scholarly journals Blood Bacterial DNA Load and Profiling Differ in Colorectal Cancer Patients Compared to Tumor-Free Controls

Cancers ◽  
2021 ◽  
Vol 13 (24) ◽  
pp. 6363
Author(s):  
Massimiliano Mutignani ◽  
Roberto Penagini ◽  
Giorgio Gargari ◽  
Simone Guglielmetti ◽  
Marcello Cintolo ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Colon Cancer ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Negative Binomial ◽  
Conditional Logistic Regression ◽  
Community Diversity ◽  
Rrna Gene ◽  
Barrier Dysfunction ◽  
Gene Copies

Inflammation and immunity are linked to intestinal adenoma (IA) and colorectal cancer (CRC) development. The gut microbiota is associated with CRC risk. Epithelial barrier dysfunction can occur, possibly leading to increased intestinal permeability in CRC patients. We conducted a case-control study including 100 incident histologically confirmed CRC cases, and 100 IA and 100 healthy subjects, matched to cases by center, sex and age. We performed 16S rRNA gene analysis of blood and applied conditional logistic regression. Further analyses were based on negative binomial distribution normalization and Random Forest algorithm. We found an overrepresentation of blood 16S rRNA gene copies in colon cancer as compared to tumor-free controls. For high levels of gene copies, community diversity was higher in colon cancer cases than controls. Bacterial taxa and operational taxonomic unit abundances were different between groups and were able to predict CRC with an accuracy of 0.70. Our data support the hypothesis of a higher passage of bacteria from gastrointestinal tract to bloodstream in colon cancer. This result can be applied on non-invasive diagnostic tests for colon cancer control.

scholarly journals Blood bacterial DNA, intestinal adenoma and colorectal cancer

2021 ◽  
Author(s):  
Massimiliano Mutignani ◽  
Roberto Penagini ◽  
Giorgio Gargari ◽  
Simone Guglielmetti ◽  
Marcello Cintolo ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Colon Cancer ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Healthy Subjects ◽  
Bacterial Load ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Gene Copies ◽  
Intestinal Adenoma

Objective We aimed to investigate the relation of blood bacterial DNA load and profiling with intestinal adenoma (IA) and colorectal cancer (CRC) patients. Design We performed 16S rRNA gene analysis of blood from 100 incident histologically confirmed CRC cases, 100 IA and 100 healthy subjects, matched to cases by centre, sex and age. Bacterial load was analysed using multiple conditional logistic regression. Differences in terms of abundance of bacteria between groups were estimated through analysis based on negative binomial distribution normalization. Random Forest was applied to predict the group assignment. Results We found an overrepresentation of blood 16S rRNA gene copies in colon cancer as compared to tumor-free controls (IA and healthy subjects). The odds ratio of colon cancer for the highest versus the lowest three quintiles of gene copies was 2.62. (95% confidence interval=1.22-5.65). No difference was found for rectal cancer and IA. For high 16S rRNA, community diversity was higher in colon cancers than controls. CRC cases had an enrichment of Peptostreptococcaceae and Acetobacteriaceae and a reduced abundance of Bacteroidaceae, Lachnospiraceae, and Ruminococcaceae. Identified variables predicted CRC from control and IA patients with an accuracy of 0.70. Conclusion Colon cancer patients had a higher DNA bacterial load and a different bacterial profiling as compared to healthy subjects, IA and rectal cancers, indicating a higher passage of bacteria from gastrointestinal tract to bloodstream. Further studies are needed to confirm this result and exploit it to conceive new non-invasive techniques for an early diagnosis of CRC.

scholarly journals Intragenomic and intraspecific heterogeneity in rrs may surpass interspecific variability in a natural population of Veillonella

Microbiology ◽  
2010 ◽  
Vol 156 (7) ◽  
pp. 2080-2091 ◽  
Author(s):  
Anne-Laure Michon ◽  
Fabien Aujoulat ◽  
Laurent Roudière ◽  
Olivier Soulier ◽  
Isabelle Zorgniotti ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Gene Diversity ◽  
Natural Populations ◽  
Variable Region ◽  
Housekeeping Gene ◽  
Single Copy ◽  
Population Based ◽  
Rrna Gene ◽  
Gene Copies

As well as intraspecific heterogeneity, intragenomic heterogeneity between 16S rRNA gene copies has been described for a range of bacteria. Due to the wide use of 16S rRNA gene sequence analysis for taxonomy, identification and metagenomics, evaluating the extent of these heterogeneities in natural populations is an essential prerequisite. We investigated inter- and intragenomic 16S rRNA gene heterogeneity of the variable region V3 in a population of 149 clinical isolates of Veillonella spp. of human origin and in 13 type or reference Veillonella strains using PCR-temporal temperature gel electrophoresis (TTGE). 16S rRNA gene diversity was high in the studied population, as 45 different banding patterns were observed. Intragenomic heterogeneity was demonstrated for 110 (74 %) isolates and 8 (61.5 %) type or reference strains displaying two or three different gene copies. Polymorphic nucleotide positions accounted for 0.5–2.5 % of the sequence and were scattered in helices H16 and H17 of the rRNA molecule. Some of them changed the secondary structure of H17. Phylotaxonomic structure of the population based on the single-copy housekeeping gene rpoB was compared with TTGE patterns. The intragenomic V3 heterogeneity, as well as recombination events between strains or isolates of different rpoB clades, impaired the 16S rRNA-based identification for some Veillonella species. Such approaches should be conducted in other bacterial populations to optimize the interpretation of 16S rRNA gene sequences in taxonomy and/or diversity studies.

scholarly journals Six species of nontuberculous mycobacteria carry non-identical 16S rRNA gene copies

2018 ◽  
Vol 155 ◽  
pp. 34-36
Author(s):  
Keita Takeda ◽  
Kinuyo Chikamatsu ◽  
Yuriko Igarashi ◽  
Yuta Morishige ◽  
Yoshiro Murase ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Nontuberculous Mycobacteria ◽  
Rrna Gene ◽  
Gene Copies

scholarly journals Fecal Short-Chain Fatty Acids Are Not Predictive of Colonic Tumor Status and Cannot Be Predicted Based on Bacterial Community Structure

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Marc A. Sze ◽  
Begüm D. Topçuoğlu ◽  
Nicholas A. Lesniak ◽  
Mack T. Ruffin ◽  
Patrick D. Schloss
Keyword(s):  
Colorectal Cancer ◽  
Fatty Acids ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Sequence Data ◽  
Short Chain Fatty Acids ◽  
Tumor Burden ◽  
Short Chain ◽  
Rrna Gene ◽  
Chain Fatty Acids

ABSTRACT Colonic bacterial populations are thought to have a role in the development of colorectal cancer with some protecting against inflammation and others exacerbating inflammation. Short-chain fatty acids (SCFAs) have been shown to have anti-inflammatory properties and are produced in large quantities by colonic bacteria that produce SCFAs by fermenting fiber. We assessed whether there was an association between fecal SCFA concentrations and the presence of colonic adenomas or carcinomas in a cohort of individuals using 16S rRNA gene and metagenomic shotgun sequence data. We measured the fecal concentrations of acetate, propionate, and butyrate within the cohort and found that there were no significant associations between SCFA concentration and tumor status. When we incorporated these concentrations into random forest classification models trained to differentiate between people with healthy colons and those with adenomas or carcinomas, we found that they did not significantly improve the ability of 16S rRNA gene or metagenomic gene sequence-based models to classify individuals. Finally, we generated random forest regression models trained to predict the concentration of each SCFA based on 16S rRNA gene or metagenomic gene sequence data from the same samples. These models performed poorly and were able to explain at most 14% of the observed variation in the SCFA concentrations. These results support the broader epidemiological data that questions the value of fiber consumption for reducing the risks of colorectal cancer. Although other bacterial metabolites may serve as biomarkers to detect adenomas or carcinomas, fecal SCFA concentrations have limited predictive power. IMPORTANCE Considering that colorectal cancer is the third leading cancer-related cause of death within the United States, it is important to detect colorectal tumors early and to prevent the formation of tumors. Short-chain fatty acids (SCFAs) are often used as a surrogate for measuring gut health and for being anticarcinogenic because of their anti-inflammatory properties. We evaluated the fecal SCFA concentrations of a cohort of individuals with different colonic tumor burdens who were previously analyzed to identify microbiome-based biomarkers of tumors. We were unable to find an association between SCFA concentration and tumor burden or use SCFAs to improve our microbiome-based models of classifying people based on their tumor status. Furthermore, we were unable to find an association between the fecal community structure and SCFA concentrations. Our results indicate that the association between fecal SCFAs, the gut microbiome, and tumor burden is weak.

Heterogeneity between 16S ribosomal RNA gene copies borne by one Desulfitobacterium strain is caused by different 100-200 bp insertions in the 5´ region

2007 ◽  
Vol 53 (1) ◽  
pp. 116-128 ◽  
Author(s):  
Richard Villemur ◽  
Philippe Constant ◽  
Annie Gauthier ◽  
Martine Shareck ◽  
Réjean Beaudet
Keyword(s):  
In Situ Hybridization ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Ribosomal Rna ◽  
16S Ribosomal Rna ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Gene Copies

Strains of Desulfitobacterium hafniense, such as strains PCP-1, DP7, TCE1, and TCP-A, have unusual long 16S ribosomal RNA (rRNA) genes due to an insertion of approximately 100 bp in the 5' region. In this report, we analyzed the 16S rRNA genes of different Desulfitobacterium strains to determine if such an insertion is a common feature of desulfitobacteria. We amplified this region by polymerase chain reaction (PCR) from eight Desulfitobacterium strains (D. hafniense strains PCP-1, DP7, TCP-A, TCE1, and DCB-2; D. dehalogenans; D. chlororespirans; and Desulfitobacterium sp. PCE1) and resolved each PCR product by denaturing gradient gel electrophoresis (DGGE). All strains had from two to seven DGGE- migrating bands, suggesting heterogeneity in their 16S rRNA gene copies. For each strain, the 5' region of the 16S rRNA genes was amplified and a clone library was derived. Clones corresponding to most PCR–DGGE migration bands were isolated. Sequencing of representative clones revealed that the heterogeneity was generated by insertions of 100–200 bp. An insertion was found in at least one copy of the 16S rRNA gene in all examined strains. In total, we found eight different types of insertions (INS1–INS8) that varied from 123 to 193 nt in length. Two-dimensional structural analyses of transcribed sequences predicted that all insertions would form an energetically stable loop. Reverse transcriptase – PCR experiments revealed that most of the observed insertions in the Desulfitobacterium strains were excised from the mature 16S rRNA transcripts. Insertions were not commonly found in bacterial 16S rRNA genes, and having a different insertion in several 16S rRNA gene copies borne by a single bacterial species was rarely observed. The function of these insertions is not known, but their occurrence can have an important impact in deriving 16S rRNA oligonucleotidic fluorescence in situ hybridization probes, as these insertions can be excised from 16S rRNA transcripts.Key words: Desulfitobacterium, 16S ribosomal RNA genes, heterogeneity, gene insertions, fluorescence in situ hybridization.

scholarly journals Bacterial DNAemia is associated with serum zonulin levels in older subjects

2020 ◽  
Author(s):  
Giorgio Gargari ◽  
Valentina Taverniti ◽  
Cristian Del Bo’ ◽  
Stefano Bernardi ◽  
Cristina Andres-Lacueva ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Serum Levels ◽  
Gene Profiling ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Blood Samples ◽  
Gene Copies ◽  
Older Subjects ◽  
Pcr Targeting

AbstractThe increased presence of bacteria in blood is a plausible contributing factor in the development and progression of aging-associated diseases. In this context, we performed the quantification and the taxonomic profiling of the bacterial DNA in blood samples collected from a group of forty-three older subjects enrolled in a nursing home. Quantitative PCR targeting the 16S rRNA gene revealed that all the older volunteers contained detectable amounts of bacterial DNA in their blood. The total amount of 16S rRNA gene copies varied considerably between subjects. Correlation analyses revealed that the bacterial DNAemia (expressed as concentration of 16S rRNA gene copies in blood) significantly correlated with the serum levels of zonulin, an emerging marker of intestinal permeability. This result was confirmed by the analysis of a second set of blood samples collected after approximately four months from the same subjects. Analyses of 16S rRNA gene profiling revealed that most of the bacterial DNA detected in blood was ascribable to the phylum Proteobacteria with a predominance of Pseudomonadaceae and Enterobacteriaceae. Several control samples were also analyzed to assess the influence exerted by contaminant bacterial DNA potentially originating from reagents and materials. The date reported here suggest that para-cellular permeability of epithelial (and potentially also endothelial) cell layers may play an important role in bacterial migration into the bloodstream. Bacterial DNAemia is likely to impact on several aspects of host physiology and could underpin the development and prognosis of various diseases in older subjects.

scholarly journals Leveraging Existing 16S rRNA Gene Surveys To Identify Reproducible Biomarkers in Individuals with Colorectal Tumors

mBio ◽  
2018 ◽  
Vol 9 (3) ◽  
Author(s):  
Marc A. Sze ◽  
Patrick D. Schloss
Keyword(s):  
Colorectal Cancer ◽  
Random Forest ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Meta Analysis ◽  
Tumor Diagnosis ◽  
Rrna Gene ◽  
Modeling Framework ◽  
Data Set ◽  
Colonic Microbiota

ABSTRACTAn increasing body of literature suggests that both individual and collections of bacteria are associated with the progression of colorectal cancer. As the number of studies investigating these associations increases and the number of subjects in each study increases, a meta-analysis to identify the associations that are the most predictive of disease progression is warranted. We analyzed previously published 16S rRNA gene sequencing data collected from feces and colon tissue. We quantified the odds ratios (ORs) for individual bacterial taxa that were associated with an individual having tumors relative to a normal colon. Among the fecal samples, there were no taxa that had significant ORs associated with adenoma and there were 8 taxa with significant ORs associated with carcinoma. Similarly, among the tissue samples, there were no taxa that had a significant OR associated with adenoma and there were 3 taxa with significant ORs associated with carcinoma. Among the significant ORs, the association between individual taxa and tumor diagnosis was equal to or below 7.11. Because individual taxa had limited association with tumor diagnosis, we trained Random Forest classification models using only the taxa that had significant ORs, using the entire collection of taxa found in each study, and using operational taxonomic units defined based on a 97% similarity threshold. All training approaches yielded similar classification success as measured using the area under the curve. The ability to correctly classify individuals with adenomas was poor, and the ability to classify individuals with carcinomas was considerably better using sequences from feces or tissue.IMPORTANCEColorectal cancer is a significant and growing health problem in which animal models and epidemiological data suggest that the colonic microbiota have a role in tumorigenesis. These observations indicate that the colonic microbiota is a reservoir of biomarkers that may improve our ability to detect colonic tumors using noninvasive approaches. This meta-analysis identifies and validates a set of 8 bacterial taxa that can be used within a Random Forest modeling framework to differentiate individuals as having normal colons or carcinomas. When models trained using one data set were tested on other data sets, the models performed well. These results lend support to the use of fecal biomarkers for the detection of tumors. Furthermore, these biomarkers are plausible candidates for further mechanistic studies into the role of the gut microbiota in tumorigenesis.

scholarly journals Six species of non-tuberculosis mycobacteria carry non-identical 16S rRNA gene copies

2018 ◽  
Author(s):  
Keita Takeda ◽  
Kinuyo Chikamatsu ◽  
Yuriko Igarashi ◽  
Yuta Morishige ◽  
Yoshiro Murase ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Rrna Gene ◽  
Target Sequence ◽  
Nucleotide Polymorphisms ◽  
16S Rrna Gene Sequences ◽  
Cloning And Sequencing ◽  
Single Nucleotide ◽  
Gene Copies ◽  
The 16S Rrna Gene

AbstractNon-tuberculosis mycobacteria (NTM) can carry two or more 16S rRNA gene copies that are, in some instances, non-identical. In this study, we used a combined cloning and sequencing approach to analyze the 16S rRNA gene sequences of six NTM species,Mycobacterium cosmeticum, M. pallens, M. hodleri, M. crocinum, M. flavescens, andM. xenopi. The approach facilitated the identification of two distinct gene copies in each species. The twoM. cosmeticumgenes had a single nucleotide difference, whereas two nucleotide polymorphisms were identified inM. hodleri, M. flavescens, andM. xenopi. M. pallenshad a difference in four nucleotides andM. crocinumin 23. Hence, we showed that the six NTM species possess at least two non-identical 16S rRNA gene copies.ImportanceThe presence of multiple 16S rRNA gene copies with nucleotide polymorphisms represents a challenge for species identification using 16S rRNA as the target sequence. Our analysis was focused on six NTM species,M. cosmeticum, M. pallens, M. hodleri, M. crocinum, M. flavescens, andM. xenopi. As a result, we generated the full-length sequences of two non-identical 16S rRNA copies for each NTM species. The data will be helpful for the sequence analysis of specimens or other samples.

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