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No association between human parvovirus B19 infection and Sjögren’s syndrome
  1. R De Stefano1,
  2. S Manganelli1,
  3. E Frati1,
  4. E Selvi1,
  5. A Azzi2,
  6. K Zakrzewska2,
  7. R Marcolongo1
  1. 1Institute of Rheumatology, University of Siena, Italy
  2. 2Department of Microbiology, University of Florence, Italy
  1. Correspondence to:
    Dr S Manganelli, Institute of Rheumatology, University of Siena, 53100 Siena, Italy; s.manganelli{at}

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The association of human parvovirus B19 (HPVB19) infection with autoimmune disease, including systemic lupus erythematosus, rheumatoid arthritis, polymyositis, and vasculitis, has been suggested, although the exact relationship between the infection and these disorders has not been fully elucidated.1,2 A recent report showed serological evidence of past B19 infection associated with the presence of cytopenia in patients with primary Sjögren’s syndrome (SS).3 To gain more information about the aetiopathogenetic role of HPVB19 for this disease, we evaluated the presence of the viral genome in minor salivary glands from patients with primary SS.

We studied 10 women with SS (mean (SD) age 45 (9) years) and 10 healthy controls matched for age (43 (6) years) and sex. SS was diagnosed according to European criteria.4 Each subject taking part in the study underwent minor salivary gland 6 mm punch biopsy under local anaesthesia. Histological evaluation of biopsy samples was carried out according to Chisholm and Mason’s classification.5 They were also analysed for the presence of DNA sequence coding for the HPVB19 non-structural protein (NS1) amplified by nested polymerase chain reaction (PCR) as a marker of infection. The outer primer pairs were P1 and P6, corresponding to nucleotides 1399–1422 and 1682–1659. In the second amplification, the P2 and P5 inner nested primer pairs, corresponding to nucleotides 1498–1525 and 1660–1576, were used. The 103 base pair (bp) diagnostic fragment was subsequently detected by ethidium bromide staining after agarose gel electrophoresis. Each sample was tested in duplicate. A 10−9 dilution of a reference serum containing about 10–100 HPVB19 genome copies was used as positive control. Negative water controls were extracted concomitantly with the diagnostic specimens in order to monitor possible contamination during the extraction step. Additional negative controls were included in each PCR run to eliminate the possibility of carryover contamination. A 268 bp fragment of the β-globin gene was amplified using primers PC04 and GH20 as a test for the absence of Taq DNA polymerase inhibitors and to estimate the quantity of DNA extracted from each minor salivary gland. A serial 10-fold dilution of DNA extracted from a known number of Hep-2 cells was used as positive control. Negative controls were simultaneously extracted water samples. PCR products were then analysed by agarose gel electrophoresis.

Blood samples from each patient were tested for the presence of anti-B19 IgM and IgG using a commercially available enzyme linked immunosorbent assay (ELISA) (Pantec, Torino-Italy).

All minor salivary gland samples of patients with SS were rated as grade III or IV according to Chisholm-Mason’s classification. In the control group, only three subjects were rated as grade I or II (subjects 13, 14, and 17, table 1).

The DNA sequence coding for NS1 of HPVB19 was found in a salivary gland specimen from one case of SS (patient 2) and from one control subject (subject 19). Both cases showed a high titre of anti-B19 IgG antibodies and the absence of specific IgM antibodies. In the patient with SS (patient 2) the presence of IgG B19 antibodies was associated with more than one focus score, whereas in the control subject (subject 19) the presence of DNAPVB19 was not associated with lymphocytic infiltrate. Anti-B19 IgG antibodies, but no anti-B19 IgM antibodies, were detected in three other subjects, including one with SS (patient 6) and two controls (subjects 12 and 17). The results of the study showed that the prevalence of past B19 infection in patients with primary SS was similar to that of the control group. Furthermore, none of the patients with SS showed serological markers of recent infection from HPVB19.

B19DNA can also be found, and can persist, in the salivary glands without inevitably inducing a lymphocytic infiltrate in this tissue. Our results suggest that B19 constituents may also be found in salivary gland tissue. However, the presence of viral DNA in the salivary glands of patients with SS appears to be incidental, and it does not support an association between SS and HPVB19 infection.

Table 1

Histological and serological features in patients with primary SS and controls. 1–10 are patients, 11–20 controls


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