BK virus encephalitis in a patient with AIDS and lymphoma

BK virus (BKV) is excreted in the urine of 8–37% of the patients with HIV infection, and BK viruria is increased with immunodeficiency [1,2]. Viral DNA in the peripheral blood mononuclear cells may also be detected in up to 50% of these patients [3]. However, clinical manifestations such as haemorrhagic cystitis or tubulointerstitial nephritis are rare [2,4]. We describe a patient with AIDS and non Hodgkin's lymphoma (NHL) suffering from a severe BKV-related encephalitis. A 44-year-old man was admitted in September 1999 for cachexia, ascites, and lymphadenopathy. Neurological examination was normal. A large cell NHL was diagnosed on the basis of a cytological examination of the bone marrow aspirate and ascites. The lumbar puncture and the brain computed tomography scan revealed no abnormalities. HIV-1 infection was diagnosed and the CD4 lymphocyte count was 65 × 106/l. After four cycles of chemotherapy a complete remission of the NHL was obtained. In November 1999, the patient developed haemorragic cystitis and progressive paraplegia. Laboratory examinations revealed an increase in the serum creatinine level to 230 μmol/l and severe pancytopenia. The bone marrow aspirate showed a reactive haemophagocytic syndrome, but no relapse of the NHL could be detected. The cerebrospinal fluid (CSF) was acellular. Magnetic resonance tomography of the brain revealed diffuse areas of increased signal intensity of the periventricular white matter. At that time, BKV DNA was detected by polymerase chain reaction (PCR) in the urine, blood, bone marrow, and CSF. The paraplegia worsened and the patient died in January 2000. A postmortem examination was performed. Tissue samples were embedded in paraffin or immediately frozen and stored at −80°C until PCR testing. Of the brain samples, only outer layers of the cerebral cortex were available for PCR. BKV DNA was sought by PCR followed by hybridization onto a specific oligonucleotide probe, using primers, probe sequences and PCR conditions previously described [5]. Sections were stained with haematoxylin and eosin, Bodian silver impregnation or Cresyl violet combined with Luxol-fast-blue. Immunocytochemistry was performed using a polyclonal anti-SV40 antibody that cross-react with most polyomaviruses, polyclonal antibodies raised against glial fibrillary acidic protein and monoclonal antibodies raised against MHC class II antigens, herpes virus antigens, the HIV protein P24, and B (CD138) and T (CD3) cells (Biosoft, Saint Quentin en Yvelines, France). In-situ hybridization was performed using biotinylated JCV, SV40 and BKV bioprobes (Enzo Diagnostics, New York, USA) and an amplification kit for BKV (GenPoint K620, Dako). Fluoresceinated probes were used to demonstrate Epstein–Barr virus EBER genome (Dako S.A., Trappes, France). Neuropathological examination showed that occasionnal CD 138-positive lymphomatous cells were present in the leptomeninges at the base of the brain. These cells did not express Epstein–Barr virus proteins or genome. Diffuse encephalomyelitis with proliferation of reactive astrocytes (Fig. 1a), and microglial activation with proliferation of rod-shaped microglia and nodules (Fig. 1b) involved the grey and white matter. It was severe in the subependymal regions of the hippocampus, thalamus and brainstem. Immunostaining for HIV and herpesviruses was negative. In contrast, immunostaining using an anti-SV40 antibody, which reacts with most polyomavirus positively stained cells in the ependymal lining, leptomeninges, subpial brain parenchyma, and choroid plexus. In-situ hybridization for BKV positively stained the nuclei of ependymal and glial cells in the subpial areas (Fig. 1c), astrocytes and microglial cells within foci of encephalitis. Staining for JCV and SV40 was negative. BKV DNA was not found by PCR in the outer layers of the cerebral cortex. Microscopic examination confirmed NHL infiltration of the kidneys, bladder, mesenteric nodes, and stomach. There was also necrosis of the tubular epithelium of the kidneys and intranuclear inclusion bodies. In-situ hybridization for BKV positively stained the nuclei of the tubular cells and of rare lymphoma cells (Fig. 1d). Postmortem PCR was positive in the kidney, bladder, mesenteric node, and stomach tissue samples.Fig. 1.: Pathological analysis of the brain and urinary tract. (a) Immunostaining of the glial fibrillary acidic protein revealed by an indirect immunoalkaline phosphatase method (APAAP) shows marked reactive astrocytosis (in red) in the hippocampus (× 250). (b) Immunostaining for MHC class II antigens (HLA-DR) revealing activated microglial cells shows marked microglial activation with a microglial nodule in the thalamus (APAAP, × 100). (c) In-situ hybridization for BK virus positively stains the nucleus of two ependymal cells and a glial cell in the underlying parenchyma (× 1000). (d) In-situ hybridization for BK virus positively stains the nucleus of a lymphoma cell within a deposit surrounding the bladder (× 1000).Our patient had a severe immunodeficiency as a result of advanced HIV disease and cytotoxic chemotherapy. BKV was initially sought in the case because of the appearance of haemorrhagic cystitis, a well-known complication of BKV infection. A clinically significant encephalitis soon developed, and disseminated BKV infection was suspected on the basis of the detection of viral DNA in CSF and bone marrow cells. The pathogenic role of BKV in severe central nervous system (CNS) disease was further supported by the postmortem findings. In situ hybridization study showed the presence of BKV genome mostly in the ventricular and meningeal brain surfaces, as previously shown [6,7]. Moreover, no other pathogen was detected in the brain. Two cases of BKV encephalitis have been reported in patients with AIDS, encephalitis and renal disease. BKV was first isolated from CSF and brain autopsy material [6]. Evidence of BKV infection in the CNS was later supported by the demonstration of viral genome and proteins in the brain and CSF [7]. Two histopathological studies have shown that BKV DNA may be detected by PCR in the brain tissues of 3–6% of HIV-infected patients with or without neurological symptoms [8,9]. Therefore, in those studies BKV was detected by PCR in the brain, but no direct evidence of BKV replication was demonstrated. In contrast, the detection of BKV genome in the CSF is rare and seems to be of particular value. Negative results were observed in 400 CSF samples from immunocompromised patients with neurological symptoms [7]. The presence of BKV genome in malignant B cells raises the question of the role of the virus in B cell transformation. In our patient, the spread of lymphoma cells in the urinary tract and leptomeninges was concordant with the presence of BKV genome within the tubular cells of the kidneys and glial cells. The demonstration of BKV-infected lymphoma cells may support the view that BKV, as is the case with the other polyomaviruses, SV40 or JCV, might be oncogenic [10,11]. In conclusion, our report adds evidence that BKV must be considered in the differential diagnosis of CNS disorders in HIV patients. The diagnosis may be suggested by PCR-detectable BKV DNA in the CSF. We suggest that additional immunosuppression such as chemotherapy may increase the risk of BKV replication and the development of encephalitis. Acknowledgements The authors would like to thank Prs J. Mikol, A. Probst and P. Gaulard for their help in the histopathological analysis. Philippe Lesprita Dominique Chaline-Lehmannb François-Jérôme Authierc Tibor Ponnellec Françoise Grayd Yves Levya