Karin Gunst

Esophageal squamous cell cancer in patients with head and neck cancer: Prevalence of human papillomavirus DNA sequences.

An etiologic role for human papillomavirus (HPV) infections in either head and neck (HNC) or esophageal carcinogenesis remains debatable. Patients with head and neck cancer are at high risk for developing a second esophageal squamous cell cancer (ESCC). The aim of our study was to determine whether HPV infections play a role in this multifocal carcinogenesis. Samples from 2 groups of HNC patients were studied: Random esophageal biopsies were collected from the first group of 60 patients who had been screened for asymptomatic ESCC. The second group consisted of 21 patients with pairs of HNC and ESCC. Both the fresh frozen biopsy samples of the first group and the paraffin‐embedded specimens of the second group were evaluated for the presence of HPV DNA sequences by PCR amplification, cloning and sequencing. HPV DNA sequences were detected in 66.7% of normal/inflammatory (34/51) and dysplastic and malignant (6/9) esophageal tissues from HNC patients being screened endoscopically. Similarly, in the second group of 21 patients with both HNC and ESCC, HPV DNA sequences were demonstrated in 13 (61.9%) of the HNC biopsies and in 14 (66.7%) of the ESCC biopsies. The prevalence of high‐risk‐type HPV 16 was low (5/51, 9.8%) in normal/inflammatory esophageal mucosa but higher (10/24, 47.6%) in ESCC. The low‐risk HPV 11 was present in 37.3% (19/51) of normal/inflammatory, 66.7% (4/6) of dysplastic and 28.9% (13/45) of the carcinoma samples. The same HPV type was present in only 3/21 pairs of HNC and ESCC samples, suggesting that a clonal expansion from the HNC to a subsequent ESCC, or visa versa, is unlikely. The high prevalence of “low‐risk” HPV infections points to the need for studies on possible interactions of these infections with the use of alcohol and tobacco in the pathogenesis of these tumors.

In Vivo and In Vitro Intragenomic Rearrangement of TT Viruses

ABSTRACT The in vitro replication of the Torque teno virus (TT virus) tth8 full-length genome and particle formation in a Hodgkins lymphoma-derived cell line after transfection with cloned viral DNA were demonstrated. Analyses of the transcription patterns of tth8 and tth7 TT virus isolates in a number of lymphoma and T-cell leukemia cell lines indicated differential additional splicing events and intragenomic rearrangement generating open reading frames which could not be deducted from the genomic sequence. We also demonstrated the presence of rearranged TT virus genomes in vivo in sera taken from pregnant mothers whose children later developed childhood leukemia, as well as sera from control mothers. Control experiments using religated cloned genomic tth8 DNA mixed with cellular DNA did not result in such subviral molecules. These subviral isolates ranged from 172 bp to full-length TT virus genomes. Possible in vivo selection for specific rearranged molecules was indicated by the presence of one isolate (561 bp) in 11 serum samples. It remains to be clarified whether selected rearranged subviral components resulting from specific TT virus types may contribute to the initiation of disease. These data demonstrate new features of TT viruses suggesting possible similarities to plant viruses of the family Geminiviridae, as well as raise questions about the documented plurality and diversity of anelloviruses.

Characterization of seven novel human papillomavirus types isolated from cutaneous tissue, but also present in mucosal lesions.

Seven novel human papillomavirus (HPV) types were isolated and characterized. HPV 94 is related most closely to HPV 10 and belongs to the genus Alphapapillomavirus, whereas HPV 98, HPV 99, HPV 100, HPV 104, HPV 105 and HPV 113 all belong to the genus Betapapillomavirus. These HPV types were isolated from and demonstrated in cutaneous tissue, but HPV 98, HPV 100, HPV 104 and HPV 113 were also detected in malignant oesophageal and oral lesions. The general prevalence of these HPV types in lesions is infrequent.

The Diversity of Torque Teno Viruses: In Vitro Replication Leads to the Formation of Additional Replication-Competent Subviral Molecules

ABSTRACT The family Anelloviridae comprises torque teno viruses (TTVs) diverse in genome structure and organization. The isolation of a large number of TTV genomes (TTV Heidelberg [TTV-HD]) of 26 TTV types is reported. Several isolates from the same type indicate sequence variation within open reading frame 1 (ORF1), resulting in considerably modified open reading frames. We demonstrate in vitro replication of 12 full-length genomes of TTV-HD in 293TT cells. Propagation of virus was achieved by several rounds of infections using supernatant and frozen whole cells of initially infected cells. Replication of virus was measured by PCR amplification and transcription analyses. Subgenomic molecules (μTTV), arising early during propagation and ranging in size from 401 to 913 bases, were cloned and characterized. Propagation of these μTTV in in vitro cultures was demonstrated in the absence of full-length genomes.

Mycovirus-Like DNA Virus Sequences from Cattle Serum and Human Brain and Serum Samples from Multiple Sclerosis Patients

ABSTRACT Myco-like viruses have been isolated from fungi, feces of various animals, and plant leaves. We report here the isolation of 3 complete genome sequences of gemycircularvirus-related viruses from healthy bovine serum and human brain and serum samples from patients with multiple sclerosis (MS). Their putative capsid proteins share similarity to Torque teno virus (TTV) open reading frame 1 (ORF1) proteins.

Intragenomic rearrangement in TT viruses: a possible role in the pathogenesis of disease.

A role for the ubiquitous Torque teno (TT) viruses in the pathogenesis of disease has not been resolved. In vivo and in vitro intragenomic rearrangement of TT virus genomes has been demonstrated. Replication in cell culture of a subviral molecule (411 bp) occurs through oligomerisation of RNA transcripts. Although the functions of the respective TT viral genes, as well as the newly formed genes in the rearranged subviral molecules, are largely unknown, certain similarities to genes of plant viruses of the family Geminiviridae will be described. A degree of similarity to certain cellular genes poses the question as to a role of molecular mimicry in the pathogenesis of autoimmune disease and diabetes.

Novel replication-competent circular DNA molecules from healthy cattle serum and milk and multiple sclerosis-affected human brain tissue

ABSTRACT Epidemiological data point to the involvement of a cow milk factor in the etiology of multiple sclerosis (MS). Eleven circular DNA molecules closely related to transmissible spongiform encephalopathy (TSE)-associated isolate Sphinx 1.76 were isolated from healthy cattle serum, cow milk, and serum and brain tissue from MS patients.

Isolation of protein-associated circular DNA from healthy cattle serum.

ABSTRACT Three replication-competent single-stranded DNA molecules sharing nucleotide similarity to transmissible spongiform encephalopathy (TSE)-associated isolate Sphinx 2.36 were isolated from healthy bovine serum.

Isolation of Bacterial Plasmid-Related Replication-Associated Circular DNA from a Serum Sample of a Multiple Sclerosis Patient

ABSTRACT Psychrobacter species are considered to be opportunistic human pathogens. We report here the isolation of a circular DNA molecule, MSSI1.162, from a serum sample taken from a multiple sclerosis patient during relapse. This isolate is distantly related to known Psychrobacter species and their plasmids.

Expression and replication of virus-like circular DNA in human cells

The consumption of bovine milk and meat is considered a risk factor for colon- and breast cancer formation, and milk consumption has also been implicated in an increased risk for developing Multiple Sclerosis (MS). A number of highly related virus-like DNAs have been recently isolated from bovine milk and sera and from a brain sample of a MS patient. As a genetic activity of these Acinetobacter-related bovine milk and meat factors (BMMFs) is unknown in eukaryotes, we analyzed their expression and replication potential in human HEK293TT cells. While all analyzed BMMFs show transcriptional activity, the MS brain isolate MSBI1.176, sharing homology with a transmissible spongiform encephalopathy-associated DNA molecule, is transcribed at highest levels. We show expression of a replication-associated protein (Rep), which is highly conserved among all BMMFs, and serological tests indicate a human anti-Rep immune response. While the cow milk isolate CMI1.252 is replication-competent in HEK293TT cells, replication of MSBI1.176 is complemented by CMI1.252, pointing at an interplay during the establishment of persistence in human cells. Transcriptome profiling upon BMMF expression identified host cellular gene expression changes related to cell cycle progression and cell viability control, indicating potential pathways for a pathogenic involvement of BMMFs.