infection for several months and years mainly due to impairment of B cell and years mainly due to impairment of B cell function.function.

Siegl and Cassinotti, 1998Siegl and Cassinotti, 1998

The frequency of B19-DNA blood donors has The frequency of B19-DNA blood donors has been estimated at rates of 1:167 to 1:35,000 been estimated at rates of 1:167 to 1:35,000

Heegaard & Heegaard &

Brown,2002Brown,2002

GlobulinGlobulin

AlbuminAlbumin

Clotting factorsClotting factors

High seroconversion rates and some High seroconversion rates and some cases of symptomatic illness have been cases of symptomatic illness have been due to blood products (eg immune due to blood products (eg immune globulin, clotting factors) prepared from globulin, clotting factors) prepared from B19-containing plasma pools with reports B19-containing plasma pools with reports of prominent B19 infection and worsening of prominent B19 infection and worsening the patient’s condition as a resultthe patient’s condition as a result

Hayakawa et al, 2002; Yee et al, 1995;Hayakawa et al, 2002; Yee et al, 1995;

Zanella et al 1995; Mortimer et al, 1983Zanella et al 1995; Mortimer et al, 1983

Azzi et al, 1992; Yee et al 1996

Factor 8

Blood clot

Filtration column

Aim of This StudyAim of This Study

To determine the frequency of parvovirus To determine the frequency of parvovirus B19 DNA contamination in a range of B19 DNA contamination in a range of plasma derivatives commercially available plasma derivatives commercially available in the local market for clinical use.in the local market for clinical use.

MaterialMaterial

Samples from three batches of each of the Samples from three batches of each of the following types of products from 2 companies following types of products from 2 companies were tested for parvovirus DNA by nested PCR:were tested for parvovirus DNA by nested PCR:

1- human albumin1- human albumin 2- human coagulation factor VIII2- human coagulation factor VIII 3- human coagulation factor IX (one company)3- human coagulation factor IX (one company) 4- human normal IV immunoglobulins 4- human normal IV immunoglobulins Serum from a patient previously diagnosed to be Serum from a patient previously diagnosed to be

positive for parvovirus B19 DNA by PCR was positive for parvovirus B19 DNA by PCR was used as a control.used as a control.

MethodsMethods DNA from each of these products was DNA from each of these products was

extracted in triplicates from 200µl aliquots extracted in triplicates from 200µl aliquots of the tested products using the of the tested products using the QIAamp QIAamp DNA Blood MinikitDNA Blood Minikit

Nested PCR using primers Nested PCR using primers • A1 5´-AATGAAAACTTTCCATTTAATGA-3´ A1 5´-AATGAAAACTTTCCATTTAATGA-3´ • B1 5´-TCCTGAACTGGTCCCGGGGATGG-3´ B1 5´-TCCTGAACTGGTCCCGGGGATGG-3´ 590 bp590 bp• A2 5´-GCGTGGAAGTGTAGCTGTGCC-3´ A2 5´-GCGTGGAAGTGTAGCTGTGCC-3´ • B2 5´-CAGAGCTTTCACCACCACTGC-3´B2 5´-CAGAGCTTTCACCACCACTGC-3´355 bp 355 bp

Frickhofen and Young, 1989

Carriere et al, 1993

Carriere et al, 1993

ResultsResults

A representative sample of each type A representative sample of each type of product was spiked with parvovirus of product was spiked with parvovirus B19 DNA (extracted from serum of a B19 DNA (extracted from serum of a previously determined B19 positive previously determined B19 positive patient) and subjected to the same patient) and subjected to the same procedures to test for the efficiency of procedures to test for the efficiency of the technique.the technique.

Results, Results, cont.cont.

Results, Results, cont.cont.

Lanes 1 2 3 4 5 6 7 8 9

Results, Results, cont.cont.

Donor Plasma Donation Center

Begin Manufacturing

Complete Manufacturing

Patient

Safety Step 9:Post-Marketing

Surveillance

Safety Step 1:Donor

Screening

Safety Step 2:Testing Donation

Safety Step 3:Inventory Hold and Lookback

Safety Step 4:Plasma Pool

Testing

Safety Step 7:Virus Inactivation

Virus/TSE Removal

Safety Step 5 & 6:Quality

AssuranceGMP

Safety Step 8:GCP

Packaging Guidance

Manufacturing SafeguardsManufacturing SafeguardsPathogen Safety is a comprehensive approach with effective redundant measures that provide a high margin of safety

The Plasma Protein Therapeutics Association

Pre-implementation

Implementation of Parvovirus B19 NAT Testing

PROCESSVIRAL REDUCTION

PROCESSVIRAL REDUCTION

No Testing UnknownMargin ofSafety

IncreasedMargin ofSafety

Reduction in Parvovirus B19 Viral LoadsReduction in Parvovirus B19 Viral Loads

101- 109

IU/mL

Production Pool

<105

IU/mL

Production PoolMini-pool Testing96-1200 donation samples

High Titer Plasma

ConclusionsConclusions It is true that results obtained from the present It is true that results obtained from the present

study seem reassuring, probably due to study seem reassuring, probably due to implementation of strict screening of blood implementation of strict screening of blood donors or plasma pools for parvovirus B19 DNA donors or plasma pools for parvovirus B19 DNA by the plasma products' manufacturing by the plasma products' manufacturing companies. companies.

Nevertheless, continuous monitoring of blood Nevertheless, continuous monitoring of blood products should be a routine procedure to products should be a routine procedure to ensure safety of such products for recipients ensure safety of such products for recipients especially those at risk to avoid the hazardous especially those at risk to avoid the hazardous possibilities of serious infections.possibilities of serious infections.

Conclusions, Conclusions, cont.cont.

Human bocavirus HBoVHuman bocavirus HBoV B 19 genotypes 2 (A6) & 3 (V9)B 19 genotypes 2 (A6) & 3 (V9)

Thank YouThank You