A case study of salivary microbiome in smokers and non-smokers in Hungary: analysis by shotgun metagenome sequencing

Objective To investigate the role of cigarette smoking in disease-development through altering the composition of the oral microbial community. Periodontitis and oral cancer are highly prevalent in Hungary; therefore, the salivary microbiome of smoker and non-smoker Hungarian adults was characterized. Methods Shotgun metagenome sequencing of salivary DNA samples from 22 individuals (11 non-smokers and 11 current smokers) was performed using the Ion Torrent PGMTM platform. Quality-filtered reads were analysed by both alignment-based sequence similarity searches and genome-centric binning. Results Prevotella, Veillonella and Streptococcus were the predominant genera in the saliva of both groups. Although the overall composition and diversity of the microbiota were similar, Prevotella was significantly more abundant in salivary samples of current smokers compared to non-smokers. Members of the genus Prevotella were implicated in the development of inflammatory diseases and oral cancer. The abundance of the genus Megasphaera also increased in current smokers, whereas the genera Neisseria, Oribacterium, Capnocytophaga and Porphyromonas were significantly reduced. The data generated by read-based taxonomic classification and genome-centric binning mutually validated the two distinct metagenomic approaches. Conclusion Smoking-associated dysbiosis of the salivary microbiome in current cigarette smokers, especially increased abundance of Prevotella and Megasphaera genera, may facilitate disease development

A case study of salivary microbiome in smokers and non-smokers in Hungary: analysis by shotgun metagenome sequencing

Objective To investigate the role of cigarette smoking in disease-development through altering the composition of the oral microbial community. Periodontitis and oral cancer are highly prevalent in Hungary; therefore, the salivary microbiome of smoker and non-smoker Hungarian adults was characterized. Methods Shotgun metagenome sequencing of salivary DNA samples from 22 individuals (11 non-smokers and 11 current smokers) was performed using the Ion Torrent PGMTM platform. Quality-filtered reads were analysed by both alignment-based sequence similarity searches and genome-centric binning. Results Prevotella, Veillonella and Streptococcus were the predominant genera in the saliva of both groups. Although the overall composition and diversity of the microbiota were similar, Prevotella was significantly more abundant in salivary samples of current smokers compared to non-smokers. Members of the genus Prevotella were implicated in the development of inflammatory diseases and oral cancer. The abundance of the genus Megasphaera also increased in current smokers, whereas the genera Neisseria, Oribacterium, Capnocytophaga and Porphyromonas were significantly reduced. The data generated by read-based taxonomic classification and genome-centric binning mutually validated the two distinct metagenomic approaches. Conclusion Smoking-associated dysbiosis of the salivary microbiome in current cigarette smokers, especially increased abundance of Prevotella and Megasphaera genera, may facilitate disease development

Epigenetic dysregulation in virus-associated neoplasms

The oncoproteins of human tumor viruses regularly interact with the cellular epigenetic machinery. Such interactions alter the epigenome of the host cell and reprogram its gene expression pattern. Altered levels or redistribution of (cytosine-5)-DNA methyltransferases and changes in the cellular methylome were observed in Kaposi sarcoma-associated herpesvirus (KSHV), hepatitis B virus (HBV), hepatitis D virus (HDV), hepatitis C virus (HCV), and human papillomavirus (HPV) associated neoplasms and cell lines. Methylation-mediated silencing of cellular promoters was also noted in Merkel cell polyomavirus (MCPyV) positive Merkel cell carcinomas, and, as discussed elsewhere, in EBV-associated malignancies and adenovirus-induced rodent tumors as well. Promoter activation also occurred, either associated with DNA hypomethylation or with the induction of euchromatic histone modifi cations by viral oncoproteins. It is worthy to notice that HCV infection induced large, hypomethylated blocks of cellular chromatin, although the exact molecular mechanism remains to be elucidated. In hepatoma cells expressing HBx, the oncoprotein encoded by the HBV genome, demethylation of the repetitive satellite 2 sequences was observed, due to downregulation of the de novo DNA methyltransferase DNMT3B. Tax and HBZ, the oncoproteins of human T-cell lymphotropic virus type I (HTLV-I), can both activate and silence distinct cellular promoters by interacting with cellular enzymes involved in histone modification. © Springer International Publishing Switzerland 2016

Wild type HBx and truncated HBx: Pleiotropic regulators driving sequential genetic and epigenetic steps of hepatocarcinogenesis and progression of HBV-associated neoplasms

Hepatitis B virus (HBV) is one of the causative agents of hepatocellular carcinoma. The molecular mechanisms of tumorigenesis are complex. One of the host factors involved is apparently the long-lasting inflammatory reaction which accompanies chronic HBV infection. Although HBV lacks a typical viral oncogene, the HBx gene encoding a pleiotropic regulatory protein emerged as a major player in liver carcinogenesis. Here we review the tumorigenic functions of HBx with an emphasis on wild type and truncated HBx variants, and their role in the transcriptional dysregulation and epigenetic reprogramming of the host cell genome. We suggest that HBx acquired by the HBV genome during evolution acts like a cellular proto-onc gene that is activated by deletion during hepatocarcinogenesis. The resulting viral oncogene (v-onc gene) codes for a truncated HBx protein that facilitates tumor progression. Copyright (c) 2015 John Wiley & Sons, Ltd

The Formation of Amyloid-Like Fibrils of alpha-Chymotrypsin in Different Aqueous Organic Solvents

The formation of amyloid-like fibrils of alpha-chymotrypsin was studied in aqueous ethanol, methanol, tert-butanol, dimethylformamide and acetonitrile. Thioflavin T (ThT), Congo red (CR) and 1-anilino-8-naphthalenesulfonic acid (ANS) binding, turbidity, intrinsic fluorescence and far-UV circular dichroism measurements were employed to characterize the amyloid fibril formation. The greatest extent of fibril formation after incubation for 24 h at pH 7.0 and at 24 degrees C was in ethanol at 55%, in methanol and dimethylformamide (DMF) at 60-70% and in tert-butanol at 60-80%. The ANS binding and intrinsic fluorescence results showed that the hydrophobic residues are more solvent-exposed in the aggregated form of alpha-chymotrypsin. The ThT, CR binding and far-UV CD measurements indicated that the formation of the cross-beta structure of alpha-chymotrypsin depends on the polarity of the organic solvent. To determine the role of surface charges in the aggregation, chemically modified forms of alpha-chymotrypsin were prepared. The citraconylated and succinylated enzymes exhibited a higher and the enzyme forms modified with aliphatic aldehydes a lower propensity for aggregation. These results suggest the important role of surface charges in the aggregation of alpha-chymotrypsin

Detection and Phylogenetic Analysis of Torque Teno Virus in Salivary and Tumor Biopsy Samples from Head and Neck Carcinoma Patients

Objectives: Because torque teno virus (TTV) has been implicated in tumorigenesis as a cocarcinogen, we studied TTV prevalence in saliva and biopsy samples from head and neck cancer (HNCC) patients, patients with premalignant lesions of oral cancer, and controls. We also wished to determine the TTV genotypes in HNCC patients. Methods: A seminested polymerase chain reaction (PCR) amplifying the N22 region of the TTV genome, as well as direct sequencing of PCR fragments, was used. Results: TTV prevalence was higher in HNCC patients (saliva: 27/71, 38%; tumor biopsy: 22/74, 30%) than in controls (saliva: 8/56, 14%; oral mucosa: 1/19, 5%). TTV prevalence was also high in patients with premalignant lesions of oral carcinoma (saliva: 9/18, 50%; biopsy: 5/21, 24%). By phylogenetic analysis, TTV belonging mostly to genotypes 1 and 2 was found in HNCC patients. In most of the cases, identical TTV strains were present in the biopsy and salivary sample of the same HNCC patient. In addition, the same TTV strain was detected in 2 laryngeal carcinoma biopsies obtained from 2 independent patients. Conclusions: Our data are compatible with the idea that TTV might act as a cocarcinogen in certain cases of HNCC. Alternatively, HNCC may facilitate either TTV replication or TTV entry into the saliva. © 2016 S. Karger AG, Base

Infectious Agents Associated with Head and Neck Carcinomas

In addition to traditional risk factors such as smoking habits and alcohol consumption, certain microbes also play an important role in the generation of head and neck carcinomas. Infection with high-risk human papillomavirus types is strongly associated with the development of oropharyngeal carcinoma, and Epstein-Barr virus appears to be indispensable for the development of non-keratinizing squamous cell carcinoma of the nasopharynx. Other viruses including torque teno virus and hepatitis C virus may act as co-carcinogens, increasing the risk of malignant transformation. A shift in the composition of the oral microbiome was associated with the development of oral squamous cell carcinoma, although the causal or casual role of oral bacteria remains to be clarified. Conversion of ethanol to acetaldehyde, a mutagenic compound, by members of the oral microflora as well as by fungi including Candida albicans and others is a potential mechanism that may increase oral cancer risk. In addition, distinct Candida spp. also produce NBMA (N-nitrosobenzylmethylamine), a potent carcinogen. Inflammatory processes elicited by microbes may also facilitate tumorigenesis in the head and neck region

Prevalence and genotypes of human papillomavirus in saliva and tumor samples of head and neck cancer patients in Hungary

Abstract In addition to traditional risk factors such as smoking, alcohol consumption and betel nut use, human papillomavirus (HPV) infection also plays a role in the development of head and neck squamous cell carcinomas (HNSCCs). Although among European countries the highest incidence and mortality rates of head and neck cancer types were recorded in Hungary, data regarding HPV prevalence in HNSCCs is scarce. We collected biopsy and saliva samples from patients diagnosed with HNSCC or oral potentially malignant disorders (OPMDs) and tested them for the presence of HPV using the PCR consensus primer set MY09/11 and the GP5+/6+ primer pair. HPV genotypes were assessed by sequencing of the amplified PCR fragments. Oral mucosa and saliva samples from tumor- and OPMD-free individuals were also analysed. HPV was detected in 11 out of 60 HNSCC samples (18%). All of the HPV positive tumors carried HPV type 16. 5 out of the 57 saliva samples collected from HNSCC patients was HPV positive (8.8%); among them, in addition to HPV16, HPV13 was also detected. Tumors located to the oropharynx had the highest HPV positivity rate with 50% (7 out of 14), which was significantly higher than the HPV prevalence in oral mucosa samples collected from controls (0 out of 20; p > 0.001) or in OPMD biopsies (0 out of 21, p > 0.001). 2 out of 57 control saliva samples (3.5%, subtype HPV13 and 11) and 3 out of 39 saliva samples from OPMD patients (7.7%, subtype HPV18, 81 and 10) were HPV positive. Our data suggested that HPV16 infection may contribute, in concert with cigarette smoking, to the development of a subset of head and neck cancers in Hungary. HPV16 infection per se does not account, however, for the high HNSCC incidence rate recorded in this country