Parvovirus B19 infection in Tunisian patients with sickle-cell anemia and acute erythroblastopenia

Abstract

Background: Human parvovirus B19 is the etiologic agent of erythema infectiosum in children. It is also associated with other clinical manifestations in different target groups. Patients with chronic hemolytic anemia are at high risk of developing acute erythroblastopenia following infection by the virus. They usually become highly viremic and pose an increased risk of virus transmission. Close monitoring of such high risk groups is required for epidemiologic surveillance and disease prevention activities. Here we report a molecular epidemiological study on B19 virus infection in Tunisian patients with chronic hemolytic anemia.

Methods: This study was conducted on 92 young chronic hemolytic anemia patients who attended the same ward at the National Bone Marrow Transplantation Center of Tunis and 46 controls from a different hospital. Screening for IgM and IgG anti-B19 antibodies was performed using commercially available enzyme immunoassays and B19 DNA was detected by nested PCR in the overlapping VP1/VP2 region. DNA was sequenced using dideoxy-terminator cycle sequencing technology.

Results: Anti-parvovirus B19 IgG antibodies were detected in 26 of 46 sickle-cell anemia patients, 18 of 46 beta-thalassemia and 7 of 46 controls. Anti-parvovirus B19 IgM antibodies were detected only in 4 of the sickle-cell anemia patients: two siblings and two unrelated who presented with acute erythroblastopenia at the time of blood collection for this study and had no history of past transfusion. B19 DNA was detected only in sera of these four patients and the corresponding 288 bp nested DNA amplicons were sequenced. The sequences obtained were all identical and phylogenetic analysis showed that they belonged to a new B19 virus strain of Genotype1.

Conclusion: A new parvovirus B19 strain of genotype1 was detected in four Tunisian patients with sickle-cell anemia. Virus transmission appeared to be nosocomial and resulted in acute erythroblastopenia in the four patients. The possibility of independent transmission of this B19 variant to the patients is unlikely in light of the present epidemiological data. However this possibility cannot be ruled out because of the low genetic variability of the virus.

Figure 1

Agarose gel electrophoresis pattern of parvovirus B19 DNA amplicons. (A) 398 bp DNA bands obtained in the first round of PCR; (B) 288 bp DNA bands obtained in the second round of PCR. M: molecular weight marker (100 bp ladder, Invitrogen). P: positive control; lanes 1 to 4: amplicons of sera from patients 1 to 4, respectively.

Figure 2

Alignment of the partial DNA sequence of the four B19 isolates with reference strains. The partial VP1/VP2 DNA sequence (R) representing the four B19 virus isolates R1 to R4 was aligned with representative sequences of the three human parvovirus genotypes retrieved from GenBank. Accession numbers were as follows. Genotype 1: OsFr [GenBank: DQ225150.1]; Kati 4 [GenBank: AF161226.1]; AnTo [GenBank: DQ225151.1]; SN807 [GenBank: DQ225149.1]; NAN [GenBank: AY504945.1]; J35 [GenBank: AY386330.1]; B19-Au [GenBank: M13178.1]. Genotype 2: LaLi [GenBank: AY044266.1]; Berlin [GenBank: AJ717293.1]; BN31.2 [GenBank: DQ333426.1]; IM-81 [GenBank: AY903437.1]; A6c8 [GenBank: AY064476.1]. Genotype 3: V9 [GenBank: AY345134.1]; D91.1 [GenBank: AY083234.1]; BN30.3 [GenBank: DQ408305.1]; R0748 [GenBank: DQ234779.1].

Figure 3

Phylogenetic tree. The phylogenetic tree of sequence R representing the four B19 virus isolates R1 to R4 was visualized by TreeView following alignment in BioEdit and Clustal W as shown in Figure 2. See legend to Figure 2 for accession numbers.