DNA affinity purification sequencing and transcriptional profiling reveal new aspects of nitrogen regulation in a filamentous fungus.

Sensing available nutrients and efficiently utilizing them is a challenge common to all organisms. The model filamentous fungus Neurospora crassa is capable of utilizing a variety of inorganic and organic nitrogen sources. Nitrogen utilization in N. crassa is regulated by a network of pathway-specific transcription factors that activate genes necessary to utilize specific nitrogen sources in combination with nitrogen catabolite repression regulatory proteins. We identified an uncharacterized pathway-specific transcription factor, amn-1, that is required for utilization of the nonpreferred nitrogen sources proline, branched-chain amino acids, and aromatic amino acids. AMN-1 also plays a role in regulating genes involved in responding to the simple sugar mannose, suggesting an integration of nitrogen and carbon metabolism. The utilization of nonpreferred nitrogen sources, which require metabolic processing before being used as a nitrogen source, is also regulated by the nitrogen catabolite regulator NIT-2. Using RNA sequencing combined with DNA affinity purification sequencing, we performed a survey of the role of NIT-2 and the pathway-specific transcription factors NIT-4 and AMN-1 in directly regulating genes involved in nitrogen utilization. Although previous studies suggested promoter binding by both a pathway-specific transcription factor and NIT-2 may be necessary for activation of nitrogen-responsive genes, our data show that pathway-specific transcription factors regulate genes involved in the catabolism of specific nitrogen sources, while NIT-2 regulates genes involved in utilization of all nonpreferred nitrogen sources, such as nitrogen transporters. Together, these transcription factors form a nutrient sensing network that allows N. crassa cells to regulate nitrogen utilization.

Classifying Studies on Workplace Violence for Visiting Nurses Using the Social-Ecological Model: A Scoping Review.

PURPOSE: To prevent workplace violence (WV) against visiting nurses (VNs), understanding the influencing factors is crucial. To better comprehend potential violence prevention strategies, the U.S. Centers for Disease Control and Prevention has presented the four-level social-ecological model (SEM) at the 1) individual, 2) relationship, 3) community, and 4) societal levels. This study aims to quantify WV studies against VNs, examine the influencing factors, identify gaps based on SEM levels, and propose future research and policy directions. DESIGN: A scoping review was conducted following the five-stage protocol proposed by Arksey and O'Malley in 2005. METHODS: Systematic searches, including manual searches, were performed using English and Korean databases. Published journal articles including editorials on WV against VNs were included, irrespective of the publication date. FINDINGS: Sixty journal articles were finally selected. Until the 1990s, most of the literature comprised editorials, with empirical research emerging after the 2000s. Classifying studies by SEM level, many studies have focused on individual (86.7%) and community (66.7%) factors, but fewer have addressed relationship (21.7%) and societal (16.7%) factors. CONCLUSIONS: The study revealed that much research has focused on individual-centered training, with gaps in risk assessment tool development, training incorporating relational aspects, standardized protocols, and understanding of the impact of legal rights and policies. This article advocates a comprehensive approach that considers all SEM levels to address WV against VNs. CLINICAL EVIDENCE: The findings confirm a research gap, which suggests the direction for future research and policies. Stakeholders should be urged to implement evidence-based strategies that contribute to safer work environments for VNs.

Development of a parent questionnaire to assess treatment adherence for a child or adolescent with epilepsy.

PURPOSE: This study aimed to develop a questionnaire to assess parents' treatment adherence for their child or adolescent with epilepsy (PQ-TAE). MATERIAL AND METHODS: Seventy-one preliminary items were developed as a first draft and refined based on reviews from experts and cognitive interviews with parents. Then, an online survey was conducted from June 15 to July 15, 2020, to test the psychometric properties of the questionnaire and to modify it. Reliability and validity were additionally tested using exploratory factor analysis, internal consistency, and statistical relationships including parents' partnership with healthcare providers, educational attainment, and economic status. RESULTS: The 4-factor model was adopted as the final factor structure of the PQ-TAE (CMIN/df = 2.15, RMSEA = 0.07, SRMR = 0.04, CFI = 0.93, TLI = 0.90). The reliability of the PQ-TAE was statistically acceptable (Cronbach's alpha = 0.93). The PQ-TAE score was statistically related to the partnership with healthcare providers (r = 0.58, p < 0.001), educational attainment (t = -2.12, p = 0.037), and economic status (t = -4.05, p < 0.001). CONCLUSIONS: The PQ-TAE using a 5-point Likert scale consists of 25 items classified into four factors. The higher the score (Range 25-125), the better the treatment adherence of the parent of a child or adolescent with epilepsy.

The regulatory and transcriptional landscape associated with carbon utilization in a filamentous fungus.

Filamentous fungi, such as Neurospora crassa, are very efficient in deconstructing plant biomass by the secretion of an arsenal of plant cell wall-degrading enzymes, by remodeling metabolism to accommodate production of secreted enzymes, and by enabling transport and intracellular utilization of plant biomass components. Although a number of enzymes and transcriptional regulators involved in plant biomass utilization have been identified, how filamentous fungi sense and integrate nutritional information encoded in the plant cell wall into a regulatory hierarchy for optimal utilization of complex carbon sources is not understood. Here, we performed transcriptional profiling of N. crassa on 40 different carbon sources, including plant biomass, to provide data on how fungi sense simple to complex carbohydrates. From these data, we identified regulatory factors in N. crassa and characterized one (PDR-2) associated with pectin utilization and one with pectin/hemicellulose utilization (ARA-1). Using in vitro DNA affinity purification sequencing (DAP-seq), we identified direct targets of transcription factors involved in regulating genes encoding plant cell wall-degrading enzymes. In particular, our data clarified the role of the transcription factor VIB-1 in the regulation of genes encoding plant cell wall-degrading enzymes and nutrient scavenging and revealed a major role of the carbon catabolite repressor CRE-1 in regulating the expression of major facilitator transporter genes. These data contribute to a more complete understanding of cross talk between transcription factors and their target genes, which are involved in regulating nutrient sensing and plant biomass utilization on a global level.

Development and evaluation of a problem-based learning simulation module for home-visit nursing.

OBJECTIVES: Although home-visit healthcare programs in Korea are expected to expand, providing hands-on experience to nursing students may be limited. This study aimed to develop and evaluate a problem-based learning (PBL) simulation module that reflects home-visit healthcare services provided by public health centers for pre-frail older adults. DESIGN AND SAMPLE: The simulation module, including PBL as prebriefing, was developed by the researchers and revised based on expert reviews. The module was evaluated using a mixed-method embedded one-group post-test-only design with focus group interviews (FGIs). Quantitative data (n = 29) were collected between April and June, 2021. FGIs (n = 10) were conducted twice in June 2021, and qualitative data were analyzed using an inductive content analysis approach. RESULTS: The average score of the Simulation Design Scale was 4.67 ± 0.36. The overall mean score of the Educational Practices Questionnaire was 4.75 ± 0.37. Three themes emerged from the FGIs: immersive learning experience, changes in perspective on nursing, and enhanced nursing competency. CONCLUSION: This PBL-based simulation module was evaluated as a systematic learning process in which nursing students could become self-directed learners, interacting and collaborating with colleagues, instructors, and environments. The module encourages them to practice home visit services.

Broad Genomic Sampling Reveals a Smut Pathogenic Ancestry of the Fungal Clade Ustilaginomycotina.

Ustilaginomycotina is home to a broad array of fungi including important plant pathogens collectively called smut fungi. Smuts are biotrophs that produce characteristic perennating propagules called teliospores, one of which, Ustilago maydis, is a model genetic organism. Broad exploration of smut biology has been hampered by limited phylogenetic resolution of Ustilaginiomycotina as well as an overall lack of genomic data for members of this subphylum. In this study, we sequenced eight Ustilaginomycotina genomes from previously unrepresented lineages, deciphered ordinal-level phylogenetic relationships for the subphylum, and performed comparative analyses. Unlike other Basidiomycota subphyla, all sampled Ustilaginomycotina genomes are relatively small and compact. Ancestral state reconstruction analyses indicate that teliospore formation was present at the origin of the subphylum. Divergence time estimation dates the divergence of most extant smut fungi after that of grasses (Poaceae). However, we found limited conservation of well-characterized genes related to smut pathogenesis from U. maydis, indicating dissimilar pathogenic mechanisms exist across other smut lineages. The genomes of Malasseziomycetes are highly diverged from the other sampled Ustilaginomycotina, likely due to their unique history as mammal-associated lipophilic yeasts. Despite extensive genomic data, the phylogenetic placement of this class remains ambiguous. Although the sampled Ustilaginomycotina members lack many core enzymes for plant cell wall decomposition and starch catabolism, we identified several novel carbohydrate active enzymes potentially related to pectin breakdown. Finally, ∼50% of Ustilaginomycotina species-specific genes are present in previously undersampled and rare lineages, highlighting the importance of exploring fungal diversity as a resource for novel gene discovery.

Correction: Heated tobacco product use and its relationship to quitting combustible cigarettes in Korean adults.

[This corrects the article DOI: 10.1371/journal.pone.0251243.].

Cigarette smoke induces promoter methylation of single-stranded DNA-binding protein 2 in human esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is the sixth most frequent cause of cancer death in the world, and cigarette smoke is a key factor in esophageal carcinogenesis. To identify molecular changes during cigarette smoke-induced ESCC, we examined the methylation status of 13 gene promoters in the human immortalized, nontumorigenic esophageal epithelial cell line (Het-1A) that were exposed to mainstream (MSE) or sidestream cigarette smoke extract (SSE) for 6 months in culture. The promoter of sequence-specific single-stranded DNA-binding protein 2 (SSBP2) was methylated in the Het-1A cells exposed to MSE (MSE-Het-1A). Promoter methylation (86%, 56/70) and downregulation of SSBP2 expression were frequently detected in tumor tissues from ESCC patients. In addition, reintroduction of SSBP2 in an ESCC cell line (TE1) that does not express SSBP2 and in the MSE-Het-1A cells inhibited expression of LRP6 and Dvl3, which are mediators of the Wnt signaling pathway. SSBP2 expression markedly decreased the colony-forming ability of ESCC cell lines and significantly inhibited cell growth of the MSE-Het-1A cells. Our results indicate that cigarette smoking is a cause of SSBP2 promoter methylation and that SSBP2 harbors a tumor suppressive role in ESCC through inhibition of the Wnt signaling pathway.

DNA methylation markers in colorectal cancer.

Colorectal cancer (CRC) arises as a consequence of the accumulation of genetic and epigenetic alterations in colonic epithelial cells during neoplastic transformation. Epigenetic modifications, particularly DNA methylation in selected gene promoters, are recognized as common molecular alterations in human tumors. Substantial efforts have been made to determine the cause and role of aberrant DNA methylation ("epigenomic instability") in colon carcinogenesis. In the colon, aberrant DNA methylation arises in tumor-adjacent, normal-appearing mucosa. Aberrant methylation also contributes to later stages of colon carcinogenesis through simultaneous methylation in key specific genes that alter specific oncogenic pathways. Hypermethylation of several gene clusters has been termed CpG island methylator phenotype and appears to define a subgroup of colon cancer distinctly characterized by pathological, clinical, and molecular features. DNA methylation of multiple promoters may serve as a biomarker for early detection in stool and blood DNA and as a tool for monitoring patients with CRC. DNA methylation patterns may also be predictors of metastatic or aggressive CRC. Therefore, the aim of this review is to understand DNA methylation as a driving force in colorectal neoplasia and its emerging value as a molecular marker in the clinic.

Presence of 5-methylcytosine in CpNpG trinucleotides in the human genome.

While the methylation machinery of mammalian cells has been shown to be capable of both maintenance and de novo methylation at CpNpG sites, CpNpG methylation in the human genome has not been demonstrated. Here, we report the first observation of 5-methylcytosines in CpNpG triplets in the human genome. We identify the existence of CpNpG methylation in a number of genes which contain trinucleotide repeat regions, including the androgen receptor (AR). We further analyzed DNA extracted from primary tissue samples and found the same pattern of CpNpG methylation. To confirm our results, we performed Southern blot analysis by analyzing the cleavage sites of restriction enzymes within exon 1 of the AR gene and found direct evidence of the presence of 5mCs in CpNpG triplets in the human genome. Our results also suggest that this methylation pattern may be due to the human DNA methyltransferases DNMT1 and DNMT3A. Although the functional significance needs to be tested further, the discovery of inheritable CpNpG methylation in the human genome may have important implications in our understanding of gene regulation and of the development of various diseases, including cancer.

Validation of the Korean Version of Impact of Event Scale-Revised (IES-R) in Korean Nurses during the COVID-19 Pandemic.

Nurses working amid the COVID-19 pandemic are at increased risk of developing post-traumatic stress disorder (PTSD). This study was conducted to verify the reliability and validity of the Korean version of Impact of Event Scale-Revised (IES-R), one of the most used tools for assessing trauma. Secondary data of 249 nurses who performed face-to-face nursing tasks during the COVID-19 pandemic, collected through an online survey, were analyzed by conducting a factor analysis of the K-IES-R and testing the internal consistency and concurrent validity with the Perceived Stress Scale (PSS), Generalized Anxiety Disorder Screener (GAD-7), and Dimensions of Anger Reactions-5 (DAR-5). The result of an exploratory factor analysis of the K-IES-R supported a three-factor structure of intrusion, avoidance, and sleep disturbance, with CMIN/DF = 2.98, RMSEA = 0.09, SRMR = 0.03, CFI = 0.93, and TLI = 0.90. The Cronbach's alpha of each subscale was 0.88-0.94. The total K-IES-R score and each factor's value showed a significant correlation (moderate or higher) with the PSS, GAD-7, and DAR-5. The K-IES-R was verified as a useful tool for assessing post-traumatic stress symptoms in nurses who directly perform nursing tasks in crises such as COVID-19. This study suggests the tool be used for early assessment of post-traumatic stress symptoms in nurses and providing appropriate interventions.

Heated tobacco product use and its relationship to quitting combustible cigarettes in Korean adults.

OBJECTIVE: We assessed the prevalence of, and factors associated with, heated tobacco product (HTP) use and analysed the association between HTP use and quitting combustible cigarettes (CCs) in Korean adults. METHODS: We conducted an online survey with 7,000 adults (males, 2,300; females, 4,700; ages 20-69) out of 70,000 age-, sex- and provincial-distribution-matched individuals based on 2018 national population statistics. Females were oversampled because the prevalence of tobacco product use is very low among women in Korea. Chi-square tests were used for bivariate analyses, and odds ratios were assessed after adjusting for sociodemographic variables. RESULTS: The prevalence of current CC, electronic cigarette (EC), and HTP use was 24.8% (males, 40.4%; females, 9.3%), 6.8% (males, 10.1%; females, 3.4%), and 10.2% (males, 16.2%; females, 4.3%), respectively. Among the 574 current HTP users, 77 (13.4%) were HTP-only users and >80% were either dual users of HTP and CC/EC, or triple users of HTP, EC, and CC. Among the current CC users, the odds of having attempted to quit CCs in the past year were greater among EC-only users (aOR 2.92; 95% CI 1.81-4.69) and dual users of HTPs and ECs (aOR 8.42; 95% CI 4.85-14.62) than among non-HTP and non-EC users. Among 2,121 ever CC smokers, the likelihood of being a former CC smoker was 0.19 (95% CI 0.15-0.24) for HTP users, 0.29 (95% CI 0.20-0.42) for EC users, and 0.03 (95% CI 0.01-0.06) for users of both HTPs and ECs compared with non-HTP and non-EC users. CONCLUSION: EC-only use and dual use of HTPs and ECs were associated with increased attempts to quit CCs; however, HTP and EC use was associated with lower odds of CC smoking abstinence.

Aspects of the Neurospora crassa Sulfur Starvation Response Are Revealed by Transcriptional Profiling and DNA Affinity Purification Sequencing.

Accurate nutrient sensing is important for rapid fungal growth and exploitation of available resources. Sulfur is an important nutrient source found in a number of biological macromolecules, including proteins and lipids. The model filamentous fungus Neurospora crassa is capable of utilizing sulfur found in a variety of sources from amino acids to sulfate. During sulfur starvation, the transcription factor CYS-3 is responsible for upregulation of genes involved in sulfur uptake and assimilation. Using a combination of RNA sequencing and DNA affinity purification sequencing, we performed a global survey of the N. crassa sulfur starvation response and the role of CYS-3 in regulating sulfur-responsive genes. The CYS-3 transcription factor bound the promoters and regulated genes involved in sulfur metabolism. Additionally, CYS-3 directly activated the expression of a number of uncharacterized transporter genes, suggesting that regulation of sulfur import is an important aspect of regulation by CYS-3. CYS-3 also directly regulated the expression of genes involved in mitochondrial electron transfer. During sulfur starvation, genes involved in nitrogen metabolism, such as amino acid and nucleic acid metabolic pathways, along with genes encoding proteases and nucleases that are necessary for scavenging nitrogen, were activated. Sulfur starvation also caused changes in the expression of genes involved in carbohydrate metabolism, such as those encoding glycosyl hydrolases. Thus, our data suggest a connection between sulfur metabolism and other aspects of cellular metabolism. IMPORTANCE Identification of nutrients present in the environment is a challenge common to all organisms. Sulfur is an important nutrient source found in proteins, lipids, and electron carriers that are required for the survival of filamentous fungi such as Neurospora crassa. Here, we transcriptionally profiled the response of N. crassa to characterize the global response to sulfur starvation. We also used DNA affinity purification sequencing to identify the direct downstream targets of the transcription factor responsible for regulating genes involved in sulfur uptake and assimilation. Along with genes involved in sulfur metabolism, this transcription factor regulated a number of uncharacterized transporter genes and genes involved in mitochondrial electron transfer. Our data also suggest a connection between sulfur, nitrogen, and carbon metabolism, indicating that the regulation of a number of metabolic pathways is intertwined.

Cellular transformation by cigarette smoke extract involves alteration of glycolysis and mitochondrial function in esophageal epithelial cells.

Cigarette-smoking increases the risk of developing various types of human cancers including esophageal cancers. To test the effects of chronic cigarette smoke exposure directly on esophageal epithelium, cellular resistance to mainstream extract (MSE), or sidestream smoke extract (SSE) was developed in chronically exposed nonmalignant Het-1A cells. Anchorage-independent growth, in vitro invasion capacity and proliferation of the resistant cells increased compared with the unexposed, sensitive cells. An epithelial marker E-cadherin was down-regulated and mesenchymal markers N-cadherin and vimentin were up-regulated in the resistant cells. Het-1A cells resistant to MSE or SSE consumed more glucose, and produced more lactate than the sensitive cells. The increased anchorage-independent cell growth of the resistant cells was suppressed by a glycolysis inhibitor, 2-deoxy-D-glucose, indicating that these cells are highly dependent on the glycolytic pathway for survival. Decreased mitochondrial membrane potential and ATP production in the resistant cells indicate the presence of mitochondrial dysfunction induced by chronic exposure of cigarette smoke extract. Increased expression of nuclear genes in the glycolytic pathway and decreased levels of mitochondrial genes in the resistant cells support the notion that cigarette smoking significantly contributes to the transformation of nonmalignant esophageal epithelial cells into a tumorigenic phenotype.

Author Correction: Genome sequencing of Rigidoporus microporus provides insights on genes important for wood decay, latex tolerance and interspecific fungal interactions.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Genome sequencing of Rigidoporus microporus provides insights on genes important for wood decay, latex tolerance and interspecific fungal interactions.

Fungal plant pathogens remain a serious threat to the sustainable agriculture and forestry, despite the extensive efforts undertaken to control their spread. White root rot disease is threatening rubber tree (Hevea brasiliensis) plantations throughout South and Southeast Asia and Western Africa, causing tree mortality and severe yield losses. Here, we report the complete genome sequence of the basidiomycete fungus Rigidoporus microporus, a causative agent of the disease. Our phylogenetic analysis confirmed the position of R. microporus among the members of Hymenochaetales, an understudied group of basidiomycetes. Our analysis further identified pathogen's genes with a predicted role in the decay of plant cell wall polymers, in the utilization of latex components and in interspecific interactions between the pathogen and other fungi. We also detected putative horizontal gene transfer events in the genome of R. microporus. The reported first genome sequence of a tropical rubber tree pathogen R. microporus should contribute to the better understanding of how the fungus is able to facilitate wood decay and nutrient cycling as well as tolerate latex and utilize resinous extractives.

Role of human aquaporin 5 in colorectal carcinogenesis.

While overexpression of several aquaporins (AQPs) has been reported in different types of human cancer, the role of AQPs in carcinogenesis has not been clearly defined. Here, by immunochemistry, we have found expression of AQP5 protein in 62.8% (59/94) of resected colon cancer tissue samples as well as association of AQP5 with liver metastasis. We then demonstrated that overexpression of human AQP5 (hAQP5) induces cell proliferation in colon cancer cells. Overexpression of wild-type hAQP5 increased proliferation and phosphorylation of extracellular signal-regulated kinase-1/2 in HCT116 colon cancer cells whereas these phenomena in hAQP5 mutants (N185D and S156A) were diminished, indicating that both membrane association and serine/threonine phosphorylation of AQP5 are required for proper function. Interestingly, overexpression of AQP1 and AQP3 showed no differences in extracellular signal-regulated kinase-1/2 phosphorylation, suggesting that AQP5, unlike AQP1, may be involved in signal transduction. Moreover, hAQP5-overexpressing cells showed an increase in retinoblastoma protein phosphorylation through the formation of a nuclear complex with cyclin D1 and CDK4. Small interfering RNA analysis confirmed that hAQP5 activates the Ras signaling pathway. These data not only describe the induction of hAQP5 expression during colorectal carcinogenesis but also provide a molecular mechanism for colon cancer development through the interaction of hAQP5 with the Ras/extracellular signal-regulated kinase/retinoblastoma protein signaling pathway, identifying hAQP5 as a novel therapeutic target.

ssDNA-binding protein 2 is frequently hypermethylated and suppresses cell growth in human prostate cancer.

PURPOSE: Prostate cancer is a major cause of cancer death among men and the development of new biomarkers is important to augment current detection approaches. EXPERIMENTAL DESIGN: We identified hypermethylation of the ssDNA-binding protein 2 (SSBP2) promoter as a potential DNA marker for human prostate cancer based on previous bioinformatics results and pharmacologic unmasking microarray. We then did quantitative methylation-specific PCR in primary prostate cancer tissues to confirm hypermethylation of the SSBP2 promoter, and analyzed its correlation with clinicopathologic data. We further examined SSBP2 expression in primary prostate cancer and studied its role in cell growth. RESULTS: Quantitative methylation-specific PCR results showed that the SSBP2 promoter was hypermethylated in 54 of 88 (61.4%) primary prostate cancers versus 0 of 23 (0%) in benign prostatic hyperplasia using a cutoff value of 120. Furthermore, we found that expression of SSBP2 was down-regulated in primary prostate cancers and cancer cell lines. Hypermethylation of the SSBP2 promoter and its expression were closely associated with higher stages of prostate cancer. Reactivation of SSBP2 expression by the demethylating agent 5-aza-2'-deoxycytidine in prostate cancer cell lines confirmed epigenetic inactivation as one major mechanism of SSBP2 regulation. Moreover, forced expression of SSBP2 inhibited prostate cancer cell proliferation in the colony formation assay and caused cell cycle arrest. CONCLUSION: SSBP2 inhibits prostate cancer cell proliferation and seems to represent a novel prostate cancer-specific DNA marker, especially in high stages of human prostate cancer.

Evaluation of promoter hypermethylation detection in body fluids as a screening/diagnosis tool for head and neck squamous cell carcinoma.

PURPOSE: To evaluate aberrant promoter hypermethylation of candidate tumor suppressor genes as a means to detect epigenetic alterations specific to solid tumors, including head and neck squamous cell carcinoma (HNSCC). EXPERIMENTAL DESIGN: Using promoter regions identified via a candidate gene and discovery approach, we evaluated the ability of an expanded panel of CpG-rich promoters known to be differentially hypermethylated in HNSCC in detection of promoter hypermethylation in serum and salivary rinses associated with HNSCC. We did preliminary evaluation via quantitative methylation-specific PCR (Q-MSP) using a panel of 21 genes in a limited cohort of patients with HNSCC and normal controls. Using sensitivity and specificity for individual markers as criteria, we selected panels of eight and six genes, respectively, for use in salivary rinse and serum detection and tested these in an expanded cohort including up to 211 patients with HNSCC and 527 normal controls. RESULTS: Marker panels in salivary rinses showed improved detection when compared with single markers, including a panel with 35% sensitivity and 90% specificity and a panel with 85% sensitivity and 30% specificity. A similar pattern was noted in serum panels, including a panel with 84.5% specificity with 50.0% sensitivity and a panel with sensitivity of 81.0% with specificity of 43.5%. We also noted that serum and salivary rinse compartments showed a differential pattern of methylation in normal subjects that influenced the utility of individual markers. CONCLUSIONS: Q-MSP detection of HNSCC in serum and salivary rinses using multiple targets offers improved performance when compared with single markers. Compartment-specific methylation in normal subjects affects the utility of Q-MSP detection strategies.

Shotgun metagenome data of a defined mock community using Oxford Nanopore, PacBio and Illumina technologies.

Metagenomic sequence data from defined mock communities is crucial for the assessment of sequencing platform performance and downstream analyses, including assembly, binning and taxonomic assignment. We report a comparison of shotgun metagenome sequencing and assembly metrics of a defined microbial mock community using the Oxford Nanopore Technologies (ONT) MinION, PacBio and Illumina sequencing platforms. Our synthetic microbial community BMock12 consists of 12 bacterial strains with genome sizes spanning 3.2-7.2 Mbp, 40-73% GC content, and 1.5-7.3% repeats. Size selection of both PacBio and ONT sequencing libraries prior to sequencing was essential to yield comparable relative abundances of organisms among all sequencing technologies. While the Illumina-based metagenome assembly yielded good coverage with few misassemblies, contiguity was greatly improved by both, Illumina + ONT and Illumina + PacBio hybrid assemblies but increased misassemblies, most notably in genomes with high sequence similarity to each other. Our resulting datasets allow evaluation and benchmarking of bioinformatics software on Illumina, PacBio and ONT platforms in parallel.