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      A decade of metaproteomics: Where we stand and what the future holds

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          Abstract

          We are living through exciting times during which we are able to unravel the “microbial dark matter” in and around us through the application of high‐resolution “meta‐omics”. Metaproteomics offers the ability to resolve the major catalytic units of microbial populations and thereby allows the establishment of genotype‐phenotype linkages from in situ samples. A decade has passed since the term “metaproteomics” was first coined and corresponding analyses were carried out on mixed microbial communities. Since then metaproteomics has yielded many important insights into microbial ecosystem function in the various environmental settings where it has been applied. Although initial progress in analytical capacities and resulting numbers of proteins identified was extremely fast, this trend slowed rapidly. Here, we discuss several representative metaproteomic investigations of activated sludge, acid mine drainage biofilms, freshwater and seawater microbial communities, soil, and human gut microbiota. By using these case studies, we highlight current challenges and possible solutions for metaproteomics to realize its full potential, i.e. to enable conclusive links between microbial community composition, physiology, function, interactions, ecology, and evolution in situ.

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          Most cited references63

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          Pyrosequencing enumerates and contrasts soil microbial diversity.

          Estimates of the number of species of bacteria per gram of soil vary between 2000 and 8.3 million (Gans et al., 2005; Schloss and Handelsman, 2006). The highest estimate suggests that the number may be so large as to be impractical to test by amplification and sequencing of the highly conserved 16S rRNA gene from soil DNA (Gans et al., 2005). Here we present the use of high throughput DNA pyrosequencing and statistical inference to assess bacterial diversity in four soils across a large transect of the western hemisphere. The number of bacterial 16S rRNA sequences obtained from each site varied from 26,140 to 53,533. The most abundant bacterial groups in all four soils were the Bacteroidetes, Betaproteobacteria and Alphaproteobacteria. Using three estimators of diversity, the maximum number of unique sequences (operational taxonomic units roughly corresponding to the species level) never exceeded 52,000 in these soils at the lowest level of dissimilarity. Furthermore, the bacterial diversity of the forest soil was phylum rich compared to the agricultural soils, which are species rich but phylum poor. The forest site also showed far less diversity of the Archaea with only 0.009% of all sequences from that site being from this group as opposed to 4%-12% of the sequences from the three agricultural sites. This work is the most comprehensive examination to date of bacterial diversity in soil and suggests that agricultural management of soil may significantly influence the diversity of bacteria and archaea.
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            Computational improvements reveal great bacterial diversity and high metal toxicity in soil.

            The complexity of soil bacterial communities has thus far confounded effective measurement. However, with improved analytical methods, we show that the abundance distribution and total diversity can be deciphered. Reanalysis of reassociation kinetics for bacterial community DNA from pristine and metal-polluted soils showed that a power law best described the abundance distributions. More than one million distinct genomes occurred in the pristine soil, exceeding previous estimates by two orders of magnitude. Metal pollution reduced diversity more than 99.9%, revealing the highly toxic effect of metal contamination, especially for rare taxa.
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              A Core Human Microbiome as Viewed through 16S rRNA Sequence Clusters

              We explore the microbiota of 18 body sites in over 200 individuals using sequences amplified V1–V3 and the V3–V5 small subunit ribosomal RNA (16S) hypervariable regions as part of the NIH Common Fund Human Microbiome Project. The body sites with the greatest number of core OTUs, defined as OTUs shared amongst 95% or more of the individuals, were the oral sites (saliva, tongue, cheek, gums, and throat) followed by the nose, stool, and skin, while the vaginal sites had the fewest number of OTUs shared across subjects. We found that commonalities between samples based on taxonomy could sometimes belie variability at the sub-genus OTU level. This was particularly apparent in the mouth where a given genus can be present in many different oral sites, but the sub-genus OTUs show very distinct site selection, and in the vaginal sites, which are consistently dominated by the Lactobacillus genus but have distinctly different sub-genus V1–V3 OTU populations across subjects. Different body sites show approximately a ten-fold difference in estimated microbial richness, with stool samples having the highest estimated richness, followed by the mouth, throat and gums, then by the skin, nasal and vaginal sites. Richness as measured by the V1–V3 primers was consistently higher than richness measured by V3–V5. We also show that when such a large cohort is analyzed at the genus level, most subjects fit the stool “enterotype” profile, but other subjects are intermediate, blurring the distinction between the enterotypes. When analyzed at the finer-scale, OTU level, there was little or no segregation into stool enterotypes, but in the vagina distinct biotypes were apparent. Finally, we note that even OTUs present in nearly every subject, or that dominate in some samples, showed orders of magnitude variation in relative abundance emphasizing the highly variable nature across individuals.
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                Author and article information

                Journal
                Proteomics
                Proteomics
                10.1002/(ISSN)1615-9861
                PMIC
                Proteomics
                John Wiley and Sons Inc. (Hoboken )
                1615-9853
                1615-9861
                10 September 2015
                October 2015
                : 15
                : 20 , Microbiomes ( doiID: 10.1002/pmic.v15.20 )
                : 3409-3417
                Affiliations
                [ 1 ] Luxembourg Centre for Systems BiomedicineUniversity of Luxembourg Esch‐sur‐AlzetteLuxembourg
                [ 2 ] Advanced Water Management CentreUniversity of Queensland BrisbaneAustralia
                Author notes
                [*] [* ] Correspondence: Professor Paul Wilmes, Luxembourg Centre for Systems Biomedicine, University of Luxembourg, 7 avenue des Hauts‐Fourneaux, L‐4362 Esch‐sur‐Alzette, Luxembourg

                E‐mail: paul.wilmes@ 123456uni.lu

                Fax: +352‐46 66 44 6949

                [†]

                Additional corresponding author: Dr. Philip L. Bond,

                E‐mail: phil.bond@ 123456awmc.uq.edu.au

                Article
                PMIC12114
                10.1002/pmic.201500183
                5049639
                26315987
                1b440251-7f3e-4ab9-a586-de41a2a8acde
                © 2015 The Authors. PROTEOMICS published by Wiley‐VCH Verlag GmbH & Co. KGaA, Weinheim.

                This is an open access article under the terms of the Creative Commons Attribution‐NonCommercial‐NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non‐commercial and no modifications or adaptations are made.

                History
                : 15 May 2015
                : 06 July 2015
                : 05 August 2015
                Page count
                Pages: 9
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                2.0
                pmic12114
                October 2015
                Converter:WILEY_ML3GV2_TO_NLMPMC version:4.9.4 mode:remove_FC converted:04.10.2016

                Molecular biology
                integrated omics,metagenomics,metaproteomics,microbial community,microbial systems ecology,microbiology

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