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101. PLoS One. 2013 Jun 6;8(6):e65520. doi: 10.1371/journal.pone.0065520. Print 2013.

 

Oral microbiome of deep and shallow dental pockets in chronic periodontitis.

 

Ge X(1), Rodriguez R, Trinh M, Gunsolley J, Xu P.

 

Author information:

(1)VCU Philips Institute of Oral and Craniofacial Molecular Biology, Virginia

Commonwealth University, Richmond, Virginia, United States of America.

 

We examined the subgingival bacterial biodiversity in untreated chronic

periodontitis patients by sequencing 16S rRNA genes. The primary purpose of the

study was to compare the oral microbiome in deep (diseased) and shallow (healthy)

sites. A secondary purpose was to evaluate the influences of smoking, race and

dental caries on this relationship. A total of 88 subjects from two clinics were

recruited. Paired subgingival plaque samples were taken from each subject, one

from a probing site depth >5 mm (deep site) and the other from a probing site

depth ≤3mm (shallow site). A universal primer set was designed to amplify the

V4-V6 region for oral microbial 16S rRNA sequences. Differences in genera and

species attributable to deep and shallow sites were determined by statistical

analysis using a two-part model and false discovery rate. Fifty-one of 170 genera

and 200 of 746 species were found significantly different in abundances between

shallow and deep sites. Besides previously identified periodontal

disease-associated bacterial species, additional species were found markedly

changed in diseased sites. Cluster analysis revealed that the microbiome

difference between deep and shallow sites was influenced by patient-level effects

such as clinic location, race and smoking. The differences between clinic

locations may be influenced by racial distribution, in that all of the African

Americans subjects were seen at the same clinic. Our results suggested that there

were influences from the microbiome for caries and periodontal disease and these

influences are independent.

 

DOI: 10.1371/journal.pone.0065520

PMCID: PMC3675156

PMID: 23762384  [PubMed - indexed for MEDLINE]

 

 

102. PLoS One. 2012;7(8):e42770. doi: 10.1371/journal.pone.0042770. Epub 2012 Aug 10.

 

The relation between oral Candida load and bacterial microbiome profiles in Dutch

older adults.

 

Kraneveld EA(1), Buijs MJ, Bonder MJ, Visser M, Keijser BJ, Crielaard W, Zaura E.

 

Author information:

(1)Department of Preventive Dentistry, Academic Centre for Dentistry Amsterdam,

University of Amsterdam, Amsterdam, The Netherlands.

 

Currently there are no evidence-based ecological measures for prevention of

overgrowth and subsequent infection by fungi in the oral cavity. The aim of this

study was to increase our knowledge on fungal-bacterial ecological interactions.

Salivary Candida abundance of 82 Dutch adults aged 58-80 years was established

relative to the bacterial load by quantitative PCR analysis of the Internal

Transcribed (ITS) region (Candida) and 16S rDNA gene (bacteria). The salivary

microbiome was assessed using barcoded pyrosequencing of the bacterial

hypervariable regions V5-V7 of 16S rDNA. Sequencing data was preprocessed by

denoising and chimera removal, clustered in Operational Taxonomic Units (OTUs)

and assigned to taxonomy. Both OTU-based (PCA, diversity statistics) and

phylogeny-based analyses (UniFrac, PCoA) were performed. Saliva of Dutch older

adults contained 0-4 × 10(8) CFU/mL Candida with a median Candida load of 0.06%.

With increased Candida load the diversity of the salivary microbiome decreased

significantly (p<0.001). Increase in the Candida load correlated positively with

class Bacilli, and negatively with class Fusobacteria, Flavobacteria, and

Bacteroidia. Microbiomes with high Candida load were less diverse and had a

distinct microbial composition towards dominance by saccharolytic and acidogenic

bacteria--streptococci. The control of the acidification of the oral environment

may be a potential preventive measure for Candida outgrowth that should be

evaluated in longitudinal clinical intervention trials.

 

DOI: 10.1371/journal.pone.0042770

PMCID: PMC3416775

PMID: 22900048  [PubMed - indexed for MEDLINE]

 

 

103. BMC Microbiol. 2014 Dec 17;14:316. doi: 10.1186/s12866-014-0316-1.

 

Comparative analysis of the human saliva microbiome from different climate zones:

Alaska, Germany, and Africa.

 

Li J(1,)(2), Quinque D(3,)(4), Horz HP(5), Li M(6), Rzhetskaya M(7), Raff

JA(8,)(9), Hayes MG(10,)(11,)(12), Stoneking M(13).

 

Author information:

(1)Max Planck Institute for Evolutionary Anthropology, Deutscher Platz 6,

D-04103, Leipzig, Germany. jingli1978@gmail.com. (2)Max Planck Independent

Research Group on Population Genomics, Chinese Academy of Sciences and Max Planck

Society (CAS-MPG) Partner Institute for Computational Biology, Shanghai

Institutes for Biological Sciences, Chinese Academy of Sciences, 320 Yueyang

Road, Shanghai, 200031, China. jingli1978@gmail.com. (3)Max Planck Institute for

Evolutionary Anthropology, Deutscher Platz 6, D-04103, Leipzig, Germany.

dquinque@googlemail.com. (4)Current address: Department of Genetics, Harvard

Medical School, 77 Louis Pasteur Avenue, Boston, MA, 02115, USA.

dquinque@googlemail.com. (5)Division of Virology, Institute of Medical

Microbiology, RWTH Aachen University Hospital, Pauwelsstrasse 30, D-52057,

Aachen, Germany. jhorz@ukaachen.de. (6)Max Planck Institute for Evolutionary

Anthropology, Deutscher Platz 6, D-04103, Leipzig, Germany. fengzys@gmail.com.

(7)Division of Endocrinology, Metabolism, and Molecular Medicine, Department of

Medicine, Northwestern University Feinberg School of Medicine, Chicago, IL,

60611, USA. m-rzhetskaya@northwestern.edu. (8)Division of Endocrinology,

Metabolism, and Molecular Medicine, Department of Medicine, Northwestern

University Feinberg School of Medicine, Chicago, IL, 60611, USA.

jenniferraff@utexas.edu. (9)Current address: Department of Anthropology,

University of Texas, Austin, TX, 78712, USA. jenniferraff@utexas.edu.

(10)Division of Endocrinology, Metabolism, and Molecular Medicine, Department of

Medicine, Northwestern University Feinberg School of Medicine, Chicago, IL,

60611, USA. ghayes@northwestern.edu. (11)Department of Anthropology, Northwestern

University, Evanston, IL, 60208, USA. ghayes@northwestern.edu. (12)Center for

Genetic Medicine, Northwestern University Feinberg School of Medicine, Chicago,

IL, 60611, USA. ghayes@northwestern.edu. (13)Max Planck Institute for

Evolutionary Anthropology, Deutscher Platz 6, D-04103, Leipzig, Germany.

stonekg@eva.mpg.de.

 

BACKGROUND: Although the importance of the human oral microbiome for health and

disease is increasingly recognized, variation in the composition of the oral

microbiome across different climates and geographic regions is largely

unexplored.

RESULTS: Here we analyze the saliva microbiome from native Alaskans (76

individuals from 4 populations), Germans (10 individuals from 1 population), and

Africans (66 individuals from 3 populations) based on next-generation sequencing

of partial 16S rRNA gene sequences. After quality filtering, a total of 67,916

analyzed sequences resulted in 5,592 OTUs (defined at ≥97% identity) and 123

genera. The three human groups differed significantly by the degree of diversity

between and within individuals (e.g. beta diversity: Africans > Alaskans >

Germans; alpha diversity: Germans > Alaskans > Africans). UniFrac, network,

ANOSIM, and correlation analyses all indicated more similarities in the saliva

microbiome of native Alaskans and Germans than between either group and Africans.

The native Alaskans and Germans also had the highest number of shared bacterial

interactions. At the level of shared OTUs, only limited support for a core

microbiome shared across all three continental regions was provided, although

partial correlation analysis did highlight interactions involving several pairs

of genera as conserved across all human groups. Subsampling strategies for

compensating for the unequal number of individuals per group or unequal sequence

reads confirmed the above observations.

CONCLUSION: Overall, this study illustrates the distinctiveness of the saliva

microbiome of human groups living under very different climatic conditions.

 

DOI: 10.1186/s12866-014-0316-1

PMCID: PMC4272767

PMID: 25515234  [PubMed - indexed for MEDLINE]

 

 

104. MBio. 2014 Apr 22;5(2):e00889. doi: 10.1128/mBio.00889-14.

 

Revealing the bacterial butyrate synthesis pathways by analyzing (meta)genomic

data.

 

Vital M, Howe AC, Tiedje JM.

 

Butyrate-producing bacteria have recently gained attention, since they are

important for a healthy colon and when altered contribute to emerging diseases,

such as ulcerative colitis and type II diabetes. This guild is polyphyletic and

cannot be accurately detected by 16S rRNA gene sequencing. Consequently,

approaches targeting the terminal genes of the main butyrate-producing pathway

have been developed. However, since additional pathways exist and alternative,

newly recognized enzymes catalyzing the terminal reaction have been described,

previous investigations are often incomplete. We undertook a broad analysis of

butyrate-producing pathways and individual genes by screening 3,184 sequenced

bacterial genomes from the Integrated Microbial Genome database. Genomes of 225

bacteria with a potential to produce butyrate were identified, including many

previously unknown candidates. The majority of candidates belong to distinct

families within the Firmicutes, but members of nine other phyla, especially from

Actinobacteria, Bacteroidetes, Fusobacteria, Proteobacteria, Spirochaetes, and

Thermotogae, were also identified as potential butyrate producers. The

established gene catalogue (3,055 entries) was used to screen for butyrate

synthesis pathways in 15 metagenomes derived from stool samples of healthy

individuals provided by the HMP (Human Microbiome Project) consortium. A high

percentage of total genomes exhibited a butyrate-producing pathway (mean, 19.1%;

range, 3.2% to 39.4%), where the acetyl-coenzyme A (CoA) pathway was the most

prevalent (mean, 79.7% of all pathways), followed by the lysine pathway (mean,

11.2%). Diversity analysis for the acetyl-CoA pathway showed that the same few

firmicute groups associated with several Lachnospiraceae and Ruminococcaceae were

dominating in most individuals, whereas the other pathways were associated

primarily with Bacteroidetes. IMPORTANCE Microbiome research has revealed new,

important roles of our gut microbiota for maintaining health, but an

understanding of effects of specific microbial functions on the host is in its

infancy, partly because in-depth functional microbial analyses are rare and

publicly available databases are often incomplete/misannotated. In this study, we

focused on production of butyrate, the main energy source for colonocytes, which

plays a critical role in health and disease. We have provided a complete database

of genes from major known butyrate-producing pathways, using in-depth genomic

analysis of publicly available genomes, filling an important gap to accurately

assess the butyrate-producing potential of complex microbial communities from

"-omics"-derived data. Furthermore, a reference data set containing the abundance

and diversity of butyrate synthesis pathways from the healthy gut microbiota was

established through a metagenomics-based assessment. This study will help in

understanding the role of butyrate producers in health and disease and may assist

the development of treatments for functional dysbiosis.

 

DOI: 10.1128/mBio.00889-14

PMCID: PMC3994512

PMID: 24757212  [PubMed - indexed for MEDLINE]

 

 

105. Am J Clin Nutr. 2015 Jan;101(1):55-64. doi: 10.3945/ajcn.114.092064. Epub 2014

Nov 12.

 

Fiber supplementation influences phylogenetic structure and functional capacity

of the human intestinal microbiome: follow-up of a randomized controlled trial.

 

Holscher HD(1), Caporaso JG(1), Hooda S(1), Brulc JM(1), Fahey GC Jr(1), Swanson

KS(1).

 

Author information:

(1)From the Department of Animal Sciences and Division of Nutritional Sciences,

University of Illinois, Urbana, IL (HDH, SH, GCF, and KSS); the Department of

Biological Sciences and Computer Sciences, Northern Arizona University,

Flagstaff, AZ (JGC); and General Mills Inc., Bell Institute of Health and

Nutrition, Minneapolis, MN (JMB).

 

Comment in

    Am J Clin Nutr. 2015 Jan;101(1):1-2.

 

BACKGROUND: In our published randomized, double-blind, placebo-controlled,

3-period crossover trial, healthy adult men (n = 21) consumed bars containing no

supplemental fiber (placebo; NFC), polydextrose (21 g/d), and soluble corn fiber

(SCF; 21 g/d) for 21 d each. Fecal specimens were collected between days 16 and

21 for fermentative end-product analysis and 16S ribosomal RNA bacterial gene

amplification for bacterial taxa identification. Fiber supplementation decreased

fecal putrefaction compounds and shifted abundances of several bacterial taxa.

OBJECTIVE: The objective was to perform whole-genome shotgun 454 pyrosequencing

on the same fecal specimens collected in that clinical trial to obtain

comprehensive fecal bacterial genome sequencing coverage and explore the full

range of bacterial genetic information in the fecal microbiome, thereby using a

systematic approach to study the impact of dietary fiber supplementation on fecal

metabolites, bacterial taxa, and bacterial metagenomes.

DESIGN: Fecal samples were subjected to whole-genome shotgun 454 pyrosequencing

to identify both fecal bacterial populations present and their functional genetic

capacity.

RESULTS: Whole-genome shotgun sequencing results revealed that fiber consumption

shifted the Bacteroidetes:Firmicutes ratio, increasing the relative abundance of

Bacteroidetes 12 ± 2% and 13 ± 2% with polydextrose and SCF, respectively,

compared with NFC. Bivariate correlations showed a positive correlation between

the Bacteroidetes:Firmicutes ratio and total dietary fiber intake but not body

mass index. Principal coordinates analysis of Bray-Curtis distances indicated

that bacterial gene composition was more similar in participants consuming fibers

(polydextrose and SCF combined) in comparison with NFC. Shifts in bacterial gene

abundances after polydextrose and SCF supplementation included genes associated

with carbohydrate, amino acid, and lipid metabolism, as well as metabolism of

cofactors and vitamins.

CONCLUSION: This study conveys novel information about the impact of dietary

fiber supplementation on the phylogenetic structure and functional capacity of

the fecal microbiome of healthy adults.

 

© 2015 American Society for Nutrition.

 

DOI: 10.3945/ajcn.114.092064

PMID: 25527750  [PubMed - indexed for MEDLINE]

 

 

106. PLoS One. 2016 May 12;11(5):e0155362. doi: 10.1371/journal.pone.0155362.

eCollection 2016.

 

Colorectal Cancer and the Human Gut Microbiome: Reproducibility with Whole-Genome

Shotgun Sequencing.

 

Vogtmann E(1,)(2), Hua X(1), Zeller G(3), Sunagawa S(3), Voigt AY(3,)(4,)(5,)(6),

Hercog R(7), Goedert JJ(1), Shi J(1), Bork P(3,)(6,)(8,)(9), Sinha R(1).

 

Author information:

(1)Division of Cancer Epidemiology & Genetics, National Cancer Institute,

Bethesda, Maryland, United States of America. (2)Division of Cancer Prevention,

National Cancer Institute, Bethesda, Maryland, United States of America.

(3)Structural and Computational Biology Unit, European Molecular Biology

Laboratory, Heidelberg, Germany. (4)Department of Applied Tumor Biology,

Institute of Pathology, University Hospital Heidelberg, Heidelberg, Germany.

(5)Clinical Cooperation Unit Applied Tumor Biology, German Cancer Research Center

(DKFZ), Heidelberg, Germany. (6)Molecular Medicine Partnership Unit (MMPU),

University Hospital Heidelberg and European Molecular Biology Laboratory,

Heidelberg, Germany. (7)Genomics Core Facility, European Molecular Biology

Laboratory, Heidelberg, Germany. (8)Max Delbrück Centre for Molecular Medicine,

Berlin, Germany. (9)Department of Bioinformatics Biocenter, University of

Würzburg, Würzburg, Germany.

 

Accumulating evidence indicates that the gut microbiota affects colorectal cancer

development, but previous studies have varied in population, technical methods,

and associations with cancer. Understanding these variations is needed for

comparisons and for potential pooling across studies. Therefore, we performed

whole-genome shotgun sequencing on fecal samples from 52 pre-treatment colorectal

cancer cases and 52 matched controls from Washington, DC. We compared findings

from a previously published 16S rRNA study to the metagenomics-derived taxonomy

within the same population. In addition, metagenome-predicted genes, modules, and

pathways in the Washington, DC cases and controls were compared to cases and

controls recruited in France whose specimens were processed using the same

platform. Associations between the presence of fecal Fusobacteria, Fusobacterium,

and Porphyromonas with colorectal cancer detected by 16S rRNA were reproduced by

metagenomics, whereas higher relative abundance of Clostridia in cancer cases

based on 16S rRNA was merely borderline based on metagenomics. This demonstrated

that within the same sample set, most, but not all taxonomic associations were

seen with both methods. Considering significant cancer associations with the

relative abundance of genes, modules, and pathways in a recently published French

metagenomics dataset, statistically significant associations in the Washington,

DC population were detected for four out of 10 genes, three out of nine modules,

and seven out of 17 pathways. In total, colorectal cancer status in the

Washington, DC study was associated with 39% of the metagenome-predicted genes,

modules, and pathways identified in the French study. More within and between

population comparisons are needed to identify sources of variation and disease

associations that can be reproduced despite these variations. Future studies

should have larger sample sizes or pool data across studies to have sufficient

power to detect associations that are reproducible and significant after

correction for multiple testing.

 

DOI: 10.1371/journal.pone.0155362

PMCID: PMC4865240

PMID: 27171425  [PubMed - in process]

 

 

107. Clin Microbiol Infect. 2012 Dec;18(12):1185-93. doi: 10.1111/1469-0691.12023.

Epub 2012 Oct 3.

 

Microbial culturomics: paradigm shift in the human gut microbiome study.

 

Lagier JC(1), Armougom F, Million M, Hugon P, Pagnier I, Robert C, Bittar F,

Fournous G, Gimenez G, Maraninchi M, Trape JF, Koonin EV, La Scola B, Raoult D.

 

Author information:

(1)Aix Marseille Université, URMITE, UM63, CNRS 7278, IRD 198, INSERM 1095,

Marseille, France.

 

Comprehensive determination of the microbial composition of the gut microbiota

and the relationships with health and disease are major challenges in the 21st

century. Metagenomic analysis of the human gut microbiota detects mostly

uncultured bacteria. We studied stools from two lean Africans and one obese

European, using 212 different culture conditions (microbial culturomics), and

tested the colonies by using mass spectrometry and 16S rRNA amplification and

sequencing. In parallel, we analysed the same three samples by pyrosequencing 16S

rRNA amplicons targeting the V6 region. The 32 500 colonies obtained by

culturomics have yielded 340 species of bacteria from seven phyla and 117 genera,

including two species from rare phyla (Deinococcus-Thermus and Synergistetes,

five fungi, and a giant virus (Senegalvirus). The microbiome identified by

culturomics included 174 species never described previously in the human gut,

including 31 new species and genera for which the genomes were sequenced,

generating c. 10 000 new unknown genes (ORFans), which will help in future

molecular studies. Among these, the new species Microvirga massiliensis has the

largest bacterial genome so far obtained from a human, and Senegalvirus is the

largest virus reported in the human gut. Concurrent metagenomic analysis of the

same samples produced 698 phylotypes, including 282 known species, 51 of which

overlapped with the microbiome identified by culturomics. Thus, culturomics

complements metagenomics by overcoming the depth bias inherent in metagenomic

approaches.

 

© 2012 The Authors. Clinical Microbiology and Infection © 2012 European Society

of Clinical Microbiology and Infectious Diseases.

 

DOI: 10.1111/1469-0691.12023

PMID: 23033984  [PubMed - indexed for MEDLINE]

 

 

108. Antimicrob Agents Chemother. 2015 Feb;59(2):1182-92. doi: 10.1128/AAC.04506-14.

Epub 2014 Dec 8.

 

Microbiome changes in healthy volunteers treated with GSK1322322, a novel

antibiotic targeting bacterial peptide deformylase.

 

Arat S(1), Spivak A(2), Van Horn S(3), Thomas E(3), Traini C(3), Sathe G(3), Livi

GP(3), Ingraham K(4), Jones L(5), Aubart K(4), Holmes DJ(4), Naderer O(5), Brown

JR(6).

 

Author information:

(1)Computational Biology, GSK R&D, Collegeville, Pennsylvania, USA Department of

Mathematics, Virginia Tech, Blacksburg, Virginia, USA. (2)Computational Biology,

GSK R&D, Collegeville, Pennsylvania, USA. (3)Target and Pathway Validation, GSK

R&D, Collegeville, Pennsylvania, USA. (4)Antibacterial Discovery Performance

Unit, GSK R&D, Collegeville, Pennsylvania, USA. (5)Antibacterial Discovery

Performance Unit, GSK R&D, Research Triangle Park, North Carolina, USA.

(6)Computational Biology, GSK R&D, Collegeville, Pennsylvania, USA

james.r.brown@gsk.com.

 

GSK1322322 is a novel antibacterial agent under development, and it has known

antibacterial activities against multidrug-resistant respiratory and skin

pathogens through its inhibition of the bacterial peptide deformylase. Here, we

used next-generation sequencing (NGS) of the bacterial 16S rRNA genes from stool

samples collected from 61 healthy volunteers at the predosing and end-of-study

time points to determine the effects of GSK1322322 on the gastrointestinal (GI)

microbiota in a phase I, randomized, double-blind, and placebo-controlled study.

GSK1322322 was administered either intravenously (i.v.) only or in an oral-i.v.

combination in single- and repeat-dose-escalation infusions. Analysis of the 16S

rRNA sequence data found no significant changes in the relative abundances of GI

operational taxonomic units (OTUs) between the prestudy and end-of-study samples

for either the placebo- or i.v.-only-treated subjects. However, oral-i.v.

treatment resulted in significant decreases in some bacterial taxa, the

Firmicutes and Bacteroidales, and increases in others, the Betaproteobacteria,

Gammaproteobacteria, and Bifidobacteriaceae. Microbiome diversity plots clearly

differentiated the end-of-study oral-i.v.-dosed samples from all others

collected. The changes in genome function as inferred from species composition

suggest an increase in bacterial transporter and xenobiotic metabolism pathways

in these samples. A phylogenetic analysis of the peptide deformylase protein

sequences collected from the published genomes of clinical isolates previously

tested for GSK1322322 in vitro susceptibility and GI bacterial reference genomes

suggests that antibiotic target homology is one of several factors that

influences the response of GI microbiota to this antibiotic. Our study shows that

dosing regimen and target class are important factors when considering the impact

of antibiotic usage on GI microbiota. (This clinical trial was registered at the

GlaxoSmithKline Clinical Study Register under study identifier PDF 113376.).

 

Copyright © 2015, American Society for Microbiology. All Rights Reserved.

 

DOI: 10.1128/AAC.04506-14

PMCID: PMC4335841

PMID: 25487798  [PubMed - indexed for MEDLINE]

 

 

109. PLoS Comput Biol. 2013;9(1):e1002863. doi: 10.1371/journal.pcbi.1002863. Epub

2013 Jan 10.

 

A guide to enterotypes across the human body: meta-analysis of microbial

community structures in human microbiome datasets.

 

Koren O(1), Knights D, Gonzalez A, Waldron L, Segata N, Knight R, Huttenhower C,

Ley RE.

 

Author information:

(1)Department of Microbiology, Cornell University, Ithaca, New York, United

States of America.

 

Recent analyses of human-associated bacterial diversity have categorized

individuals into 'enterotypes' or clusters based on the abundances of key

bacterial genera in the gut microbiota. There is a lack of consensus, however, on

the analytical basis for enterotypes and on the interpretation of these results.

We tested how the following factors influenced the detection of enterotypes:

clustering methodology, distance metrics, OTU-picking approaches, sequencing

depth, data type (whole genome shotgun (WGS) vs.16S rRNA gene sequence data), and

16S rRNA region. We included 16S rRNA gene sequences from the Human Microbiome

Project (HMP) and from 16 additional studies and WGS sequences from the HMP and

MetaHIT. In most body sites, we observed smooth abundance gradients of key genera

without discrete clustering of samples. Some body habitats displayed bimodal

(e.g., gut) or multimodal (e.g., vagina) distributions of sample abundances, but

not all clustering methods and workflows accurately highlight such clusters.

Because identifying enterotypes in datasets depends not only on the structure of

the data but is also sensitive to the methods applied to identifying clustering

strength, we recommend that multiple approaches be used and compared when testing

for enterotypes.

 

DOI: 10.1371/journal.pcbi.1002863

PMCID: PMC3542080

PMID: 23326225  [PubMed - indexed for MEDLINE]

 

 

 

PLoS One. 2016 Sep 19;11(9):e0162659. doi: 10.1371/journal.pone.0162659. eCollection 2016.

Identification of Immunity-Related Genes in Dialeurodes citri against Entomopathogenic Fungus Lecanicillium attenuatum by RNA-Seq Analysis.

Yu S1, Ding L1, Luo R1, Li X1, Yang J1, Liu H1, Cong L1, Ran C1.

Author information

Abstract

Dialeurodes citri is a major pest in citrus producing areas, and large-scale outbreaks have occurred increasingly often in recent years. Lecanicillium attenuatum is an important entomopathogenic fungus that can parasitize and kill D. citri. We separated the fungus from corpses of D. citri larvae. However, the sound immune defense system of pests makes infection by an entomopathogenic fungus difficult. Here we used RNA sequencing technology (RNA-Seq) to build a transcriptome database for D. citri and performed digital gene expression profiling to screen genes that act in the immune defense of D. citri larvae infected with a pathogenic fungus. De novo assembly generated 84,733 unigenes with mean length of 772 nt. All unigenes were searched against GO, Nr, Swiss-Prot, COG, and KEGG databases and a total of 28,190 (33.3%) unigenes were annotated. We identified 129 immunity-related unigenes in transcriptome database that were related to pattern recognition receptors, information transduction factors and response factors. From the digital gene expression profile, we identified 441 unigenes that were differentially expressed in D. citri infected with L. attenuatum. Through calculated Log2Ratio values, we identified genes for which fold changes in expression were obvious, including cuticle protein, vitellogenin, cathepsin, prophenoloxidase, clip-domain serine protease, lysozyme, and others. Subsequent quantitative real-time polymerase chain reaction analysis verified the results. The identified genes may serve as target genes for microbial control of D. citri.

PMID: 27644092 DOI: 10.1371/journal.pone.0162659

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8.

Sci Rep. 2016 Sep 19;6:33660. doi: 10.1038/srep33660.

Characterization of the mechanism of prolonged adaptation to osmotic stress of Jeotgalibacillus malaysiensis via genome and transcriptome sequencing analyses.

Yaakop AS1, Chan KG2, Ee R2, Lim YL2, Lee SK2, Manan FA1, Goh KM1.

Author information

Abstract

Jeotgalibacillus malaysiensis, a moderate halophilic bacterium isolated from a pelagic area, can endure higher concentrations of sodium chloride (NaCl) than other Jeotgalibacillus type strains. In this study, we therefore chose to sequence and assemble the entire J. malaysiensis genome. This is the first report to provide a detailed analysis of the genomic features of J. malaysiensis, and to perform genetic comparisons between this microorganism and other halophiles. J. malaysiensis encodes a native megaplasmid (pJeoMA), which is greater than 600 kilobases in size, that is absent from other sequenced species of Jeotgalibacillus. Subsequently, RNA-Seq-based transcriptome analysis was utilised to examine adaptations of J. malaysiensis to osmotic stress. Specifically, the eggNOG (evolutionary genealogy of genes: Non-supervised Orthologous Groups) and KEGG (Kyoto Encyclopaedia of Genes and Genomes) databases were used to elucidate the overall effects of osmotic stress on the organism. Generally, saline stress significantly affected carbohydrate, energy, and amino acid metabolism, as well as fatty acid biosynthesis. Our findings also indicate that J. malaysiensis adopted a combination of approaches, including the uptake or synthesis of osmoprotectants, for surviving salt stress. Among these, proline synthesis appeared to be the preferred method for withstanding prolonged osmotic stress in J. malaysiensis.

PMID: 27641516 DOI: 10.1038/srep33660

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9.

J Oral Microbiol. 2016 Sep 16;8:32383. doi: 10.3402/jom.v8.32383. eCollection 2016.

Actinomyces spp. gene expression in root caries lesions.

Dame-Teixeira N1, Parolo CC2, Maltz M2, Tugnait A3, Devine D3, Do T4.

Author information

Abstract

BACKGROUND:

The studies of the distribution of Actinomyces spp. on carious and non-carious root surfaces have not been able to confirm the association of these bacteria with root caries, although they were extensively implicated as a prime suspect in root caries.

OBJECTIVE:

The aim of this study was to observe the gene expression of Actinomyces spp. in the microbiota of root surfaces with and without caries.

DESIGN:

The oral biofilms from exposed sound root surface (SRS; n=10) and active root caries (RC; n=30) samples were collected. The total bacterial RNA was extracted, and the mRNA was isolated. Samples with low RNA concentration were pooled, yielding a final sample size of SRS=10 and RC=9. Complementary DNA (cDNA) libraries were prepared and sequenced on an Illumina(®) HiSeq 2500 system. Sequence reads were mapped to eight Actinomyces genomes. Count data were normalized using DESeq2 to analyse differential gene expression applying the Benjamini-Hochberg correction (false discovery rate [FDR]<0.001).

RESULTS:

Actinomyces spp. had similar numbers of reads (Mann-Whitney U-test; p>0.05), except for Actinomyces OT178 (p=0.001) and Actinomyces gerencseriae (p=0.004), which had higher read counts in the SRS. Genes that code for stress proteins (clp, dnaK, and groEL), enzymes of glycolysis pathways (including enolase and phosphoenolpyruvate carboxykinase), adhesion (Type-2 fimbrial and collagen-binding protein), and cell growth (EF-Tu) were highly - but not differentially (p>0.001) - expressed in both groups. Genes with the most significant upregulation in RC were those coding for hypothetical proteins and uracil DNA glycosylase (p=2.61E-17). The gene with the most significant upregulation in SRS was a peptide ABC transporter substrate-binding protein (log2FC=-6.00, FDR=2.37E-05).

CONCLUSION:

There were similar levels of Actinomyces gene expression in both sound and carious root biofilms. These bacteria can be commensal in root surface sites but may be cariogenic due to survival mechanisms that allow them to exist in acid environments and to metabolize sugars, saving energy.

KEYWORDS:

Actinomyces spp.; RNA-seq; differential expression; root caries; transcriptome

PMID: 27640531

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10.

Comp Biochem Physiol Part D Genomics Proteomics. 2016 Aug 28;20:101-110. doi: 10.1016/j.cbd.2016.08.002. [Epub ahead of print]

De novo sequencing and transcriptome analysis of female venom glands of ectoparasitoid Bracon hebetor (Say.) (Hymenoptera: Braconidae).

Manzoor A1, UlAbdin Z2, Webb BA3, Arif MJ1, Jamil A4.

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Abstract

Venom is a key-factor in the regulation of host physiology by parasitic Hymenoptera and a potentially rich source of novel bioactive substances for biotechnological applications. The limited study of venom from the ectoparasitoid Bracon hebetor, a tiny wasp that attacks larval pest insects of field and stored products and is thus a potential insect control agent, has not described the full complement and composition of these biomolecules. To have a comprehensive picture of genes expressed in the venom glands of B. hebetor, a venom gland transcriptome was assembled by using next generation sequencing technologies followed by de novo assemblies of the 10.81 M sequence reads yielded 22,425 contigs, of which 10,581 had significant BLASTx hits to know genes. The majority of hits were to Diachasma alloeum, an ectoparasitoid from same taxonomic family, as well as other wasps. Gene ontology grouped the sequences into molecular functions in which catalytic activity with 42.2% was maximum, cellular components in which cells with 33.8% and biological processes among which metabolic process with 30% had the most representatives. In this study, we highlight the most abundant sequences, and those that are likely to be functional components of the venom for parasitization. Full length ORFs of Calreticulin, Venom Acid Phosphatase Acph-1 like protein and arginine kinase proteins were isolated and their tissue specific expression was studied by RT-PCR. Our report is the first to characterize components of the B. hebetor venom glands that may be useful for developing control tools for insect pests and other applications.

Copyright © 2016 Elsevier Inc. All rights reserved.

KEYWORDS:

Bracon hebetor; De novo assembly; Illumina technology; Next generation sequencing; RNA-seq; Transcriptome; Venom glands

PMID: 27636656 DOI: 10.1016/j.cbd.2016.08.002

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11.

Genome Biol Evol. 2016 Sep 15. pii: evw229. [Epub ahead of print]

Reptile pregnancy is underpinned by complex changes in uterine gene expression: a comparative analysis of the uterine transcriptome in viviparous and oviparous lizards.

Griffith OW1, Brandley MC2, Belov K3, Thompson MB3.

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Abstract

The evolution of new organs is difficult to study because most vertebrate organs evolved only once, more than 500 million years ago. An ideal model for understanding complex organ evolution is the placenta, a structure that is present in live bearing reptiles and mammals (amniotes), which has evolved independently more than 115 times. Using transcriptomics, we characterised the uterine gene expression patterns through the reproductive cycle of a viviparous skink lizard, Pseudemoia entrecasteauxii Then we compare these patterns with the patterns of gene expression from two oviparous skinks Lampropholis guichenoti and Lerista bougainvillii While thousands of genes are differentially expressed between pregnant and non-pregnant uterine tissue in the viviparous skink, few differentially expressed genes were identified between gravid and non-gravid oviparous skinks. This finding suggests that in P. entrecasteauxii, a pregnant specific gene expression profile has evolved, allowing for the evolution of pregnancy specific innovations in the uterus. We find substantial gene expression differences between the uterus of the chorioallantoic and yolk sac placenta in P. entrecasteauxii, suggesting these placental regions are specialized for different placental functions. In particular, the chorioallantoic placenta is likely a major site of nutrient transport by membrane bound transport proteins, whilst the yolk sac placenta also likely transports nutrients but via apocrine secretions. We discuss how the evolution of transcription factor networks are likely to underpin the evolution of the new transcriptional states in the uterine tissue of viviparous reptiles.

© The Author(s) 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

KEYWORDS:

Pseudemoia; RNA-seq; convergent evolution; lizard; placenta; viviparity