scholarly journals Detection of cell-free microbial DNA using a contaminant-controlled analysis framework

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Enrique Zozaya-Valdés ◽  
Stephen Q. Wong ◽  
Jeanette Raleigh ◽  
Athena Hatzimihalis ◽  
Sarah Ftouni ◽  
...  
Keyword(s):  
Human Microbiome ◽  
16S Rrna Gene Sequencing ◽  
Digital Pcr ◽  
Rrna Gene ◽  
Analysis Framework ◽  
Specific Detection ◽  
Melanoma Patients ◽  
Rrna Gene Sequencing ◽  
Negative Controls ◽  
Microbial Dna

Abstract Background The human microbiome plays an important role in cancer. Accumulating evidence indicates that commensal microbiome-derived DNA may be represented in minute quantities in the cell-free DNA of human blood and could possibly be harnessed as a new cancer biomarker. However, there has been limited use of rigorous experimental controls to account for contamination, which invariably affects low-biomass microbiome studies. Results We apply a combination of 16S-rRNA-gene sequencing and droplet digital PCR to determine if the specific detection of cell-free microbial DNA (cfmDNA) is possible in metastatic melanoma patients. Compared to matched stool and saliva samples, the absolute concentration of cfmDNA is low but significantly above the levels detected from negative controls. The microbial community of plasma is strongly influenced by laboratory and reagent contaminants introduced during the DNA extraction and sequencing processes. Through the application of an in silico decontamination strategy including the filtering of amplicon sequence variants (ASVs) with batch dependent abundances and those with a higher prevalence in negative controls, we identify known gut commensal bacteria, such as Faecalibacterium, Bacteroides and Ruminococcus, and also other uncharacterised ASVs. We analyse additional plasma samples, highlighting the potential of this framework to identify differences in cfmDNA between healthy and cancer patients. Conclusions Together, these observations indicate that plasma can harbour a low yet detectable level of cfmDNA. The results highlight the importance of accounting for contamination and provide an analytical decontamination framework to allow the accurate detection of cfmDNA for future biomarker studies in cancer and other diseases.

scholarly journals Fetal gut colonization: meconium does not have a detectable microbiota before birth

2021 ◽  
Author(s):  
Katherine M. Kennedy ◽  
Max J. Gerlach ◽  
Thomas Adam ◽  
Markus M. Heimesaat ◽  
Laura Rossi ◽  
...  
Keyword(s):  
Positive Culture ◽  
16S Rrna Gene Sequencing ◽  
Microbial Colonization ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Gut Colonization ◽  
First Pass ◽  
Rrna Gene Sequencing ◽  
Negative Controls ◽  
Host Metabolism

AbstractMicrobial colonization of the human intestine impacts host metabolism and immunity, however when colonization occurs is unclear. Although numerous studies have reported bacterial DNA in first-pass meconium samples, these samples are collected hours to days after birth. We investigated whether bacteria could be detected in meconium prior to birth. Fetal meconium (n = 20) was collected by rectal swab during elective breech Cesarean sections without labour prior to antibiotics and compared to technical and procedural controls (n = 5), first-pass meconium (neonatal meconium; n = 14), and infant stool (n = 25). Unlike first-pass meconium, no microbial signal distinct from negative controls was detected in fetal meconium by 16S rRNA gene sequencing. Additionally, positive aerobic (n = 10 of 20) and anaerobic (n = 12 of 20) clinical cultures of fetal meconium (13 of 20 samples positive in at least one culture) were identified as likely skin contaminants, most frequently Staphylococcus epidermidis, and not detected by sequencing in most samples (same genera detected by culture and sequencing in 2 of 13 samples with positive culture). We conclude that fetal gut colonization does not occur before birth, and that microbial profiles of neonatal meconium reflect populations acquired during and after birth.

scholarly journals Bacterial communities associated with cell phones and shoes

PeerJ ◽  
2020 ◽  
Vol 8 ◽  
pp. e9235
Author(s):  
David A. Coil ◽  
Russell Y. Neches ◽  
Jenna M. Lang ◽  
Guillaume Jospin ◽  
Wendy E. Brown ◽  
...  
Keyword(s):  
Large Scale ◽  
Cell Phones ◽  
Human Microbiome ◽  
16S Rrna Gene Sequencing ◽  
The United States ◽  
Oral Microbiome ◽  
Rrna Gene ◽  
Sporting Events ◽  
Rrna Gene Sequencing ◽  
Source And Sink

Background Every human being carries with them a collection of microbes, a collection that is likely both unique to that person, but also dynamic as a result of significant flux with the surrounding environment. The interaction of the human microbiome (i.e., the microbes that are found directly in contact with a person in places such as the gut, mouth, and skin) and the microbiome of accessory objects (e.g., shoes, clothing, phones, jewelry) is of potential interest to both epidemiology and the developing field of microbial forensics. Therefore, the microbiome of personal accessories are of interest because they serve as both a microbial source and sink for an individual, they may provide information about the microbial exposure experienced by an individual, and they can be sampled non-invasively. Findings We report here a large-scale study of the microbiome found on cell phones and shoes. Cell phones serve as a potential source and sink for skin and oral microbiome, while shoes can act as sampling devices for microbial environmental experience. Using 16S rRNA gene sequencing, we characterized the microbiome of thousands of paired sets of cell phones and shoes from individuals at sporting events, museums, and other venues around the United States. Conclusions We place this data in the context of previous studies and demonstrate that the microbiome of phones and shoes are different. This difference is driven largely by the presence of “environmental” taxa (taxa from groups that tend to be found in places like soil) on shoes and human-associated taxa (taxa from groups that are abundant in the human microbiome) on phones. This large dataset also contains many novel taxa, highlighting the fact that much of microbial diversity remains uncharacterized, even on commonplace objects.

scholarly journals Identifying prognostic pairwise relationships among bacterial species in microbiome studies

2021 ◽  
Vol 17 (11) ◽  
pp. e1009501
Author(s):  
Sean M. Devlin ◽  
Axel Martin ◽  
Irina Ostrovnaya
Keyword(s):  
Bacterial Species ◽  
Human Microbiome ◽  
16S Rrna Gene Sequencing ◽  
R Package ◽  
Major Influence ◽  
Rrna Gene ◽  
Data Set ◽  
Cancer Data ◽  
Selection Framework ◽  
Rrna Gene Sequencing

In recent literature, the human microbiome has been shown to have a major influence on human health. To investigate this impact, scientists study the composition and abundance of bacterial species, commonly using 16S rRNA gene sequencing, among patients with and without a disease or condition. Methods for such investigations to date have focused on the association between individual bacterium and an outcome, and higher-order pairwise relationships or interactions among bacteria are often avoided due to the substantial increase in dimension and the potential for spurious correlations. However, overlooking such relationships ignores the environment of the microbiome, where there is dynamic cooperation and competition among bacteria. We present a method for identifying and ranking pairs of bacteria that have a differential dichotomized relationship across outcomes. Our approach, implemented in an R package PairSeek, uses the stability selection framework with data-driven dichotomized forms of the pairwise relationships. We illustrate the properties of the proposed method using a published oral cancer data set and a simulation study.

scholarly journals Bacterial communities associated with cell phones and shoes

2019 ◽  
Author(s):  
David A Coil ◽  
Russell Y Neches ◽  
Jenna M Lang ◽  
Guillaume Jospin ◽  
Wendy E Brown ◽  
...  
Keyword(s):  
Large Scale ◽  
Cell Phones ◽  
Human Microbiome ◽  
16S Rrna Gene Sequencing ◽  
The United States ◽  
Rrna Gene ◽  
Oral Microbiota ◽  
Sporting Events ◽  
Rrna Gene Sequencing ◽  
Source And Sink

Background: Every human being carries with them a collection of microbes, a collection that is likely both unique to that person, but also dynamic as a result of significant flux with the surrounding environment. The interaction of the human microbiome (i.e., the microbes that are found directly in contact with a person in places such as the gut, mouth, and skin) and the microbiome of accessory objects (e.g., shoes, clothing, phones, jewelry) is of potential interest to both epidemiology and the developing field of microbial forensics. Therefore, the microbiome of personal accessories are of interest because they serve as both a microbial source and sink for an individual, they may provide information about the microbial exposure experienced by an individual, and they can be sampled non-invasively. Findings: We report here a large-scale study of the microbiota found on cell phones and shoes. Cell phones serve as a potential source and sink for skin and oral microbiota, while shoes can act as sampling devices for the microbial environmental experience. Using 16S rRNA gene sequencing, we characterized the microbiota of thousands of paired sets of cell phones and shoes from individuals at sporting events, museums, and other venues around the United States. Conclusions: We place this data in the context of previous studies and demonstrate that the microbiota of phones and shoes are different. This difference is driven largely by the presence of “environmental” taxa (taxa from groups that tend to be found in places like soil) on shoes and human-associated taxa (taxa from groups that are abundant in the human microbiome) on phones. This large dataset also contains many novel taxa, highlighting the fact that much of microbial diversity remains uncharacterized, even on commonplace objects.

scholarly journals Bacterial communities associated with cell phones and shoes

2019 ◽  
Author(s):  
David A Coil ◽  
Russell Y Neches ◽  
Jenna M Lang ◽  
Guillaume Jospin ◽  
Wendy E Brown ◽  
...  
Keyword(s):  
Large Scale ◽  
Cell Phones ◽  
Human Microbiome ◽  
16S Rrna Gene Sequencing ◽  
The United States ◽  
Rrna Gene ◽  
Oral Microbiota ◽  
Sporting Events ◽  
Rrna Gene Sequencing ◽  
Source And Sink

Background: Every human being carries with them a collection of microbes, a collection that is likely both unique to that person, but also dynamic as a result of significant flux with the surrounding environment. The interaction of the human microbiome (i.e., the microbes that are found directly in contact with a person in places such as the gut, mouth, and skin) and the microbiome of accessory objects (e.g., shoes, clothing, phones, jewelry) is of potential interest to both epidemiology and the developing field of microbial forensics. Therefore, the microbiome of personal accessories are of interest because they serve as both a microbial source and sink for an individual, they may provide information about the microbial exposure experienced by an individual, and they can be sampled non-invasively. Findings: We report here a large-scale study of the microbiota found on cell phones and shoes. Cell phones serve as a potential source and sink for skin and oral microbiota, while shoes can act as sampling devices for the microbial environmental experience. Using 16S rRNA gene sequencing, we characterized the microbiota of thousands of paired sets of cell phones and shoes from individuals at sporting events, museums, and other venues around the United States. Conclusions: We place this data in the context of previous studies and demonstrate that the microbiota of phones and shoes are different. This difference is driven largely by the presence of “environmental” taxa (taxa from groups that tend to be found in places like soil) on shoes and human-associated taxa (taxa from groups that are abundant in the human microbiome) on phones. This large dataset also contains many novel taxa, highlighting the fact that much of microbial diversity remains uncharacterized, even on commonplace objects.

scholarly journals Investigation of Nasal/Oropharyngeal Microbial Community of COVID-19 Patients by 16S rDNA Sequencing

2021 ◽  
Vol 18 (4) ◽  
pp. 2174
Author(s):  
Martina Rueca ◽  
Andrea Fontana ◽  
Barbara Bartolini ◽  
Pierluca Piselli ◽  
Antonio Mazzarelli ◽  
...  
Keyword(s):  
Medical Condition ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Shannon Index ◽  
Rrna Gene ◽  
Human Coronavirus ◽  
Rdna Sequencing ◽  
Rrna Gene Sequencing ◽  
Severity Of The Disease ◽  
Negative Controls

Since December 2019, SARS-CoV-2 infection has been still rapidly spreading, resulting in a pandemic, followed by an increasing number of cases in countries throughout the world. The severity of the disease depends on the patient’s overall medical condition but no appropriate markers are available to establish the prognosis of the patients. We performed a 16S rRNA gene sequencing, revealing an altered composition of the nasal/oropharyngeal (NOP) microbiota in 21 patients affected by COVID-19, paucisymptomatic or in an Intensive Care Unit (ICU), as compared to 10 controls negative for COVID-19 or eight affected by a different Human Coronavirus (HKU, NL63 and OC43). A significant decrease in Chao1 index was observed when patients affected by COVID-19 (in ICU) were compared to paucisymptomatic. Furthermore, patients who were in ICU, paucisymptomatic or affected by other Coronaviruses all displayed a decrease in the Chao1 index when compared to controls, while Shannon index significantly decreased only in patients under ICU as compared to controls and paucisymptomatic patients. At the phylum level, Deinococcus-Thermus was present only in controls as compared to SARS-CoV-2 patients admitted to ICU, paucisymptomatic or affected by other coronaviruses. Candidatus Saccharibacteria (formerly known as TM7) was strongly increased in negative controls and SARS-CoV-2 paucisymptomatic patients as compared to SARS-CoV-2 ICU patients. Other modifications were observed at a lower taxonomy level. Complete depletion of Bifidobacterium and Clostridium was exclusively observed in ICU SARS-CoV-2 patients, which was the only group characterized by the presence of Salmonella, Scardovia, Serratia and Pectobacteriaceae. In conclusion, our preliminary results showed that nasal/oropharyngeal microbiota profiles of patients affected with SARS-CoV-2 may provide valuable information in order to facilitate the stratification of patients and may open the way to new interventional strategies in order to ameliorate the outcome of the patients.

Screening and Molecular Characterization of Cellulase Producing Actinobacteria from Litchi Orchard

2019 ◽  
Vol 13 (1) ◽  
pp. 90-101
Author(s):  
Sanju Kumari ◽  
Utkarshini Sharma ◽  
Rohit Krishna ◽  
Kanak Sinha ◽  
Santosh Kumar
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Molecular Characterization ◽  
Biochemical Characterization ◽  
Evolutionary Relationship ◽  
Gene Sequencing ◽  
Cellulase Activity ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Rrna Gene Sequencing

Background: Cellulolysis is of considerable economic importance in laundry detergents, textile and pulp and paper industries and in fermentation of biomass into biofuels. Objective: The aim was to screen cellulase producing actinobacteria from the fruit orchard because of its requirement in several chemical reactions. Methods: Strains of actinobacteria were isolated on Sabouraud’s agar medium. Similarities in cultural and biochemical characterization by growing the strains on ISP medium and dissimilarities among them perpetuated to recognise nine groups of actinobacteria. Cellulase activity was measured by the diameter of clear zone around colonies on CMC agar and the amount of reducing sugar liberated from carboxymethyl cellulose in the supernatant of the CMC broth. Further, 16S rRNA gene sequencing and molecular characterization were placed before NCBI for obtaining recognition with accession numbers. Results: Prominent clear zones on spraying Congo Red were found around the cultures of strains of three groups SK703, SK706, SK708 on CMC agar plates. The enzyme assay for carboxymethylcellulase displayed extra cellulase activity in broth: 0.14, 0.82 and 0.66 µmol mL-1 min-1, respectively at optimum conditions of 35°C, pH 7.3 and 96 h of incubation. However, the specific cellulase activities per 1 mg of protein did not differ that way. It was 1.55, 1.71 and 1.83 μmol mL-1 min-1. The growing mycelia possessed short compact chains of 10-20 conidia on aerial branches. These morphological and biochemical characteristics, followed by their verification by Bergey’s Manual, categorically allowed the strains to be placed under actinobacteria. Further, 16S rRNA gene sequencing, molecular characterization and their evolutionary relationship through phylogenetics also confirmed the putative cellulase producing isolates of SK706 and SK708 subgroups to be the strains of Streptomyces. These strains on getting NCBI recognition were christened as Streptomyces glaucescens strain SK91L (KF527284) and Streptomyces rochei strain SK78L (KF515951), respectively. Conclusion: Conclusive evidence on the basis of different parameters established the presence of cellulase producing actinobacteria in the litchi orchard which can convert cellulose into fermentable sugar.

scholarly journals Diversity, Composition and Functional Inference of Gut Microbiota in Indian Cabbage white Pieris canidia (Lepidoptera: Pieridae)

Life ◽  
2020 ◽  
Vol 10 (11) ◽  
pp. 254
Author(s):  
Ying Wang ◽  
Jianqing Zhu ◽  
Jie Fang ◽  
Li Shen ◽  
Shuojia Ma ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
16S Rrna Gene Sequencing ◽  
Adult Survival ◽  
Rrna Gene ◽  
Life Stages ◽  
Microbial Composition ◽  
Significant Difference ◽  
Functional Inference ◽  
Rrna Gene Sequencing ◽  
Insect Fitness

We characterized the gut microbial composition and relative abundance of gut bacteria in the larvae and adults of Pieris canidia by 16S rRNA gene sequencing. The gut microbiota structure was similar across the life stages and sexes. The comparative functional analysis on P. canidia bacterial communities with PICRUSt showed the enrichment of several pathways including those for energy metabolism, immune system, digestive system, xenobiotics biodegradation, transport, cell growth and death. The parameters often used as a proxy of insect fitness (development time, pupation rate, emergence rate, adult survival rate and weight of 5th instars larvae) showed a significant difference between treatment group and untreated group and point to potential fitness advantages with the gut microbiomes in P. canidia. These data provide an overall view of the bacterial community across the life stages and sexes in P. canidia.

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