Human polyomavirus (BK) infection and ureteric stenosis in renal allograft recipients.

Human polyomavirus (BK) was detected in two renal allograft recipients as a result of routine examination of Papanicolaou-stained smears of urinary sediment in the light microscope. Infection with this recently identified virus was confirmed by virus isolation and electron microscopy. The cytological, histological, and ultrastructural changes due to the virus are described, and virus excretion is correlated with the clinical progress of the patients and the pathological findings. The transplant ureters in both patients were found to be ulcerated and stenosed, and virus-infected cells were observed in the ureteric epithelium. We suggest that the administration of high-dose steroids in transplantation may permit active infection with human polyomavirus to occur in ureteric epithelium which has been damaged by ischaemia or inflammation.     

Activation of human polyomavirus infection-detection by cytologic technics

Human polyomaviruses produce characteristic large basophilic nuclear inclusions in urothelial cells. Cytologic screening of urinary sediment for inclusion-bearing cells permits the identification of persons who are actively excreting these viruses. The human polyomavirus-induced changes may be differentiated from changes induced by other viruses and from cancer by transmission electron microscopy of infected cells. Cytologic evidence of virus infection was detected in five of 37 cancer patients receiving various treatments, two of whom were also diabetic, and in two of 84 adult patients who had diabetes mellitus. The monitoring of 3,648 urine samples sent for routine cytologic examination revealed 12 additional patients to have cytologic changes of human polyomavirus infection. Electron microscopy confirmed the presence of polyomavirus particles in seven of the 19 cytologically positive specimens. Previous reports of human polyomavirus excretion have been confined to describing patients whose immunity may have been impaired by drug therapy, congenital disease, or pregnancy. This study indicates that cytologic evidence of active human polyomavirus infection may be found among patients receiving various treatments for a number of medical disorders not usually associated with immunologic defects. Further studies to identify factors concerned in the reactivation of human polyomavirus are indicated.     

Early events in polyomavirus infection: fusion of monopinocytotic vesicles containing virions with mouse kidney cell nuclei

The entry of polyomavirus enclosed in monopinocytotic vesicles into mouse kidney cell nuclei was studied and evidence for a fusion mechanism was obtained. In vivo studies using the fluorescent lipophilic dye diI-C16(3) as a plasma membrane label showed that polyomavirus-infected nuclei accumulate plasma membrane, while uninfected or polyoma capsid-infected nuclei do not. Further evidence for fusion was obtained with electron microscopy of thin sections of infected mouse kidney cells. These specimens showed accumulation of plasma membrane in the outer nuclear membrane as well as evidence of recent fusion events. The polyoma virions (capsid proteins) were seen to accumulate on the inner nuclear membrane and in the nucleus and were identified by immunogold staining of the thin sections. The combined results of the in vivo dye studies and thin section immunoelectron microscopy studies provide evidence for a fusion mechanism for polyomavirus entry into mouse kidney cell nuclei.     

Characterization of a new polyomavirus (Polyomavirus papionis-2) isolated from baboon kidney cell cultures

Summary Viruses with papovavirus morphology were seen in fluids from baboon kidney cell cultures on three separate occasions (isolates A, B, and C). The size of the virions, 47.9 nm, placed the virus in the polyomavirus genus. It grew well in baboon kidney and Vero cells and less well in human embryo lung (HEL) fibroblasts.The virus could not be identified as the previously described baboon polyomavirus, SA 12, or as any of the other known primate polyomaviruses BK, JC or SV 40, the non-primate viruses mouse polyoma, K, rabbit kidney vacuolating virus (RKV) or bovine polyomavirus (FRKV) by immunofluorescence, immune electron microscopy or hemagglutination inhibition (HI) tests. A rabbit antiserum to the new virus (isolate A) reacted only with the three isolates and not with the other primate polyomaviruses studied.Thirteen percent of 118 wild-caught baboons (Papio anubis) had HI antibody to the new polyomavirus and 21 percent were seropositive for SA 12; only two baboons had antibody to both viruses. These results suggest that in baboons there are two antigenically distinct polyomaviruses which circulate independently. The two viruses may also be distinguished by their hemagglutinating properties: SA 12 agglutinated erythrocytes from a wider range of species but only the newly recognized polyomavirus agglutinated baboon erythrocytes.We propose that the two baboon viruses, SA 12 and the new virus, should be namedPolyomavirus papionis-1 andPolyomavirus papionis-2 respectively.     

BK Polyomavirus in Renal Transplants: Role of Electron Microscopy and Immunostaining in Detecting Early Infection

Reactivation of BK polyomavirus (BKV) is increasingly recognized as a cause of failure of renal allografts. Since no specific treatment is available for this infection, early diagnosis is important, as it allows for early intervention and possible recovery of renal function. Forty-four consecutive renal transplant biopsies performed over a 2-year period were included in the study. In addition to evaluation of renal biopsy tissue sections using routine histochemical stains, CD3, CD20, BK virus immunostains using the specific BK virus and the SV40 antibodies and electron microscopy studies were performed. None of the transplant cases but one exhibited classical histologic viral changes. Viral particles were seen by EM in 19%, and BK-virus positivity was identified in only 43% of these cases. CD20-rich inflammatory infiltrates predominated in cases in which either positive BK stain and/or viral particles were identified ultrastructurally. A combined approach using electron microscopic and immunohistochemical evaluation can be utilized effectively to identify BK virus-associated nephropathy at an early phase facilitating early clinical intervention.     

BK polyomavirus in renal transplants: role of electron microscopy and immunostaining in detecting early infection

Reactivation of BK polyomavirus (BKV) is increasingly recognized as a cause of failure of renal allografts. Since no specific treatment is available for this infection, early diagnosis is important, as it allows for early intervention and possible recovery of renal function. Forty-four consecutive renal transplant biopsies performed over a 2-year period were included in the study. In addition to evaluation of renal biopsy tissue sections using routine histochemical stains, CD3, CD20, BK virus immunostains using the specific BK virus and the SV40 antibodies and electron microscopy studies were performed. None of the transplant cases but one exhibited classical histologic viral changes. Viral particles were seen by EM in 19%, and BK-virus positivity was identified in only 43% of these cases. CD20-rich inflammatory infiltrates predominated in cases in which either positive BK stain and/or viral particles were identified ultrastructurally. A combined approach using electron microscopic and immunohistochemical evaluation can be utilized effectively to identify BK virus-associated nephropathy at an early phase facilitating early clinical intervention.     

Contribution of transmission electron microscopy to fine-needle aspiration biopsy diagnosis: Comparison of cytology and combined cytology and transmission electron microscopy with final histological diagnosis

This report evaluates 74 fine-needle aspiration biopsies processed for transmission electron microscopy with subsequent surgical procedure. The specificity of diagnosis obtained by cytology alone was compared to that obtained by cytology and electron microscopy, using histologic diagnosis as the gold standard. When cytology gave a diagnosis of malignancy but could not give tumor category or type, electron microscopy could correctly give both. When cytology could give tumor category but not type, electron microscopy correctly identified type in the majority of cases. When cytology gave tumor category and type, electron microscopy confirmed the diagnosis. Transmission electron microscopy is very helpful when the cytopathologist can diagnose malignancy but cannot give tumor category and/or type. When the cytopathologist is specific in his/her diagnosis, TEM is not as helpful. Diagn Cytopathol 1996;15: 282–287. © 1996 Wiley-Liss, Inc.     

Identification of mineral particles in pneumoconiotic lungs

An extraction replication technique has been used for the study, by electron microscopy, of lung tissue from a number of different cases of pneumoconiosis.The technique provides a relatively simple means of studying the surface area of replicated tissues, and any foreign particles present which are either replicated or extracted from the tissue can be identified. A set of standard micrographs of different types of mineral particles likely to be encountered should be kept for reference. Most of the particles identified by this technique were beyond the limits of resolution of optical microscopy. Furthermore, the procedure involves the minimum of chemical and physical treatment.The histochemical diagnosis and industrial history was confirmed in the first two cases described, by the electron microscopy investigations. The third case presented an uncertain histological and industrial history, but an electron microscopy study confirmed the presence of the mineral particle considered responsible for the clinical condition. The fourth and fifth cases described illustrate the use of this technique in identifying a particular type of pleomorphic mineral, and illustrating its position in situ in relation to surrounding tissue.     

Uncombable hair syndrome: a clinical report

Uncombable hair syndrome, also named "pili trianguli et canaliculi" or "cheveux incoiffables", is a rare structural anomaly of the hair shaft first reported in 1973. Both inherited and sporadic forms have been described, characterized by dry and frizzy scalp hair that is impossible to comb. Diagnosis is suspected clinically and confirmed by scanning electron microscopy. The condition is usually isolated, however, several physical abnormalities can be associated. We report the case of a 2(1/2) year old-girl presenting isolated uncombable hair syndrome suspected clinically and confirmed by scanning electron microscopy.     

Identification of acrylic cement particles in tissues

Investigation of the tissue response to acrylic cement particles is hampered by the fact that they are affected by tissue processing for transmission electron microscopy and for routine light microscopy. However, in the case of transmission electron microscopy, the present study shows that intracellular and extracellular cement sites can, in favourable circumstances, be identified by their heterogeneous appearance in the electron beam. This appearance results from the biphasic system which can form from the acrylic and the epoxy embedding medium during tissue processing. The validity of this means of identification has been confirmed (a) by observing the surface morphology of the particles preparedin vitro for the present study; (b) by use of barium sulphate as a marker in the cement; (c) by examination of control tissue samples. In the case of light microscopy, acrylic cement particles can be stained with Sudan dyes in frozen sections, as previously described by Willert and Semlitsch, or identified in paraffin sections by the presence of barium sulphate marker.     

Polyomavirus inclusion bodies in chicken caecal epithelium

In this case report we describe the light microscopic and transmission electron microscopic appearance of polyomavirus inclusions in caecal epithelial cells from a chicken with diarrhoea and intestinal parasites. The DNA in situ hybridization technique demonstrated that the polyomavirus reported here is different from polyomaviruses observed in psittaciformes. The chicken polyomavirus described may share some homologous base sequences with psittaciforme polyomavirus; however, the viruses are different. Clinicians and pathologists should be aware that polyomavirus can infect chickens and that polyomavirus inclusion bodies can be found in chicken caecal epithelial cells.     

Murine polyomavirus infection of 3T6 mouse cells shows evidence of predominant necrosis as well as limited apoptosis

The current study was developed to determine if polyomavirus infected 3T6 mouse cells evoked an apoptotic or a necrotic mechanism during infection. Infected cells were analyzed by flow cytometry, transmission electron microscopy (TEM), DNA electrophoresis and by measuring caspase-3 enzymatic activity. Infected cells that were analyzed at 72 h post-infection showed the following: flow cytometry analysis revealed a 5% increase in apoptotic cells and a 46% increase in necrotic cells when compared to uninfected cells; electron microscopy showed 10% cells with characteristic apoptotic morphology and 40% with necrotic appearance; caspase-3 activity was found to increase two fold when compared to uninfected cells and DNA fragmentation (laddering) was clearly evident late in infection. It was concluded that infected cells predominantly showed necrosis, although some cells showed apoptosis in late infection. Recombinant capsid-like particles composed of the polyomavirus structural proteins were not able to induce cell death.     

WU polyomavirus in children with acute lower respiratory tract infections, China.

BACKGROUND: WU polyomavirus (WUPyV), a new member of the genus of Polyomavirus in the family Polyomaviridae, has been found and associated with respiratory tract infections recently. However, its clinical role and pathogenicity has not been known. OBJECTIVES: To confirm that WU polyomavirus has been found in Chinese children. STUDY DESIGN: WU polyomavirus was detected and identified using PCR methods. A total of 278 specimens of nasopharyngeal aspirate were collected, and then PCR products were sequenced directly. RESULTS: One of 278 nasopharyngeal aspirates was positive for WUPyV in one child, and the positive rate was 0.4%. The results showed that the sequences of genome, LTAg and VP2 gene was identical to the reference sequences of WU polyomavirus prototype strains. CONCLUSIONS: We confirmed that WU polyomavirus had been found and identified in the respiratory secretions in China.     

Pulmonary carcinomas with a sarcomatoid element: an immunocytochemical and ultrastructural analysis

Eight primary carcinomas of the lung with a prominent spindle-cell sarcomatoid component were studied by immunocytochemical staining and electron microscopy. The eight tumors were indistinguishable by conventional light microscopy, with the exception of one unusual neoplasm that followed multiple pathways of differentiation with elements of squamous cell carcinoma, rhabdomyosarcoma, chondrosarcoma, and an undifferentiated spindle-cell population. Reticulin fiber production by individual spindle cells and a sharp demarcation of the carcinomatous and sarcomatoid domains by light microscopy were not useful differentiating features. Three of the eight tumors exhibited keratin expression in both the carcinomatous and spindle-cell components. Both immunocytochemical and electron microscopic analyses were required to detect epithelial differentiation, as in one case keratin was identified only by immunocytochemical staining and in another only by ultrastructural examination. Epithelial differentiation was undetectable in the sarcomatoid component of five tumors, and in one case immunoreactive myoglobin was identified in spindle cells; skeletal muscle differentiation was confirmed ultrastructurally. We propose that pulmonary carcinomas exhibiting evidence of epithelial differentiation in a sarcomatoid component be termed spindle-cell carcinomas and that those biphasic tumors exhibiting mesenchymal differentiation into specific tissues, such as neoplastic bone, cartilage, or striated muscle, or lacking epithelial differentiation by light microscopy, immunocytochemistry, and electron microscopy be classified as carcinosarcomas. This distinction may ultimately be unnecessary, because these two tumors may represent different points along a morphologic and biologic continuum.     

Analysis of hamster lymphomas for the presence of hamster papovavirus DNA

The hamster papovavirus (HaPV) is a polyomavirus isolated from skin epitheliomas arising spontaneously in young Syrian hamsters. It can induce lymphomas and leukaemias in newborn hamsters. Although no virus particles are detectable by electron microscopy, high amounts of monomeric and oligomeric forms of extrachromosomal HaPV DNA molecules are found in the lymphoma cells. These molecules display deletions of about 300 nucleotides in length. Their role in the lymphoma induction is discussed.     

The Quarterly Case Retroperitoneal Tumor with Liver Metastases in a 38-Year-Old Female

A correlative microscopy method for the ultrastructural analysis of focal viral tissue infections is presented. Using a confocal scanning laser microscope, foci of infection are identified in tissue sections prior to embedment; a variety of techniques can be employed for viral detection, including staining with standard histochemical reagents and fluorescently labeled antibodies. Areas of infection identified using confocal microscopy are excised from the tissue sections, embedded, and examined by transmission electron microscopy. Applications of this technique in both diagnostic and basic research settings are described.     

Generation of virus-like particles consisting of the major capsid protein VP1 of goose hemorrhagic polyomavirus and their application in serological tests

Goose hemorrhagic polyomavirus (GHPV) is the causative agent of hemorrhagic nephritis and enteritis of geese (HNEG), a fatal disease of young geese with high mortality rates. GHPV cannot be efficiently propagated in tissue culture. To provide antigens for diagnostic tests and vaccines, its major structural protein VP1 was recombinantly expressed in Sf9 insect cells and in the yeast Saccharomyces cerevisiae. As demonstrated by density gradient centrifugation and electron microscopy, GHPV-VP1 expressed in insect cells formed virus-like particles (VLPs) with a diameter of 45 nm indistinguishable from infectious polyomavirus particles. However, efficiency of VLP formation was low as compared to the monkey polyomavirus SV-40-VP1. In yeast cells, GHPV-VP1 alone formed smaller VLPs, 20 nm in diameter. Remarkably, co-expression of GHPV-VP2 resulted in VLPs with a diameter of 45 nm. All three types of GHPV-VLPs were shown to hemagglutinate chicken erythrocytes. ELISA and hemagglutination inhibition tests using the VLPs as antigen detected GHPV-specific antibodies in up to 85.7% of sera derived from flocks with HNEG but in none of the sera of a clinically healthy flock. However, GHPV-specific antibodies were also detected in sera from two other flocks without HNEG indicating a broad distribution of GHPV due to subclinical or unrecognised infections.     

The structure of avian polyomavirus reveals variably sized capsids, non-conserved inter-capsomere interactions, and a possible location of the minor capsid protein VP4

Avian polyomavirus (APV) causes a fatal, multi-organ disease among several bird species. Using cryogenic electron microscopy and other biochemical techniques, we investigated the structure of APV and compared it to that of mammalian polyomaviruses, particularly JC polyomavirus and simian virus 40. The structure of the pentameric major capsid protein (VP1) is mostly conserved; however, APV VP1 has a unique, truncated C-terminus that eliminates an intercapsomere-connecting β-hairpin observed in other polyomaviruses. We postulate that the terminal β-hairpin locks other polyomavirus capsids in a stable conformation and that absence of the hairpin leads to the observed capsid size variation in APV. Plug-like density features were observed at the base of the VP1 pentamers, consistent with the known location of minor capsid proteins VP2 and VP3. However, the plug density is more prominent in APV and may include VP4, a minor capsid protein unique to bird polyomaviruses.     

Electron microscopy in the diagnosis of BK-polyoma virus infection in the transplanted kidney

BK polyomavirus has become an important etiologic agent responsible for significant morbidity in renal transplant recipients. This virus can be detected in transitional cells in the urine (decoy cells) using cytology, but correlation with allograft function status and histologic evidence of renal involvement is poor. Accurate diagnosis of BK polyomavirus infection requires a high index of suspicion and utilization of ancillary diagnostic techniques in many cases. Electron microscopy is very sensitive in depicting the presence of BK virions, but the finding of viral particles is not by itself diagnostic of BK interstitial nephritis. Management of patients with polyoma virus nephropathy is difficult since there is no specific antiviral therapy available at this time.     

BK Polyomavirus Interstitial Nephritis in a Renal Allograft Recipient

Human polyomavirus (PV) interstitial nephritis has recently been recognized as a cause of severe renal allograft dysfunction. It occurs in immunosuppressed patients after reactivation of the latent virus PV type BK (BK virus) in the renal epithelium. BK disease is defined as a morphologically manifest renal infection with cytopathic signs accompanied by varying degrees of interstitial inflammatory cell infiltrates and functional impairment. It is also identified by the presence of cells containing viral inclusion bodies (decoy cells) in the urine. The authors report a case of BK PV interstitial nephritis in a 36-year-old renal allograft recipient. Under light microscopy the chief diagnostic indicator was detection of intranuclear viral inclusions, which were found exclusively in tubular epithelial cells. Cells with viral changes were often enlarged with nuclear atypia and chromatin basophilia. Widespread interstitial plasma cell infiltrates associated with tubulitis were present. Intranuclear paracrystalline arrays of virus particles 35-38 nm in diameter were present as characteristic ultrastructural indicators. Urine samples revealed decoy cells with ground-glass-type intranuclear inclusions positive for BK virus by electron microcopy.