Clinical interpretation of antineutrophil cytoplasmic antibodies: parvovirus B19 infection as a pitfall.
ABSTRACT While antibodies directed against proteinase 3 (PR3-ANCA) and myeloperoxidase (MPO-ANCA) have a high specificity for the diagnosis of systemic vasculitis, they may also be found as an epiphenomenon of acute viral infection.
To investigate whether positive ANCA test results may be a common feature of acute parvovirus B19 infection.
Sera were analysed from 1242 patients from a rheumatology outpatient clinic for reactivity with parvovirus B19 and EBV antibodies. They were tested for the presence of PR3-ANCA and MPO-ANCA, along with sera known to contain IgM antibodies to these viruses obtained from among 41,366 samples submitted for virological screening.
ANCA were found in 10% (5/50) of the sera positive for IgM antibodies to parvovirus and in 3/51 sera containing IgM antibodies to EBV. Three of six patients with arthritis and concomitant parvovirus infection were found positive for PR3-ANCA and two were found positive for MPO-ANCA. All six patients tested negative for ANCA after six months of follow up.
PR3-ANCA and MPO-ANCA may occur transiently in patients with acute B19 infection or infectious mononucleosis, highlighting the importance of repeated antibody tests in oligosymptomatic clinical conditions in which systemic autoimmune disease is suspected.
- [show abstract] [hide abstract]
ABSTRACT: The most effective treatment of chronic hepatitis B virus (HBV) infection is interferon alfa (IFN-alpha), a potentially severe side effect of which is the induction of autoimmunity. To assess whether IFN-alpha causes clinical or serological autoimmune manifestations, we studied 61 children randomized to receive 5 MU/m2 of IFN-alpha three times per week for 12 weeks, with or without steroid priming or no treatment. Autoantibodies to antinuclei (ANA), smooth muscle (SMA), gastric parietal cell (GPC), liver kidney microsomal type 1, mitochondrial, liver cytosolic antigen, thyroid microsomal, and thyroid globulin were detected by standard techniques. Over a median of 4 years (range, 1-5 years) from randomization, no clinical signs of autoimmunity were observed. Autoantibody positivity for nuclei, smooth muscle, and/or gastric parietal cells was observed on at least one occasion in 42 of 61 children (69%), with no overall difference in the prevalence between patients treated with interferon alone (19 of 24 [79%]), steroids plus interferon (13 of 20 [65%]), or untreated controls (10 of 17 [59%]). There was also no difference in the autoantibody prevalence before, during, and at follow-up after cessation of treatment in both interferon-treated and interferon-untreated patients. Autoantibodies are common in chronic HBV infection, and their prevalence is uninfluenced by IFN-alpha.Hepatology 10/1996; 24(3):520-3. · 12.00 Impact Factor
- [show abstract] [hide abstract]
ABSTRACT: The presence of anti-endothelial cells (AECA), smooth muscle (SMA), antinuclear (ANA) and antimitochondrial (AMA) autoantibodies, and liver/kidney microsomal antibody type 1 (LKM1) was investigated retrospectively in sera of liver transplant patients and correlated with cytomegalovirus (CMV) infection as determined by the antigenemia test and with the appearance of acute or chronic allograft rejection. Indirect immunofluorescence analysis was carried out in sequential sera from 40 liver transplant patients. Ten out of 23 antigenemia-positive and none of the antigenemia-negative patients developed serum autoantibodies (P = 0.002, Fisher's exact test). Anti-endothelial cell autoantibodies were found in nine cases and SMA in four patients. Antinuclear antibodies were detected in one otherwise autoantibody-negative patient. All but one case of autoantibody positivity were observed in the high antigenemia group (P < 0.0001, Fisher's exact test). In all but one case, autoantibodies were detected in blood during the antigenemia phase and in most cases in coincidence with or after the antigenemia peak. Even though a statistically significant correlation was not found between autoantibody production and the development of acute or chronic allograft rejection, proportionally more acute rejection cases were observed in the autoantibody-positive than in the autoantibody-negative group. It has been speculated that CMV-induced endothelial damage may be a potent antigenic stimulus, which leads to the production of anti-endothelial cells autoantibodies. Anti-endothelial cell autoantibodies may represent not only a marker of cell injury but also contribute to the progression of the inflammatory response leading to the exposure of tissue-privileged self antigens and the induction of other autoantibodies such as SMA.Journal of Medical Virology 01/2002; 66(1):56-62. · 2.37 Impact Factor
Clinical interpretation of antineutrophil cytoplasmic
antibodies: parvovirus B19 infection as a pitfall
J Hermann, U Demel, D Stu ¨nzner, E Daghofer, G Tilz, W Graninger
Ann Rheum Dis 2005;64:641–643. doi: 10.1136/ard.2004.024877
Background: While antibodies directed against proteinase 3
(PR3-ANCA) and myeloperoxidase (MPO-ANCA) have a
high specificity for the diagnosis of systemic vasculitis, they
may also be found as an epiphenomenon of acute viral
Objective: To investigate whether positive ANCA test results
may be a common feature of acute parvovirus B19 infection.
Methods: Sera were analysed from 1242 patients from a
rheumatology outpatient clinic for reactivity with parvovirus
B19 and EBV antibodies. They were tested for the presence of
PR3-ANCA and MPO-ANCA, along with sera known to
contain IgM antibodies to these viruses obtained from among
41 366 samples submitted for virological screening.
Results: ANCA were found in 10% (5/50) of the sera
positive for IgM antibodies to parvovirus and in 3/51 sera
containing IgM antibodies to EBV. Three of six patients with
arthritis and concomitant parvovirus infection were found
positive for PR3-ANCA and two were found positive for
MPO-ANCA. All six patients tested negative for ANCA after
six months of follow up.
Conclusions: PR3-ANCA and MPO-ANCA may occur tran-
siently in patients with acute B19 infection or infectious
mononucleosis, highlighting the importance of repeated
antibody tests in oligosymptomatic clinical conditions in
which systemic autoimmune disease is suspected.
viral infections such as chronic hepatitis and cytomegalovirus
infection may be associated with the production of a variety
of autoantibodies.1 2
(ANCA) with high specificity to proteinase 3 (PR3-ANCA)
and myeloperoxidase (MPO-ANCA) is now an important tool
in the diagnosis of systemic vasculitis.3Nevertheless, ANCA
have also been described in patients with chronic hepatitis C
infection, suggesting that viral infection may induce their
production.4Parvovirus B19 (B19) infection may predispose
to the production of autoantibodies against Ro, Scl-70, and
antiphospholipids.5Molecular mimicry is the most popular
hypothesis to explain autoantibody production in viral
infection, and it proposes that viral antigens that share
homologies with host antigens generate a cross reactive
immune response.6This can lead to diagnostic confusion in
daily clinical practice.
After observing an index case presenting with polyarthritis,
low grade fever, malaise, and positive PR3-ANCA and MPO-
ANCA test results (see below) eventually diagnosed as an
acute parvovirus B19 infection, we investigated whether
espite the low pretest likelihood, rheumatologists are
sometimes tempted to use specific autoantibody tests
to uncover the underlying disease. On the other hand,
positive ANCA test results might be a relatively common
feature of acute B19 infection.
PATIENTS AND METHODS
A 24 year old woman (table 1, patient 1) presented in our
outpatient clinic with polyarthritis, low grade fever, malaise,
and a positive test result for PR3-ANCA and MPO-ANCA. Her
erythrocyte sedimentation rate (ESR) was 10 mm/h, C
reactive protein was normal, and she tested negative for
rheumatoid factor and antinuclear antibodies (ANA). As
examination of the upper respiratory tract, chest x ray, and
urine sediment were normal, and as specific antibodies to
B19 were found by enzyme linked immunosorbent assay
(ELISA) and immunoblot, B19 infection was considered and
the patient was treated with a non-steroidal anti-inflamma-
tory drug and low dose glucocorticoid. The clinical symptoms
subsided three weeks later and PR3-ANCA and MPO-ANCA
were negative after six months.
In a consecutive case series from 1998 to 2000, 1242
patients with suspected arthritis of peripheral joints attend-
ing our outpatient clinic were screened for serological
evidence of acute B19 infection. Six female patients (mean
age 34 years, range 18 to 44) presenting with mono-, oligo-,
or polyarthritis, and a macular rash in two, were diagnosed as
having acute B19 infection on the basis of specific IgM and
IgG antibodies detected by ELISA and confirmed by
immunoblot using the structural proteins VP-N, VP-C, and
VB-1S, and the non-structural protein NS-1 of the B19 virus
(Mikrogen GmbH, Munich, Germany). B19 viral nucleic acid
was found in two of four patients tested by a nested
polymerase chain reaction.7
Blood was drawn from patients with B19 infection after
informed consent had been obtained and was tested for the
presence of PR3-ANCA and MPO-ANCA by specific ELISA
(Orgentec Diagnostika, Mainz, Germany). The cut off level of
the ELISA was 5 IU/ml. Specificity of the ELISA for PR3-
ANCA and MPO-ANCA was reported to be 93–100% and the
sensitivity 68% and 55%, respectively.3After six months and
two years, all six patients were re-evaluated clinically. After
the first six months serum samples were collected and
retested for the presence of B19 IgM and IgG antibodies and
PR3-ANCA and MPO-ANCA.
To further establish the frequency of ANCA in patients
with acute viral infections, 41 366 serum samples sent to the
Institute of Hygiene between 2002 and 2003 were screened
for IgM antibodies to B19 and Epstein-Barr virus (EBV).
Forty four sera with IgM antibodies to B19, indicating
acute B19 infection (11 male, 33 female; mean (SD) age, 42
(20) years), and 51 sera with IgM antibodies to EBV,
indicating infectious mononucleosis (26 male, 25 female;
age 23 (19) years) were subsequently tested for the presence
of perinuclear (p)- or cytoplasmic (c)-ANCA by indirect
Abbreviations: ANCA, antineutrophil cytoplasmic antibodies; IIF,
indirect immunofluorescence; MPO, myeloperoxidase; PR3, proteinase 3
immunofluorescence (IIF) on ethanol, formalin, and metha-
nol fixed granulocytes (Scabos Diagnostics, Vienna, Austria)
and for PR3-ANCA and MPO-ANCA by ELISA (Orgentec
Diagnostika, Mainz, Germany).
Descriptive statistics were carried out using the software
package StatView version 5.0.
Outpatients with B19 infection and detection of ANCA
Of the six patients diagnosed with acute B19 infection, two
presented with oligoarthritis, three with polyarthritis, one
with monoarthritis, and two had a macular rash (table 1).
Five patients with B19 infection reported morning stiffness
lasting between 30 and 210 minutes. The ESR was raised in
five patients and C reactive protein levels were high in three
patients. A positive rheumatoid factor at low titre was found
in two patients. Antinuclear antibodies were negative in all
patients. One patient tested positive for PR3-ANCA and two
for PR3-ANCA and MPO-ANCA (table 2). In three patients
with B19 infection, neither PR3-ANCA nor MPO-ANCA was
detectable either at study entry or after six months of follow
up. After six months, peripheral joint disease and the
macular rash—when present—had subsided in and all
patients showed B19 seroconversion, demonstrating ade-
quate immune response to the virus (table 2). No clinical
signs or symptoms of systemic vasculitis were found in those
patients positive for PR3-ANCA or MPO-ANCA, even after a
follow up of two years’ duration. Importantly, all patients
with raised PR3-ANCA and/or MPO-ANCA at study entry
tested negative after six months of follow up. Raised ESR and
C reactive protein levels normalised after six months.
Serum samples from patients with serological
constellation of acute B19 or EBV infection
Two of 44 serum samples (5%) positive for B19 IgM
antibodies tested positive for p-ANCA in the IIF at a titre of
1:320. One of the two positive samples also tested positive for
MPO-ANCA by ELISA at a level of 49.5 U/ml. Of the 51 EBV
IgM antibody positive sera, one showed an IIF pattern of p-
ANCA at a titre of 1:320 and two a titre of 1:40, but neither of
them was positive for MPO-ANCA or PR3-ANCA by ELISA.
Conversely, no serum sample negative for ANCA by IIF tested
positive for PR3-ANCA or MPO-ANCA.
Patients presenting with arthralgia or arthritis are often a
major challenge for rheumatologists because diseases as
different as viral infection or systemic vasculitis may finally
evolve from such a monosymptomatic clinical condition.
Parvovirus infection associated with self limiting non-erosive
arthritis occurs in up to 12% of adults attending an early
arthritis clinic.8Conversely, Wegener’s granulomatosis may
also present with arthralgia, and non-erosive arthritis can be
an early symptom in more than 25% of patients.9Hence,
reliable antibody tests to distinguish Wegener’s granuloma-
tosis from viral infection are needed.
In our clinical series of patients with arthritis and acute
B19 infection, a commonly used ELISA detected PR3-ANCA
and MPO-ANCA in three of six patients in the early phases of
infection. Six months later, when clinical symptoms had
subsided and seroconversion had indicated resolution of
acute B19 infection, PR3-ANCA and MPO-ANCA were no
longer detectable. When we subsequently tested 44 sera from
individuals with acute B19 infection by IIF and ELISA we
found ANCA in two of them, which reinforces the hypothesis
that false positive ANCA may occur during acute parvovirus
infection. An analogous approach with EBV antibodies
yielded similar findings. The trivial explanation for our
observation, such as a non-specific reaction of the ELISA
test system, cannot be fully excluded as we did not have
enough sera left for IIF or immunoblotting in the clinical
series of arthritis patients. However, the occurrence of ANCA
found by IIF in 5% of individuals with acute B19 infection
suggests that there is a pathophysiological context for viral
infection and the development of ANCA. This is supported
theoretically by the following: the presence of B19 viral DNA
Characteristics of six patients with acute parvovirus B19 infection at study entry (baseline)
Pt Sex Age (y) Joints involvedRash ESR (mm/h)RF ANA
(ELISA) B19 IgM/IgG-Ab (IB)B19 PCR
ANA, antinuclear antibodies; B19 IgM/IgG-Ab, parvovirus B19 IgM and IgG antibodies; ELISA, enzyme linked immunosorbent assay; ESR, erythrocyte
sedimentation rate; F, female; IB, immunoblot; nd, not done; PCR, polymerase chain reaction; Pt, patient; RF, rheumatoid factor; y, years; +, positive; –, negative.
with arthritis and acute parvovirus B19 infection at baseline and after a follow up of six
ELISA test results for the detection of PR3-ANCA and MPO-ANCA of six patients
Baseline Six months
(IU/ml)PR3-ANCA (IU/ml)MPO-ANCA (IU/ml) B19 IgM/IgG Ab
Ab, antibodies; ANCA, antineutrophil cytoplasmic antibodies; B19 IgM/IgG-Ab, parvovirus B19 IgM- and IgG
antibodies; ELISA, enzyme linked immunosorbent assay; MPO, myeloperoxidase; PR3, proteinase 3; Pt, patient; –,
negative; 2/+, IgM antibody negative and IgG antibody positive.
642Hermann, Demel, Stu ¨nzner, et al
in tissues from patients with Wegener’s granulomatosis10; the
fact that B19 can inoculate and thereby activate endothelial
cells11; and the fact that PR3 is expressed on activated
endothelial cells and neutrophils.12In support of our findings,
a Taiwanese group also reported four cases of B19 infection
with a positive p-ANCA test, and they too detected both
MPO-ANCA and PR3-ANCA in two of them.13
ANCA were found in 5% of patients with clinically and
serologically diagnosed B19 infection and in a random
sample of sera of patients with acute B19 and EBV infection
by a routine ELISA and in two cases by IIF. ANCA were no
longer detectable after acute B19 infection had subsided,
suggesting an association with the viral infection. We
therefore propose that in clinical situations with a low
pretest probability, even those autoantibody tests known to
be highly specific for autoimmune diseases should be
repeated after a certain interval if these test results are
essential for the diagnosis.
J Hermann, U Demel, G Tilz, W Graninger, Department of Internal
Medicine, Medical University, Graz, Austria
D Stu ¨nzner, E Daghofer, Institute of Hygiene, Medical University, Graz
Correspondence to: Dr Josef Hermann, Medical University, Graz,
Department of Internal Medicine, Division of Rheumatology,
Auenbruggerplatz 15, A-8036 Graz, Austria; josef.hermann@
Accepted 22 August 2004
Published Online First 14 October 2004
1 Gregorio GV, Jones H, Choudhuri K, Vegnente A, Bortolotti F, Mieli-
Vergani G, et al. Autoantibody prevalence in chronic hepatitis B virus
infection: effect on interferon alfa. Hepatology 1996;24:520–3.
2 Varani S, Muratori L, De Ruvo N, Vivarelli M, Lazzarotto T, Gabrielli L,
et al. Autoantibody appearance in cytomegalovirus-infected liver
transplant recipients: correlation with antigenemia. J Med Virol
3 Csernok E, Ahlquist D, Ullrich S, Gross WL. A critical evaluation of
commercial immunoassays for antineutrophil cytoplasmic antibodies directed
against proteinase 3 and myeloperoxidase in Wegener’s granulomatosis and
microscopic polyangiitis. Rheumatology (Oxford) 2002;41:1313–17.
4 Ohira H, Tojo J, Shinzawa J, Suzuki T, Miyata M, Nishimaki T, et al.
Antineutrophil cytoplasmic antibody in patients with antinuclear antibody-
positive chronic hepatitis C. Fukushima J Med Sci 1998;44:83–92.
5 Von Landenberg P, Lehmann HW, Knoll A, Dorsch S, Modrow S.
Antiphospholipid antibodies in pediatric and adult patients with rheumatic
disease are associated with parvovirus B19 infection. Arthritis Rheum
6 Barnett LA, Fujinami RS. Molecular mimicry: a mechanism for autoimmune
injury. FASEB J 1992;6:840–4.
7 Gassinotti P, Weitz M, Siegl G. Human parvovirus B19 infections: routine
diagnosis by a new nested polymerase chain reaction assay. J Med Virol
8 White DG, Mortimer PP, Blake DR, Woolf AD, Cohen BJ, Bacon PA. Human
parvovirus arthropathy. Lancet, 1985;i, 419–21.
9 Noritake DT, Weiner SR, Bassett LW, Paulus HE, Weisbart R. Rheumatic
manifestations of Wegener’s granulomatosis. J Rheumatol 1987;14:949–51.
10 Finkel TH, Torok TJ, Ferguson PJ, Durigon EL, Zaki SR, Leung DY, et al.
Chronic parvovirus B19 infection and systemic necrotising vasculitis:
opportunistic infection or aetiological agent? [see comments]. Lancet
11 Magro CM, Nuovo G, Ferri C, Crowson AN, Giuggioli D, Sebastiani M.
Parvoviral infection of endothelial cells and stromal fibroblasts: a possible
pathogenetic role in scleroderma. J Cutan Pathol 2004;31:43–50.
12 Mayet WJ, Schwarting A, Meyer zum Buschenfelde KH. Cytotoxic effects of
antibodies to proteinase 3 (c-ANCA) on human endothelial cells. Clin Exp
13 Chou TN, Hsu TC, Chen RM, Lin LI, Tsay GJ. Parvovirus B19 infection
associated with the production of anti-neutrophil cytoplasmic antibody
(ANCA) and anticardiolipin antibody (aCL). Lupus 2000;9:551–4.
Transiently positive ANCA during recent viral infection643