scholarly journals Cervicovaginal Microbiome Composition Is Associated with Metabolic Profiles in Healthy Pregnancy

mBio ◽  
2020 ◽  
Vol 11 (4) ◽  
Author(s):  
Andrew Oliver ◽  
Brandon LaMere ◽  
Claudia Weihe ◽  
Stephen Wandro ◽  
Karen L. Lindsay ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Reproductive Health ◽  
Microbial Communities ◽  
Carbon Sources ◽  
Amplicon Sequencing ◽  
Vaginal Microbiome ◽  
Content Type ◽  
Healthy Women ◽  
Healthy Pregnancy ◽  
Tof Ms

ABSTRACT Microbes and their metabolic products influence early-life immune and microbiome development, yet remain understudied during pregnancy. Vaginal microbial communities are typically dominated by one or a few well-adapted microbes which are able to survive in a narrow pH range and are adapted to live on host-derived carbon sources, likely sourced from glycogen and mucin present in the vaginal environment. We characterized the cervicovaginal microbiomes of 16 healthy women throughout the three trimesters of pregnancy. Additionally, we analyzed saliva and urine metabolomes using gas chromatography-time of flight mass spectrometry (GC-TOF MS) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) lipidomics approaches for samples from mothers and their infants through the first year of life. Amplicon sequencing revealed most women had either a simple community with one highly abundant species of Lactobacillus or a more diverse community characterized by a high abundance of Gardnerella, as has also been previously described in several independent cohorts. Integrating GC-TOF MS and lipidomics data with amplicon sequencing, we found metabolites that distinctly associate with particular communities. For example, cervicovaginal microbial communities dominated by Lactobacillus crispatus have high mannitol levels, which is unexpected given the characterization of L. crispatus as a homofermentative Lactobacillus species. It may be that fluctuations in which Lactobacillus dominate a particular vaginal microbiome are dictated by the availability of host sugars, such as fructose, which is the most likely substrate being converted to mannitol. Overall, using a multi-“omic” approach, we begin to address the genetic and molecular means by which a particular vaginal microbiome becomes vulnerable to large changes in composition. IMPORTANCE Humans have a unique vaginal microbiome compared to other mammals, characterized by low diversity and often dominated by Lactobacillus spp. Dramatic shifts in vaginal microbial communities sometimes contribute to the presence of a polymicrobial overgrowth condition called bacterial vaginosis (BV). However, many healthy women lacking BV symptoms have vaginal microbiomes dominated by microbes associated with BV, resulting in debate about the definition of a healthy vaginal microbiome. Despite substantial evidence that the reproductive health of a woman depends on the vaginal microbiota, future therapies that may improve reproductive health outcomes are stalled due to limited understanding surrounding the ecology of the vaginal microbiome. Here, we use sequencing and metabolomic techniques to show novel associations between vaginal microbes and metabolites during healthy pregnancy. We speculate these associations underlie microbiome dynamics and may contribute to a better understanding of transitions between alternative vaginal microbiome compositions.

scholarly journals Cervicovaginal microbiome composition drives metabolic profiles in healthy pregnancy

2019 ◽  
Author(s):  
Andrew Oliver ◽  
Brandon LaMere ◽  
Claudia Weihe ◽  
Stephen Wandro ◽  
Karen L. Lindsay ◽  
...  
Keyword(s):  
Microbial Communities ◽  
Carbon Sources ◽  
Amplicon Sequencing ◽  
Abundant Species ◽  
Metagenomic Sequencing ◽  
Vaginal Microbiome ◽  
Microbiome Composition ◽  
Aryl Hydrocarbon ◽  
Shotgun Metagenomic Sequencing ◽  
Ph Range

AbstractBackgroundMicrobes and their metabolic products influence early-life immune and microbiome development, yet remain understudied during pregnancy. Vaginal microbial communities are typically dominated by one or a few well adapted microbes, which are able to survive in a narrow pH range. In comparison to other human-associated microbes, vaginal microbes are adapted to live on host-derived carbon sources, likely sourced from glycogen and mucin present in the vaginal environment.MethodsUsing 16S rRNA and ITS amplicon sequencing, we characterized the cervicovaginal microbiomes of 18 healthy women throughout the three trimesters of pregnancy. Shotgun metagenomic sequencing permitted refinement of the taxonomy established by amplicon sequencing, and identification of functional genes. Additionally, we analyzed saliva and urine metabolomes using GC-TOF and LC-MS/MS lipidomics approaches for samples from mothers and their infants through the first year of life.ResultsAmplicon sequencing revealed most women had either a simple community with one highly abundant species of Lactobacillus or a more diverse community characterized by a high abundance of Gardnerella, as has also been previously described in several independent cohorts. Integrating GC-TOF and lipidomics data with amplicon sequencing, we found metabolites that distinctly associate with particular communities. For example, cervicovaginal microbial communities dominated by Lactobacillus crispatus also have high mannitol levels, which contradicts the basic characterization of L. crispatus as a homofermentative Lactobacillus species. It may be that fluctuations in which Lactobacillus dominate a particular vaginal microbiome are dictated by the availability of host sugars, such as fructose, which is the most likely substrate being converted to mannitol. Furthermore, indole-3-lactate (ILA) was also indicative of L. crispatus specifically. ILA has immunomodulatory properties through binding the human aryl hydrocarbon receptor (AhR), which may maintain the especially low diversity of L. crispatus dominated communities.ConclusionsOverall, using a multi-‘omic approach, we begin to address the genetic and molecular means by which a particular vaginal microbiome becomes vulnerable to large changes in composition.

scholarly journals Effects of Actinomycete Secondary Metabolites on Sediment Microbial Communities

2016 ◽  
Vol 83 (4) ◽  
Author(s):  
Nastassia V. Patin ◽  
Michelle Schorn ◽  
Kristen Aguinaldo ◽  
Tommie Lincecum ◽  
Bradley S. Moore ◽  
...  
Keyword(s):  
Microbial Community ◽  
Secondary Metabolites ◽  
Microbial Communities ◽  
Community Composition ◽  
Marine Sediments ◽  
Microbial Community Composition ◽  
Amplicon Sequencing ◽  
Next Generation ◽  
Content Type ◽  
Predatory Bacteria

ABSTRACT Marine sediments harbor complex microbial communities that remain poorly studied relative to other biomes such as seawater. Moreover, bacteria in these communities produce antibiotics and other bioactive secondary metabolites, yet little is known about how these compounds affect microbial community structure. In this study, we used next-generation amplicon sequencing to assess native microbial community composition in shallow tropical marine sediments. The results revealed complex communities comprised of largely uncultured taxa, with considerable spatial heterogeneity and known antibiotic producers comprising only a small fraction of the total diversity. Organic extracts from cultured strains of the sediment-dwelling actinomycete genus Salinispora were then used in mesocosm studies to address how secondary metabolites shape sediment community composition. We identified predatory bacteria and other taxa that were consistently reduced in the extract-treated mesocosms, suggesting that they may be the targets of allelopathic interactions. We tested related taxa for extract sensitivity and found general agreement with the culture-independent results. Conversely, several taxa were enriched in the extract-treated mesocosms, suggesting that some bacteria benefited from the interactions. The results provide evidence that bacterial secondary metabolites can have complex and significant effects on sediment microbial communities. IMPORTANCE Ocean sediments represent one of Earth's largest and most poorly studied biomes. These habitats are characterized by complex microbial communities where competition for space and nutrients can be intense. This study addressed the hypothesis that secondary metabolites produced by the sediment-inhabiting actinomycete Salinispora arenicola affect community composition and thus mediate interactions among competing microbes. Next-generation amplicon sequencing of mesocosm experiments revealed complex communities that shifted following exposure to S. arenicola extracts. The results reveal that certain predatory bacteria were consistently less abundant following exposure to extracts, suggesting that microbial metabolites mediate competitive interactions. Other taxa increased in relative abundance, suggesting a benefit from the extracts themselves or the resulting changes in the community. This study takes a first step toward assessing the impacts of bacterial metabolites on sediment microbial communities. The results provide insight into how low-abundance organisms may help structure microbial communities in ocean sediments.

scholarly journals Molecular and Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry-Based Characterization of Clinically Significant Melanized Fungi in India

2017 ◽  
Vol 55 (4) ◽  
pp. 1090-1103 ◽  
Author(s):  
Ashutosh Singh ◽  
Pradeep Kumar Singh ◽  
Anil Kumar ◽  
Jagdish Chander ◽  
Geetika Khanna ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Time Of Flight ◽  
Laser Desorption ◽  
Content Type ◽  
Maldi Tof Ms ◽  
Flight Mass Spectrometry ◽  
Melanized Fungi ◽  
Tof Ms ◽  
Matrix Assisted Laser ◽  
Matrix Assisted Laser Desorption

ABSTRACTMelanized or black fungi are a heterogeneous group of fungi causing cutaneous to systemic diseases with high mortality. These fungi are rarely reported as agents of human infections, primarily due to difficulties in their classical identification. In this study, we examined, using molecular methods and matrix-assisted laser desorption ioniazation–time of flight mass spectrometry (MALDI-TOF MS), the diversity of melanized fungi (MF) isolated from patients in 19 medical centers in India. Overall, during a 4-year period, 718 (5.3%) clinical specimens yielded MF. Of these, 72 (10%) isolates had clinical significance and were identified primarily by sequencing the internal transcribed spacer and large subunit (LSU) regions. MF represented 21 genera comprising 29 species, the majority of them belonging to the orders Pleosporales (50%) and Chaetothyriales (22%). Among the 29 fungal species identified in this study, only 6 (20%) species were identified by the MALDI-TOF MS due to the limited commercial database of Bruker Daltonics for MF. However, a 100% identification rate of 20 additional species identified in this study was obtained by constructing an in-house database using 24- to 96-h-old liquid cultures. Further, the CLSI broth microdilution method revealed low MICs for posaconazole (≤1 μg/ml) and voriconazole (≤2 μg/ml) in 96% and 95% of isolates, respectively. Skin/subcutaneous and sino-nasal and pulmonary phaeohyphomycosis due to MF were diagnosed in 21.4% (n= 15) and 28.5% (n= 20) of cases. Also, 10% of patients had central nervous system involvement (n= 7), and 3 cases of fungal osteomyelitis due toCladophialophora bantianaandCorynesporaspp. were observed.

scholarly journals Acinetobacter variabilis sp. nov. (formerly DNA group 15 sensu Tjernberg & Ursing), isolated from humans and animals

2015 ◽  
Vol 65 (Pt_3) ◽  
pp. 857-863 ◽  
Author(s):  
Lenka Krizova ◽  
Jana McGinnis ◽  
Martina Maixnerova ◽  
Matej Nemec ◽  
Laurent Poirel ◽  
...  
Keyword(s):  
Carbon Sources ◽  
Taxonomic Status ◽  
Comparative Sequence Analysis ◽  
Whole Genome ◽  
Content Type ◽  
Maldi Tof Ms ◽  
Link Type ◽  
Maldi Tof ◽  
Group 15 ◽  
Tof Ms

We aimed to define the taxonomic status of 16 strains which were phenetically congruent with Acinetobacter DNA group 15 described by Tjernberg & Ursing in 1989. The strains were isolated from a variety of human and animal specimens in geographically distant places over the last three decades. Taxonomic analysis was based on an Acinetobacter -targeted, genus-wide approach that included the comparative sequence analysis of housekeeping, protein-coding genes, whole-cell profiling based on matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS), an array of in-house physiological and metabolic tests, and whole-genome comparative analysis. Based on analyses of the rpoB and gyrB genes, the 16 strains formed respective, strongly supported clusters clearly separated from the other species of the genus Acinetobacter . The distinctness of the group at the species level was indicated by average nucleotide identity values of ≤82 % between the whole genome sequences of two of the 16 strains (NIPH 2171T and NIPH 899) and those of the known species. In addition, the coherence of the group was also supported by MALDI-TOF MS. All 16 strains were non-haemolytic and non-gelatinase-producing, grown at 41 °C and utilized a rather limited number of carbon sources. Virtually every strain displayed a unique combination of metabolic and physiological features. We conclude that the 16 strains represent a distinct species of the genus Acinetobacter , for which the name Acinetobacter variabilis sp. nov. is proposed to reflect its marked phenotypic heterogeneity. The type strain is NIPH 2171T ( = CIP 110486T = CCUG 26390T = CCM 8555T).

scholarly journals Detection of Colistin Resistance in Escherichia coli by Use of the MALDI Biotyper Sirius Mass Spectrometry System

2019 ◽  
Vol 57 (12) ◽  
Author(s):  
R. Christopher D. Furniss ◽  
Laurent Dortet ◽  
William Bolland ◽  
Oliver Drews ◽  
Katrin Sparbier ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Escherichia Coli ◽  
Cost Effective ◽  
Colistin Resistance ◽  
Content Type ◽  
Maldi Tof Ms ◽  
E Coli ◽  
Maldi Tof ◽  
Maldi Biotyper ◽  
Tof Ms

ABSTRACT Polymyxin antibiotics are a last-line treatment for multidrug-resistant Gram-negative bacteria. However, the emergence of colistin resistance, including the spread of mobile mcr genes, necessitates the development of improved diagnostics for the detection of colistin-resistant organisms in hospital settings. The recently developed MALDIxin test enables detection of colistin resistance by matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) in less than 15 min but is not optimized for the mass spectrometers commonly found in clinical microbiology laboratories. In this study, we adapted the MALDIxin test for the MALDI Biotyper Sirius MALDI-TOF MS system (Bruker Daltonics). We optimized the sample preparation protocol by using a set of 6 mobile colistin resistance (MCR) protein-expressing Escherichia coli clones and validated the assay with a collection of 40 E. coli clinical isolates, including 19 confirmed MCR protein producers, 12 colistin-resistant isolates that tested negative for commonly encountered mcr genes (i.e., likely chromosomally resistant isolates), and 9 polymyxin-susceptible isolates. We calculated polymyxin resistance ratio (PRR) values from the acquired spectra; PRR values of 0, indicating polymyxin susceptibility, were obtained for all colistin-susceptible E. coli isolates, whereas positive PRR values, indicating resistance to polymyxins, were obtained for all resistant strains, independent of the genetic basis of resistance. Thus, we report a preliminary feasibility study showing that an optimized version of the MALDIxin test adapted for the routine MALDI Biotyper Sirius system provides an unbiased, fast, reliable, cost-effective, and high-throughput way of detecting colistin resistance in clinical E. coli isolates.

scholarly journals Rare Biosphere Archaea Assimilate Acetate in Precambrian Terrestrial Subsurface at 2.2 km Depth

Geosciences ◽  
2018 ◽  
Vol 8 (11) ◽  
pp. 418 ◽  
Author(s):  
Maija Nuppunen-Puputti ◽  
Lotta Purkamo ◽  
Riikka Kietäväinen ◽  
Mari Nyyssönen ◽  
Merja Itävaara ◽  
...  
Keyword(s):  
Microbial Communities ◽  
Cell Activation ◽  
Carbon Sources ◽  
Amplicon Sequencing ◽  
Stable Isotope Probing ◽  
Deep Biosphere ◽  
Deep Subsurface ◽  
Rare Biosphere ◽  
Terrestrial Subsurface

The deep biosphere contains a large portion of the total microbial communities on Earth, but little is known about the carbon sources that support deep life. In this study, we used Stable Isotope Probing (SIP) and high throughput amplicon sequencing to identify the acetate assimilating microbial communities at 2260 m depth in the bedrock of Outokumpu, Finland. The long-term and short-term effects of acetate on the microbial communities were assessed by DNA-targeted SIP and RNA targeted cell activation. The microbial communities reacted within hours to the amended acetate. Archaeal taxa representing the rare biosphere at 2260 m depth were identified and linked to the cycling of acetate, and were shown to have an impact on the functions and activity of the microbial communities in general through small key carbon compounds. The major archaeal lineages identified to assimilate acetate and metabolites derived from the labelled acetate were Methanosarcina spp., Methanococcus spp., Methanolobus spp., and unclassified Methanosarcinaceae. These archaea have previously been detected in the Outokumpu deep subsurface as minor groups. Nevertheless, their involvement in the assimilation of acetate and secretion of metabolites derived from acetate indicated an important role in the supporting of the whole community in the deep subsurface, where carbon sources are limited.

scholarly journals Comparison of PCR-Electrospray Ionization Mass Spectrometry with 16S rRNA PCR and Amplicon Sequencing for Detection of Bacteria in Excised Heart Valves

2016 ◽  
Vol 54 (11) ◽  
pp. 2825-2831 ◽  
Author(s):  
Bart Peeters ◽  
Paul Herijgers ◽  
Kurt Beuselinck ◽  
Willy E. Peetermans ◽  
Marie-Christin Herregods ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Infective Endocarditis ◽  
16S Rrna ◽  
Heart Valves ◽  
Amplicon Sequencing ◽  
Molecular Techniques ◽  
Ionization Mass ◽  
Causative Pathogen ◽  
Content Type ◽  
Esi Ms

Identification of the causative pathogen of infective endocarditis (IE) is crucial for adequate management and therapy. A broad-range PCR-electrospray ionization mass spectrometry (PCR-ESI-MS) technique was compared with broad-spectrum 16S rRNA PCR and amplicon sequencing (16S rRNA PCR) for the detection of bacterial pathogens in 40 heart valves obtained from 34 definite infective endocarditis patients according to the modified Duke criteria and six nonendocarditis patients. Concordance between the two molecular techniques was 98% for being positive or negative, 97% for concordant identification up to the genus level, and 77% for concordant identification up to the species level. Sensitivity for detecting the causative pathogen (up to the genus level) in excised heart valves was 88% for 16S rRNA PCR and 85% for PCR-ESI-MS; the specificity was 83% for both methods. The two molecular techniques were significantly more sensitive than valve culture (18%) and accurately identified bacteria in excised heart valves. In eight patients with culture-negative IE, the following results were obtained: concordant detection ofCoxiella burnetii(n= 2),Streptococcus gallolyticus(n= 1),Propionibacterium acnes(n= 1), and viridans group streptococci (n= 1) by both molecular tests, detection ofP. acnesby PCR-ESI-MS whereas the 16S rRNA PCR was negative (n= 1), and a false-negative result by both molecular techniques (n= 2). In one case of IE caused by viridans streptococci, PCR-ESI-MS was positive forEnterococcusspp. The advantages of PCR-ESI-MS compared to 16S rRNA PCR are its automated workflow and shorter turnaround times.

scholarly journals Comparison of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry and Molecular Biology Techniques for Identification of Culicoides (Diptera: Ceratopogonidae) Biting Midges in Senegal

2014 ◽  
Vol 53 (2) ◽  
pp. 410-418 ◽  
Author(s):  
Masse Sambou ◽  
Maxence Aubadie-Ladrix ◽  
Florence Fenollar ◽  
Becaye Fall ◽  
Hubert Bassene ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Species Level ◽  
Morphological Identification ◽  
Biting Midges ◽  
Ionization Time ◽  
Content Type ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Flight Mass Spectrometry ◽  
Tof Ms

Biting midges of the genusCulicoidesare implicated as vectors for a wide variety of pathogens. The morphological identification of these arthropods may be difficult because of a lack of detailed investigation of taxonomy for this species in Africa. However, matrix-assisted laser desorption ionization−time of flight mass spectrometry (MALDI-TOF MS) profiling is efficient for arthropod identification at the species level. This study established a spectrum database ofCulicoidesspp. from Senegal using MALDI-TOF. Identification ofCulicoidesinsects to the species level before mass spectrometry was performed on the basis of morphological characters. MALDI-TOF MS reference spectra were determined for 437 field-caughtCulicoidesof 10 species. The protein profiles of all testedCulicoidesrevealed several peaks with mass ranges of 2 to 20 kDa. In a validation study, 72Culicoidesspecimens in the target species were correctly identified at the species level with a similarity of 95 to 99.9%. FourCulicoidesprotein profiles were misidentified. Nevertheless, six SuperSpectra (C. imicola,C. enderleini,C. oxystoma,C. kingi,C. magnus, andC. fulvithorax) were created. Abdomens of midges were used to amplify and sequence a portion of the mitochondrial cytochrome oxidase I gene (COI). The results obtained using the MALDI-TOF MS method were consistent with the morphological identification and similar to the genetic identification. Protein profiling using MALDI-TOF is an efficient approach for the identification ofCulicoidesspp., and it is economically advantageous for approaches that require detailed and quantitative information of vector species that are collected in field. The database of AfricanCulicoidesMS spectra created is the first database in Africa. The COI sequences of fiveCulicoidesspecies that were previously noncharacterized using molecular methods were deposited in GenBank.

scholarly journals Development of a time-series shotgun metagenomics database for monitoring microbial communities at the Pacific coast of Japan

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Kazutoshi Yoshitake ◽  
Gaku Kimura ◽  
Tomoko Sakami ◽  
Tsuyoshi Watanabe ◽  
Yukiko Taniuchi ◽  
...  
Keyword(s):  
Time Series ◽  
Microbial Communities ◽  
Pacific Coast ◽  
Three Dimensional ◽  
Amplicon Sequencing ◽  
Metagenomic Data ◽  
Shotgun Metagenomics ◽  
Functional Features ◽  
Pacific Coast Of Japan ◽  
Marine Microbial Communities

AbstractAlthough numerous metagenome, amplicon sequencing-based studies have been conducted to date to characterize marine microbial communities, relatively few have employed full metagenome shotgun sequencing to obtain a broader picture of the functional features of these marine microbial communities. Moreover, most of these studies only performed sporadic sampling, which is insufficient to understand an ecosystem comprehensively. In this study, we regularly conducted seawater sampling along the northeastern Pacific coast of Japan between March 2012 and May 2016. We collected 213 seawater samples and prepared size-based fractions to generate 454 subsets of samples for shotgun metagenome sequencing and analysis. We also determined the sequences of 16S rRNA (n = 111) and 18S rRNA (n = 47) gene amplicons from smaller sample subsets. We thereafter developed the Ocean Monitoring Database for time-series metagenomic data (http://marine-meta.healthscience.sci.waseda.ac.jp/omd/), which provides a three-dimensional bird’s-eye view of the data. This database includes results of digital DNA chip analysis, a novel method for estimating ocean characteristics such as water temperature from metagenomic data. Furthermore, we developed a novel classification method that includes more information about viruses than that acquired using BLAST. We further report the discovery of a large number of previously overlooked (TAG)n repeat sequences in the genomes of marine microbes. We predict that the availability of this time-series database will lead to major discoveries in marine microbiome research.

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