7 metagenomics sheds light on the ecology of marine microbes and their viruses

1、TIMI 1588 No. of Pages 11ReviewMetagenomics Sheds Light on the Ecology of Marine Microbes and TheirVirusesFelipe Hernandes Coutinho,1,2 Gustavo Bueno Gregoracci,3 Juline Marta Walter,1 Cristiane Carneiro Thompson,1 and Fabiano L. Thompson1,4,*Advances brought about by omics-based approaches have rev

2、olutionized our understanding of the diversity and ecological processes involving marine archaea, bacteria, and their viruses. This broad review discusses recent exam- ples of how genomics, metagenomics, and ecogenomics have been applied to reveal the ecology of these biological entities. Three majo

3、r topics are covered in this revision: (i) the novel roles of microorganisms in ecosystem processes; (ii) virushost associations; and (iii) ecological associations of microeukaryotes and other microbes. We also briey comment on the discovery of novel taxa from marine ecosystems; development of a rob

4、ust taxonomic framework for prokaryotes; breakthroughs on the diversity and ecology of cyanobacteria; and advances on ecological modelling. We conclude by discussing limitations of the eld and suggesting directions for future research.ARobustFrameworkforStudyingMicrobialEcologyWhichecologicalprocess

5、estakeplace withinthemarinemicrobiome?Answeringthisquestion has been a central goal of microbial ecology for decades. Microbial ecologists strive to understand how environmental factors act together to modulate community assembly (see Glossary) taking place within microbial communities, uxes of biog

6、eochemical cycles, and interactions between biological entities ranging from ecosystems to genes. This informa- tion provides mechanistic insights which allow us to predict the responses of microbial communities to ecosystem changes, and possibly to manipulate them to perform specic functions 1,2. U

7、nderstanding how microbial communities function has never been more relevant in light of the current scenario of global climate changes and ecosystem alterations.1Laboratory of Microbiology, Instituto de Biologia, Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil 2Evolutionary Ge

8、nomics Group, Departamento de Produccon Vegetal y Microbiologa, Universidad Miguel Hernndez (UMH), Alicante, Spain 3Department of Marine Sciences, Federal University of So Paulo (Unifesp), Santos, Brazil4Center of Technology CT2, SAGE- COPPE, Federal Universidade Federal do Rio de Janeiro (UFRJ), Ri

9、o de Janeiro, BrazilIt is impossible to comprehensively understand the functioning of any ecosystem without knowing which organisms are part of it. Thus, determining the taxonomic composition of microbial communities in situ is the standing stone on which studies of microbial ecology are built. Cult

10、ure-independent approaches (e.g., metagenomics and single-cell genomics) played a fundamental role in achieving this goal. We are now able to chart the biodiversity of ecologically relevant organisms that have long been overlooked by culture-based methods.Metagenome-assembled genomes (MAGs) and sing

11、le amplied genomes (SAGs) successfully revealed thousands of novel microbial genomes, and phylogenetic reconstruction revealed that some are entirely novel branches of the microbial tree of life. Marine ecosystems have consistently been a source of these novel evolutionary lineages. One single study

12、 has described nearly 8000 high-quality MAGs, revealing 17 novel uncultured bacterial phyla (UBP) and three novel uncultured archaeal phyla (UAP) 3. Another major advancement regarding the diversity of Archaea was the discovery of the Asgard superphylum. The implications of this*Correspondence: fabi

13、ano.thompsonbiologia.ufrj.br (F.L. Thompson)./10.1016/j.tim.2018.05.015 1 2018 Published by Elsevier Ltd.Trends in Microbiology, Month Year, Vol. xx, No. yyHighlightsMultipleassociationsof marinemicrobes with each other, and with their environ- ment, have recently been discovered, shed

14、ding light on their ecological niches and the community assembly process.Culture-independent approaches have drasticallyexpanded our knowledge of the diversity of marine bacteria, archaea, microeukaryotes and viruses, redening our understanding of the evolution of life on Earth.Omics-based approache

15、s provided deeper insights into the contributions of both novel and previously known taxa to biogeochemical cycles and other ecosystem-sustaining processes.Most ecogenomics studies are focused on Picocyanobacteria and SAR11 from the euphotic zone, while othertaxa and ecosystems have been neglected.M

16、etagenomics revealed the massive diversity of marine viruses and showed that their inuence over host commu- nities is much broader than previously conceived.TIMI 1588No. of Pages 11nding go beyond the eld of microbiology as phylogenomic data suggested that one of the phyla within the Asgard group wa

17、s the ancestor of Earths eukaryotic life 4.This novel diversity must be robustly classied and organized into units with evolutionary and ecological relevance (Box 1). Only with a robust framework for classication and nomenclature can we investigate microbial communities across ecological gradients a

18、nd integrate the obtained information into ecological models (Figure 1, Key Figure). Since most of these new genomes had little to no associated phenotype information, phylogenomics and genomic taxonomy had a fundamental role in the classication of these new taxa. One such example was the recent res

19、tructuring of the taxonomy of Cyanobacteria through an ecoge- nomics-based approach (Box 2). In this review we provide a broad overview of the most successful approaches within the eld of microbial ecology aimed at deciphering ecology of the marinemicrobiomeachievedthroughgenomics,metagenomics,andec

20、ogenomics,published between 2013 and early 2018. Through the text, the term microbe is used to refer to bacteria, archaeaandviruses, whicharethefocusofthisreview, whileunicellulareukaryotesarereferred to as microeukaryotes.NewMicrobialRolesinMarineEcosystemProcessesMicrobes play a fundamental role i

21、n sustaining processes that are essential to life on our planet, including: primary productivity, nutrient cycling, and host physiology 5,6. MAGs and SAGs have been particularly useful for identifying potential roles of prokaryotes in biogeochemical cycles.Thesenewlydiscoveredorganismsareinvolvedinc

22、yclesofcarbon,nitrogenandsulfur, so that their discovery adds new players to the list of known organisms that contribute to energy and matter cycles that were previously unaccounted for. Early studies of SAGs revealed that SAR324 gammaproteobacteria from the deep ocean rely on sulfur oxidation to po

23、wer carbon xationthroughtheCalvinBensonBasshamcycle, whichchallengedtheconceptthatcarbon xation in the dark ocean is driven by crenarchaeota using the 3-hydroxypropionate/4-hydrox- ybutyrate cycle 7. SAGs have also demonstrated that light-independent carbon xation2 Trends in Microbiology, Month Year

24、, Vol. xx, No. yyGlossaryCommunity assembly: the combination of stochastic and deterministic processes that together shape the taxonomic composition of biological communities.Culture-based methods: approaches based on isolating microorganisms or viruses from their original environment and cultivatin

25、g them in the laboratory, usually in axenic cultures. If properly executed, allows the separation of a single population from the community.Ecogenomics: a eld that aims to understand the ecology of microorganisms based on their genomes/metagenomes and the evolutionary and ecological patterns that ca

26、n be derived from them.Ecological niche: the multidimensional space volume of abiotic (environmental parameters) and biotic (other organisms and viruses) conditions in which an organism can survive, grow, and reproduce.Genomic taxonomy: classication of novel organisms or identication of known taxa (

27、e.g., genera, species, or strains) through comparative analysis of their genomic content.Lysogenic viral infection: viral infection process in which the viral genome is replicated alongside the host genome without the production of the new viral particles or cell burst, thus avoiding host death.Lyti

28、c viral infection: viral infection process that culminates in death of the host cell due to bursting and release of the newly produced viral particles.Metagenome-assembled genome (MAG): complete or partial genome sequences obtained by assembling reads from one or multiple metagenomes followed by bin

29、ning together the assembled contigs.Metagenomics: analysis of genomic sequences of a whole community, directly from a sample without any prior cultivation steps.Microbial community: an assemblage of potentially interacting microorganisms within dened spatial and temporal limits.Microeukaryotes: sing

30、le-celled, microscopic eukaryotic microbes, that is, the protists.Phylogenomics: reconstruction of phylogenies based on multiple genesBox 1. Understanding Microbial Evolution, and the Emergence of Ecological Strategies through EcogenomicsOne major impediment for studying microbial communities in sit

31、u is the taxonomic classication of microbes. Classical phenotype-basedapproaches often fail to distinguish between taxa at ecologically relevant scales. Genomic taxonomy represents an alternative to bypass the limitations of classical approaches by establishing a solid framework for the identication

32、 and classication of microbial taxa into more natural units 77. Genomic taxonomy is dened as an integrated comparative approach that covers multiple genomic traits 78, including: pan and core genome delimitation, average amino acid identity (AAI), average nucleotide identity (ANI), in silico DNADNA

33、hybridization, nucleotide signature analysis 79, and genotype-to-phenotype predictions 80, which dene the presence of diagnostic phe- notypes on the basis of gene content. Genomic taxonomy and phylogenomics are increasingly prevalent approaches for the discovery of new taxa and for the reclassicatio

34、n of previously known groups of microbes 8183. The shift from phenotype- to genotype-based taxonomy has redened the species concept of Bacteria and Archaea. Under a genomic perspective, microbial taxa can be dened as monophyletic clusters of organisms sharing higher genomic similarity with each othe

35、r than with any other clades. Thus, taxa can be viewed as clusters of gene-ow units subjected to natural selection 84. Evolutionary models and genome analysis showed that, in spite of the potential for horizontal gene transfer (HGT), natural selection is required for cluster formation 77. Thus, sele

36、ction acts on gene frequencies over generations, optimizing the tness of taxa totheir ecosystem. This is exemplied by the rapid evolution of adaptive traits among copiotrophic bacteria of the genus Vibrio, in which closely related populations that belong to the same species can display very distinct

37、 ecological strategies. Comparative genomics demonstrated that theexible portion of Vibrio genomes evolves quickly through point mutations and homologous recombination. These changes lead to fast adaptations to rapidly changing environmental conditions, allowing populations to differ into two major

38、ecological niches: particle-attached and swimmers 86. Alternatively, the process of niche partitioning among this genus can also be driven by HGT, as demonstrated by the fast differentiation of niches among alginate-degrading communities of Vibrio, that over short evolutionary times can specialize i

39、nto roles of pioneers, harvesters, and scavengers 106. Findings such as these demonstrate that even the microscale resource gradients of the marine ecosystem can produce gene-ow barriers that eventually lead to speciation.TIMI 1588 No. of Pages 11Key FigureApproaches to Study the Ocean Microbiome th

40、rough Ecogenomics and MetagenomicsOcean microbiomePhysicalmeasurementsChemicalanalysesShotgunmetagenomics (taxonomy + funcons)Ampliconmetagenomics(taxonomy)SAGs(new microbes or viruses + metabolism)MAGs(new microbes or viruses + metabolism)GenomictaxonomyEcogenomicsCo-occurrencenetworksMetabolicMeta

41、bolicmodelling Trophic strategiesstrategySymbiocinteraconsEcologicalnicheClassificaonphylogenyEcological modelsBloom forecasngDisease outbreak prevenon Climate change responses(See figure legend on the bottom of the next page.)Trends in Microbiology, Month Year, Vol. xx, No. yy 3(often the entire co

42、re genome of a group) yielding more robust results than strategies based on a single or a small number of phylogenetic markers.Single amplied genome (SAG): complete or partial genome sequence obtained by sequencing the amplied DNA of a previously isolated single cell or viral particle.TIMI 1588 No.

43、of Pages 11strategies (e.g., the WoodLjungdahl pathway) are much more widespread in members of the Bacteria and Archaea than originally believed 8. Multiple MAGs of the candidate phyla radiation were obtained from terrestrial subsurface samples. Among those were included multiple genomes postulated

44、to be involved in processes of carbon, sulfur and nitrogen cycling, such as CO2 and N2 xation, and H2 production and oxidation 9. This discovery provided a new picture of the occurrence of these metabolisms among the microbial diversity. Yet, the presence and activity levels of the members of the ca

45、ndidate phyla radiation in marine ecosystems have not been reported so far. Thus, the contributions of these organisms to nutrient cycles that take place in marine habitats have not been elucidated.Paradigms about the metabolic routes of well characterized processes have been challenged by newly dis

46、covered MAGs. For example, the two-step process of ammonia oxidation to nitrate, through nitrite, was originally believed to be carried out separately by consortia of ammonia-oxidizing and nitrite-oxidizing microorganisms. But MAGs of the phylum Nitrospira were shown to be capable of performing both

47、 steps of the ammonia oxidation process 10. Finally, recovery of MAGs from a deep aquifer led to the discovery of the phylum Bathyarchaeota 11. The genome content of these organisms revealed that methane metab- olism is not restricted to archaea of the phylum Euryarchaeota, as previously believed 11

48、.Ecogenomics has also revealed new ecological roles for microorganisms that have been the subject of extensive research for decades, such as SAR11. This clade was originally believed to be limitedtooxicmarine environments, buta combinationofsingle-cellgenomics andmetagenomics revealed that some line

49、ages within SAR11 thrive in oxygen-minimum zones 12. This lineage of SAR11 has a functional nitrate reductase enzyme that performs the rst step of the denitrication process, demonstrating that members of SAR11 can contribute to oceanic nitrogen loss. The strong similarity between oxic and anoxic lin

50、eages suggests that adaptation to anoxic environ- ments has emerged recently during the evolution of this group12. Similarly, surface and deep- ocean lineages of SAR11 were shown to have a surprisingly similar genetic content but subtle differences in genome size, intergenic spacers, and amino-acid

51、content that might be linked to adaptations to these two different ecological niches 13. These subtle differences between oxic/anoxic and surface/deep lineages of SAR11 demonstrate that this group can rapidly evolve changes that allow these bacteria to expand their range of niches. For a comprehensi

52、ve review on the ecology and evolution of SAR11 readers are referred to 14.Figure 1. Microbial concentration strategies often involve ltration. Viral concentration strategies often involve tangential ow ltration or iron chloride occulation, the latter offering quantitative samples. MAG, metagenome-a

53、ssembled genome; SAG, single amplied genome.4 Trends in Microbiology, Month Year, Vol. xx, No. yyBox 2. Ecogenomics Redenes the Taxonomy and Ecology of CyanobacteriaCyanobacteria are the main primary producers in marine habitats and therefore have a fundamental role on sustaining marine ecosystems.

54、The taxonomy of this phylum was originally established based on morphological traits. Multiple phylogenetic reconstructions demonstrated that this taxonomy disagreed with the phylogeny of this phylum, and that many of the taxa within Cyanobacteria were not monophyletic and were in need of revision 8

55、789. Thus, phyloge- nomics and genomic taxonomy were applied to redene the taxonomy of this phylum. This strategy led to the delineation of 57 genera (of which 28 are new), and 87 species (32 new) 83. In addition, based on the abundance of the newly established taxa in a global-scale metagenomic dat

56、aset, and their associations with environmental parameters, members of these taxa were assigned to three major ecological niches: Low Temperature, Low-Tem- perature Copiotrophs, and High-Temperature Oligotrophs. These groups were coherently linked to taxa dened through genomic taxonomy and phylogeno

57、mics, elucidating how different patterns of niche occupancy are distributed among this phylum, thus expanding the ecotype concept that was originally limited to Prochlorococcus and Para- synechococcus (formerly marine Synechococcus) 90 to the whole of this phylum.TIMI 1588No. of Pages 11The Neglecte

58、d Majority: Ecology of Marine VirusesFor decades, viruses were neglected by microbial ecologists due to a lack of efcient methods for their quantication and taxonomic classication. Since the discovery that viruses are the most abundant biological entities in the oceans 15, microbiologists began to characterize their ecological roles, especially of those which infect bacteria and archaea. Viruses are more than mere infectious particles (virions), and their rich biology emerges when they interact with their hosts, which are thought to i