KI Polyomaviruses Research Articles

Quantitative detection of the new polyomaviruses KI, WU and Merkel cell virus in transbronchial biopsies from lung transplant recipients

BackgroundRecently, three new polyomaviruses—KI, WU and Merkel cell (MCV)—have been discovered and their detection has been reported in different types of specimens, including respiratory samples, suggesting their shedding in the...

KI and WU Polyomavirus detection in tonsils of immunocompetent patients

The Polyomaviruses KI and WU have been identified in 2007 in nasopharyngeal aspirates of patients with acute respiratory tract infections.They have been further isolated in faecal samples, lymphoid tissue, bronchoalveolar lavages and blood, although their pathological associations, molecular epidemiology and tissue tropism are still not widely known.The aim of this study was to determine the presence of KIV and WUV in tonsillar tissue of immunocompetent patients to assess whether the tonsils may represent a site of infection and/or persistence.The presence of Polyomavirus BKV, JCV and SV40 was also evaluated. The presence of Polyomavirus KI,WU, BK, JC and SV40-DNA was evaluated in a prospective study on tonsillar samples obtained from 29 immunocompetent patients (adults and children undergoing tonsillectomy) by a commercially available Taqman Real-time quantitative PCR (BKV Q-PCR Alert Kit, Nanogen) and a home-made system (for KIV,WUV, SV40, JCV).The amplification conditions were optimized and standardized. KIV-DNA was detected in 2/29 (6.9%) tonsil tissue (obtained from an adult and a child), while WU, BK, JC and SV40 were negative in all cases. Our prevalence results are consistent with those reported in the literature on samples from the respiratory tract and lymphoid tissue such as tonsils. Further studies are needed to understand the significance of the presence of KIV in tonsil tissue and assess the possibility that the virus can establish latent and/or persistent infection at this level.

Newly described human polyomaviruses Merkel Cell, KI and WU are present in urban sewage and may represent potential environmental contaminants

Recently, three new polyomaviruses (KI, WU and Merkel cell polyomavirus) have been reported to infect humans. It has also been suggested that lymphotropic polyomavirus, a virus of simian origin, infects humans. KI and WU polyomaviruses have been detected mainly in specimens from the respiratory tract while Merkel cell polyomavirus has been described in a very high percentage of Merkel cell carcinomas. The distribution, excretion level and transmission routes of these viruses remain unknown.Here we analyzed the presence and characteristics of newly described human polyomaviruses in urban sewage and river water in order to assess the excretion level and the potential role of water as a route of transmission of these viruses. Nested-PCR assays were designed for the sensitive detection of the viruses studied and the amplicons obtained were confirmed by sequencing analysis. The viruses were concentrated following a methodology previously developed for the detection of JC and BK human polyomaviruses in environmental samples. JC polyomavirus and human adenoviruses were used as markers of human contamination in the samples. Merkel cell polyomavirus was detected in 7/8 urban sewage samples collected and in 2/7 river water samples. Also one urine sample from a pregnant woman, out of 4 samples analyzed, was positive for this virus. KI and WU polyomaviruses were identified in 1/8 and 2/8 sewage samples respectively. The viral strains detected were highly homologous with other strains reported from several other geographical areas. Lymphotropic polyomavirus was not detected in any of the 13 sewage neither in 9 biosolid/sludge samples analyzed.This is the first description of a virus isolated from sewage and river water with a strong association with cancer. Our data indicate that the Merkel cell polyomavirus is prevalent in the population and that it may be disseminated through the fecal/urine contamination of water. The procedure developed may constitute a useful tool for studying the excreted strains, prevalence and transmission of these recently described polyomaviruses.

Genetic variability of the small t antigen of the novel KI, WU and MC polyomaviruses

Recently, three novel human polyomaviruses were discovered: KIPyV, WUPyV and MCPyV. To study the genetic variability of these viruses, an evolutionary analysis of the large T antigen, small t antigen, VP1, VP2 and VP3 genes was carried out. Of the genes analyzed, only the small t antigen of KIPyV and MCPyV was found to be under statistically supported selective pressure. In KIPyV, 31 selected sites were found under positive selection (ω(2)=1.54), with posterior probabilities above 0.75. Of these sites, 17 are located in the J domain and two of these are located in the HPDGG box. Thirteen of these sites were identified as being under positive selection at the 99% confidence level. Moreover, seven codons were under significant purifying selection, particularly in the J domain. In MCPyV, we confirmed the five sites already found to be under positive selection (posterior probabilities 0.99). Selective pressure analysis may provide useful information on viral evolution.

Prevalence and Clinical Impact of Polyomaviruses KI and WU in Lung Transplant Recipients

The newly discovered polyomaviruses KI and WU (KIV and WUV) were isolated from secretions of patients with respiratory symptoms as well as in blood, spleen, lymphoid tissues, and stools, especially in immunocompromised conditions. The aim of this work was to evaluate the prevalence of KIV and WUV in bronchoalveolar lavage (BAL) from lung transplant recipients. We also examined potential correlations between these viruses and occurrences of pneumonia, acute respiratory insufficiency, or other acute respiratory conditions and acute rejection episodes. Discharge diagnosis was based on the International Classification of Diseases–Italian version 2002, based on the 9th-revision clinical modification. A rejection episode was diagnosed by transbronchial lung biopsy in accordance with the 2007 International Society for Heart and Lung Transplantation Working Formulation. Overall, we analyzed 53 BALs obtained from 24 transplant recipients. Positive polymerase chain reaction results were observed in 6 samples (11.3%) from 6 patients (25%), versus 7 samples (13.2%) from 7 patients (29.2%) for KIV and WUV, respectively. Regarding the diagnosis of pneumonia, the prevalence was 22.2% and 33.3% for KIV and WUV, respectively. In cases of acute respiratory insufficiency or other acute respiratory conditions, 2 out of 9 samples were positive for KIV (22.2%) and 4 out of 9 for WUV (44.4%). An Acute rejection episode (ARE) was diagnosed in 7 instances among 6 lung transplant patients: The corresponding BAL specimens showed positive results for KIV in 3 out of 7 (42.8%) cases with ARE vs 3 out of 46 (6.5%) without an ARE ( P < .05), and for WUV in 3 out of 7 (42.8%) vs 4 out of 46 (8.7%) ( P < .05), respectively. Although the small number of specimens limits the statistical analysis, our results showed a higher prevalence of WUV compared with KIV. The compromised pulmonary environment in the lung allograft may cause reactivation of these viruses. Their roles in this context need to be further evaluated.

WU and KI Polyomaviruses Remain Orphans in Adults

To the Editor—We read with interest the brief report by Barzon et al [1] showing that WU and KI polyomaviruses (WUPyV and KIPyV) reactivate and are frequently detectable in the brains of human immunodeficiency virus (HIV)–positive patients without being associated with progressive multifocal leukoencephalopathy. We also recently disproved such an association between novel polyomaviruses (including lymphotropic polyomavirus [LPyV] and Merkel cell carcinoma polyomavirus [MCPyV]) and progressive multifocal leukoencephalopathy in HIV-negative patients [2]. However, because the prevalence of novel polyomaviruses remains high and because they have been shown to reactivate under conditions of immune deficiency [3], they could still prove to be useful as surrogate markers of functional immunodeficiency, as was recently proposed for another orphan virus, torquetenovirus (TTV) [4]. Therefore, we analyzed the occurrence of WUPyV, KIPyV, LPyV, and MCPyV viremia (by the same methods used previously [3]) at the time of peak TTV viremia after high-dose chemotherapy supported by autologous stem cell transplant in 17 patients with multiple myeloma, 1 with non-Hodgkin lymphoma, and 1 with systemic sclerosis. All samples tested negative. It remains difficult to identify the body site with the highest prevalence and, hence, the greatest sensitivity for these viruses; it could be feces [5], in contrast to urine for JC and BK polyomaviruses [6]. Given that many polyomaviruses are lymphotropic and retain oncogenic potential, we investigated their occurrence in lymph node biopsy samples from patients with lymphoma or leukemia. We could not find any occurrence of WUPyV, KIPyV, LPyV, and MCPyV in any of 49 lymph node samples tested (from, namely, 11 patients with follicular lymphoma, 14 with diffuse large B cell lymphoma, 10 with B cell chronic lymphocytic leukemia, 7 with Hodgkin lymphoma, 2 with hairy cell leukemia, 2 with mantle cell lymphoma, 1 with Burkitt lymphoma, 1 with splenic marginal zone lymphoma, and 1 with B cell acute lymphoblastic leukemia).

Survey for the presence of BK, JC, KI, WU and Merkel cell polyomaviruses in human brain tissues

Background Recently three previously unknown polyomaviruses (KI, WU and Merkel cell polyomaviruses) have been identified from human specimens. The spectrum of clinical manifestations and their tissue tropism are currently unknown. Since a member of this virus family, JC virus, is well-known for its capacity to establish latency in human brain tissue where reactivation in immunocompromised individuals can result in fatal progressive multifocal leukoencephalopathy, we sought to examine for the presence of all the five known human polyomaviruses in a series of human brain tissues. Objectives To investigate the possibility of neuropersistence of the newly identified human polyomaviruses. Study design Autopsy brain tissues were collected from 10 different brain regions of 30 individuals who died from diseases unrelated to viral infections. Nested PCR was used to assess the presence or absence of viral DNA. Results Ten samples collected from five individuals were found to harbour JCV DNA. In contrast, none of the 300 brain tissues examined showed positive results for BK, KI, WU or Merkel cell polyomavirus. Conclusion The newly identified KI, WU and Merkel cell polyomaviruses either do not, or have a much lower neuropersistent potential compared to JCV.

DNA from KI, WU and Merkel cell polyomaviruses is not detected in childhood central nervous system tumours or neuroblastomas.

BackgroundBK and JC polyomaviruses (BKV and JCV) are potentially oncogenic and have in the past inconclusively been associated with tumours of the central nervous system (CNS), while BKV has been hinted, but not confirmed to be associated with neuroblastomas. Recently three new polyomaviruses (KIPyV, WUPyV and MCPyV) were identified in humans. So far KIPyV and WUPyV have not been associated to human diseases, while MCPyV was discovered in Merkel Cell carcinomas and may have neuroepithelial cell tropism. However, all three viruses can be potentially oncogenic and this compelled us to investigate for their presence in childhood CNS and neuroblastomas.MethodologyThe presence of KI, WU and MCPyV DNA was analysed, by a joint WU and KI specific PCR (covering part of VP1) and by a MCPyV specific regular and real time quantitative PCR (covering part of Large T) in 25 CNS tumour biopsies and 31 neuroblastoma biopsies from the Karolinska University Hospital, Sweden. None of the three new human polyomaviruses were found to be associated with any of the tumours, despite the presence of PCR amplifiable DNA assayed by a S14 housekeeping gene PCR.ConclusionIn this pilot study, the presence of MCPyV, KI and WU was not observed in childhood CNS tumours and neuroblastomas. Nonetheless, we suggest that additional data are warranted in tumours of the central and peripheral nervous systems and we do not exclude that other still not yet detected polyomaviruses could be present in these tumours.

Merkel Cell Polyomavirus

The recently described Merkel cell polyomavirus (MCPyV) is reportedly present in 50% to 80% of Merkel cell carcinomas (MCC). Although the virus has been shown to be absent from other cutaneous neoplasms, its association with malignancies that are histologically similar to MCC, specifically small cell carcinoma of the lung and other high-grade neuroendocrine tumors, has yet to be thoroughly investigated. To address this issue, we identified a set of 74 cases of visceral high-grade neuroendocrine tumors from a variety of anatomic sites, including 32 cases from the lung, 16 cases from the gastrointestinal tract, 20 cases from the female reproductive system, 3 cases from soft tissue, 2 cases from the head and neck region, and 1 case from the bladder. Using a set of primers optimized to detect MCPyV in formalin-fixed tissue, polymerase chain reaction (PCR)-based testing showed evidence of MCPyV DNA in only 1 of the 74 tumors; however, clinicopathologic review of the positive case (a neuroendocrine tumor of the small intestine) disclosed that the patient had a history of primary MCC of the buttock. PCR-based testing also showed no evidence of the related WU and KI polyomaviruses in the set of 74 cases. We conclude that, when evaluated by PCR-based testing, MCPyV is a specific marker for MCC that can be helpful in distinguishing cases of metastatic MCC from other high-grade neuroendocrine tumors. Our results also suggest that MCPyV does not have a role in the oncogenesis of visceral high-grade neuroendocrine tumors.

DNA from BK Virus and JC Virus and from KI, WU, and MC Polyomaviruses as Well as from Simian Virus 40 Is Not Detected in Non-UV-Light-Associated Primary Malignant Melanomas of Mucous Membranes

DNA from BK Virus and JC Virus and from KI, WU, and MC Polyomaviruses as Well as from Simian Virus 40 Is Not Detected in Non-UV-Light-Associated Primary Malignant Melanomas of Mucous Membranes

KI and WU Polyomaviruses in Patients Infected with HIV-1, Italy

To the Editor: Before 2007, two human polyomaviruses were known to infect humans: BK virus and JC virus (1,2). Recently, 2 novel polyomaviruses, KI polyomavirus (KIPyV) and WU polyomavirus (WUPyV), were identified in the respiratory secretions of children with signs of acute respiratory signs (3,4); little evidence exists to suggest that these viruses are causative agents of respiratory tract disease (5). To determine the prevalence of WUPyV and KIPyV in the plasma of HIV-1–infected patients, we screened 62 persons who were HIV-1 positive by using PCR to detect the 2 viruses. We also conducted phylogenetic analysis of the identified strains.

Serologic Evidence of Frequent Human Infection with WU and KI Polyomaviruses

WU polyomavirus (WUPyV) and KI polyomavirus (KIPyV) are novel human polyomaviruses. They were originally identified in human respiratory secretions, but the extent of human infection caused by these viruses has not been described to date. To determine the seroepidemiology of WUPyV and KIpyIV, we used an ELISA to screen serum samples from 419 patients at the St. Louis Children's Hospital and Barnes-Jewish Hospital during 2007-2008. The age-stratified deidentified samples were examined for antibodies to the major capsid proteins of WUPyV and KIPyV. Seropositivity for each virus was similar; antibody levels were high in the youngest age group (<6 months), decreased to a nadir in the next age group (6 to <12 months), and then steadily increased with subsequent age groups, eventually reaching a plateau of approximate, equals 80% for WUPyV and approximate, equals 70% for KIPyV. These results demonstrate that both KIPyV and WUPyV cause widespread infection in the human population.

Real Time PCR TaqMan assays for detection of polyomaviruses KIV and WUV in clinical samples

Recently, polyomaviruses KI and WU were identified in the airways of patients with acute respiratory symptoms. The epidemiology and pathogenesis of these two viruses are not fully understood, and the development of molecular assays, such as Real Time PCR, was useful for examining their biology and role in different clinical syndromes. The evaluation of different target regions for the amplification of polyomaviruses KI and WU, comparing published primer/probe sets and sets designed in the laboratory is described and was used for testing 175 clinical specimens (84 stools and 91 tonsils). The results showed that the laboratory designs were more sensitive for the detection of polyomaviruses KI and WU DNA in clinical samples. The choice of the primer/probe set, and primarily of the region for amplification, may be relevant for understanding the pathogenic role of viruses such as polyomaviruses KI and WU.

Excretion of the novel polyomaviruses KI and WU in the stool of patients with hematological disorders

Infection with human polyomaviruses BKV and JCV is asymptomatic, and lifelong and widespread, among the general population. However, in the setting of immunosuppression, secondary to medications or viral infection, for example, with HIV, reactivation can occur and result in severe disease. In this study, stool specimens from 31 patients with hematological disorders (25 transplanted and 6 non-transplanted) were examined prospectively to determine whether the novel polyomaviruses KIV and WUV reactivated and were excreted in the gastrointestinal tract. Reactivation was correlated with the appearance of gastrointestinal and respiratory symptoms. Of the 31 patients examined, KIV and WUV were detected in 13 transplanted patients as single infection or in combination with BKV, cytomegalovirus (CMV), and adenovirus (Adv). Because of frequent co-infections, a clear correlation between novel polyomaviruses and clinical symptoms could not be established. There was no correlation between demographic variables and detection of KIV and WUV. Phylogenetic analysis of the small t-antigen gene of KIV and WUV isolates showed that the novel polyomaviruses identified in feces clustered with those identified in the respiratory tract suggesting an oral-fecal transmission of these viruses. The novel polyomaviruses KI and WU may have a pathogenic role in immunocompromised patients.

Identification of the novel KI and WU polyomaviruses in human tonsils

Background Three novel polyomaviruses have been recently discovered: KI, WU and MC polyomaviruses. Their role in human pathology is debated while tissue tropism and site of latency remain unknown. Objective To test the hypothesis that KI, WU and MC polyomaviruses can infect human tonsils. Study design Archival paraffin-embedded tonsils from 91 patients affected by different tonsil diseases were screened by polymerase chain reaction to detect viral DNA of KIV, WUV, MCV, BKV and JCV. Phylogenetic and evolutionary analysis of the identified polyomaviruses was carried out. Results Of the 91 tested specimens, 11 contained KIV DNA (12%), 4 WUV DNA (4.4%), 5 BKV DNA (5.5%). MCV and JCV were not detected. Phylogenetic analysis showed that KIVs identified in tonsils fall into a clade distinct from that containing KIVs isolated from respiratory secretions, respiratory tissue and feces. Moreover, four positively selected sites (4.5% of t-Ag sites) were found under strong positive selection ( ω = 11.4), with posterior probabilities above 0.99. All the sites were located in the N-terminal region of the small t antigen. Conclusions The results suggest that the novel KI and WU polyomaviruses can infect human tonsils. Future studies are needed to define their role in tonsil diseases.

Detection of BK, JC, WU, or KI polyomaviruses in faecal, urine, blood, cerebrospinal fluid and respiratory samples

BackgroundThe recently described WU (WUV) and KI (KIV) polyomaviruses have been primarily detected in respiratory samples, however other body sites have not been extensively investigated to date. The related human polyomaviruses JCV and BKV in contrast, have been detected in a wide range of sample types, leading to increased knowledge about their biology and pathogenesis. ObjectivesThe aim of the study was to investigate and compare the presence of JCV, BKV, WUV, and KIV in a variety of patient samples. Study designNasopharyngeal aspirates (NPAs), bronchoalveolar lavages (BALs), cerebrospinal fluid (CSF), blood, faeces and urine from paediatric and adult immunocompetent and compromised patients were screened for the presence of the polyomaviruses by real-time PCR. The non-translated region (NTR) and VP1 of select WUV and KIV positive samples were sequenced and analysed. ResultsWUV and KIV were predominantly detected in NPA, BAL, and faeces from paediatric patients. JCV and BKV were primarily detected in blood, urine and faeces from adult patients. WUV and KIV NTR/VP1 sequence similarity ranged from 99.5% to 100% and 97.5–100%, respectively. ConclusionsOverall, WUV and KIV were detected in paediatric respiratory tract samples, in contrast to JCV and BKV, in which respiratory detections were uncommon. Additionally, the lack of WUV and KIV detections in blood, CSF, urine and adult faeces reinforces the parallel in divergent genomic phylogeny and apparent tissue tropism between JCV and BKV, and WUV and KIV. NTR/VP1 sequence variation did not appear to be associated with site of WUV or KIV detection.

Reactivation and Mutation of Newly Discovered WU, KI, and Merkel Cell Carcinoma Polyomaviruses in Immunosuppressed Individuals

Infection with the human polyomaviruses BK (BKV) and JC (JCV) is almost ubiquitous, asymptomatic, and lifelong. However, reactivation during immunosuppression, associated with mutations in the transcriptional control region (TCR) that up-regulates viral replication, can cause life-threatening disease. In this study, we investigated whether the recently discovered WU and KI polyomaviruses (WUPyV and KIPyV) and Merkel cell polyomavirus (MCPyV) could, like BKV and JCV, persist, mutate, and reactivate in immunodeficient subjects. Autopsy samples of lymphoid tissue from 42 AIDS-immunosuppressed subjects and 55 control samples were screened by polymerase chain reaction for all 5 polyomaviruses. TCR sequences from KIPyV and WUPyV recovered from both immunosuppressed and nonimmunosuppressed subjects were compared. Combined polyomavirus detection frequencies were much higher for the immunosuppressed group, compared with the nonimmunosuppressed group (35.7% vs. 3.6%), with viral loads in lymphoid tissues ranging from < or = 8.4 x 10(5) to > 1.5 x 10(5) viral genome copies per 10(6) cells. MCPyV was recovered from only 1 HIV-negative study subject. TCR sequences from reactivated WUPyV and KIPyV variants showed a number of point mutations and insertions that were absent in viruses recovered from respiratory tract specimens obtained from nonimmunosuppressed subjects. KIPyV and WUPyV show reactivation frequencies comparable to those of BKV and JCV during immunosuppression. TCR changes that potentially lead to transcriptional dysregulation may have pathogenic consequences equivalent in severity to those observed for JCV and BKV.

P.042 Comparative analysis of two Adenovirus TaqMan PCRs

Three novel polyomaviruses have been recently discovered: KI, WU and MC polyomaviruses. Their role in human pathology is debated while tissue tropism and site of latency remain unknown.To test the hypothesis that KI, WU and MC polyomaviruses can infect human tonsils.Archival paraffin-embedded tonsils from 91 patients affected by different tonsil diseases were screened by polymerase chain reaction to detect viral DNA of KIV, WUV, MCV, BKV and JCV. Phylogenetic and evolutionary analysis of the identified polyomaviruses was carried out.Of the 91 tested specimens, 11 contained KIV DNA (12%), 4 WUV DNA (4.4%), 5 BKV DNA (5.5%). MCV and JCV were not detected. Phylogenetic analysis showed that KIVs identified in tonsils fall into a clade distinct from that containing KIVs isolated from respiratory secretions, respiratory tissue and feces. Moreover, four positively selected sites (4.5% of t-Ag sites) were found under strong positive selection (ω = 11.4), with posterior probabilities above 0.99. All the sites were located in the N-terminal region of the small t antigen.The results suggest that the novel KI and WU polyomaviruses can infect human tonsils. Future studies are needed to define their role in tonsil diseases.

A single-tube, real-time PCR assay for detection of the two newly characterized human KI and WU polyomaviruses

Background Three new human polyomaviruses have been recently described, and investigating their in vivo biology and pathogenicity will require sensitive and rational detection assays. Objectives To develop and evaluate a sensitive and rational assay for detection of the newly identified KI and WU polyomaviruses. Study design A single-tube, dual-probe, real-time PCR assay for simultaneous detection and discrimination of KI and WU polyomaviruses was developed. Results The assay had near single-molecule sensitivity for both viruses and no cross-reactivity was observed. A panel of 637 nasopharyngeal aspirates was screened, resulting in a frequency of 1.4% for KIPyV and 1.3% for WUPyV. Conclusions The dual-probe assay provides a rational approach for further studies of KIPyV and WUPyV.

Prevalence and Pathogenicity of WU and KI Polyomaviruses in Children, the Netherlands

A longitudinal study in 2004 and 2005 detected polyomaviruses WU and KI in 44% and 17% of children with and without respiratory symptoms, respectively, in the Netherlands. In some children both viruses were detected for long periods. In several symptomatic children no other respiratory pathogen was detected.