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Microbes drive ecosystem functioning and their viruses modulate these impacts through mortality, gene transfer and metabolic reprogramming. Despite the importance of virus-host interactions and likely variable infection efficiencies of individual phages across hosts, such variability is seldom quantified. Here, we quantify infection efficiencies of 38 phages against 19 host strains in aquatic Cellulophaga (Bacteroidetes) phage-host model systems. Binary data revealed that some phages infected only one strain while others infected 17, whereas quantitative data revealed that efficiency of infection could vary 10 orders of magnitude, even among phages within one population. This provides a baseline for understanding and modeling intrapopulation host range variation. Genera specific host ranges were also informative. For example, the Cellulophaga Microviridae, showed a markedly broader intra-species host range than previously observed in Escherichia coli systems. Further, one phage genus, Cba41, was examined to investigate nonheritable changes in plating efficiency and burst size that depended on which host strain it most recently infected. While consistent with host modification of phage DNA, no differences in nucleotide sequence or DNA modifications were detected, leaving the observation repeatable, but the mechanism unresolved. Overall, this study highlights the importance of quantitatively considering replication variations in studies of phage-host interactions.
Assuntos
Bacteriófagos/fisiologia , Bacteroidetes/virologia , Microviridae/fisiologia , Bacteriófagos/genética , Bacteroidetes/genética , Bacteroidetes/fisiologia , Replicação do DNA , Escherichia coli/fisiologia , Escherichia coli/virologia , Especificidade de Hospedeiro , Microviridae/genética , Replicação ViralRESUMO
Bacterial viruses (phages) influence global biogeochemical cycles by modulating bacterial mortality, metabolic output and evolution. However, our understanding of phage infections is limited by few methods and environmentally relevant model systems. Prior work showed that Cellulophaga baltica phage Ï38:1 infects its original host lytically, and an alternative host either delayed lytically or lysogenically. Here we investigate these infections through traditional and marker-based approaches, and introduce geneELISA for high-throughput examination of phage-host interactions. All methods confirmed the lytic, original host infection (70-80 min latent period; approximately eight phages produced per cell), but alternative host assays were more challenging. A 4.5 h experiment detected no phage production by plaque assay, whereas phageFISH and geneELISA revealed phage genome replication and a latent period ≥ 150 min. Longer experiments (26 h) suggested an 11 h latent period and a burst size of 871 by plaque assay, whereas phageFISH identified cell lysis starting at < 5 h and lasting to 11 h, but for only 7% to 21.5% of infected cells, respectively, and with â¼ 39 phages produced per cell. These findings help resolve the nature of the alternative host infection as delayed lytic and offer solutions to methodological challenges for studying inefficient phage-host interactions.
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Bacteriólise , Bacteroidetes/virologia , Especificidade de Hospedeiro/fisiologia , Interações Hospedeiro-Patógeno/fisiologia , Podoviridae/patogenicidade , Bacteroidetes/metabolismo , Ensaio de Imunoadsorção Enzimática , Hibridização in Situ Fluorescente , Dados de Sequência Molecular , Podoviridae/genéticaRESUMO
Bacterial viruses (phages) are abundant, ecologically important biological entities. However, our understanding of their impact is limited by model systems that are primarily not well represented in nature, e.g. Enterophages and their hosts. Here, we investigate genomic characteristics and infection strategies among six aquatic Bacteroidetes phages that represent two genera of exceptionally large (â¼70-75 kb genome) podoviruses, which were isolated from the same seawater sample using Cellulophaga baltica as host. Quantitative host range studies reveal that these genera have contrasting narrow (specialist) and broad (generalist) host ranges, with one-step growth curves revealing reduced burst sizes for the generalist phages. Genomic comparisons suggest candidate genes in each genus that might explain this host range variation, as well as provide hypotheses about receptors in the hosts. One generalist phage, φ38:1, was more deeply characterized, as its infection strategy switched from lytic on its original host to either inefficient lytic or lysogenic on an alternative host. If lysogenic, this phage was maintained extrachromosomally in the alternative host and could not be induced by mitomycin C. This work provides fundamental knowledge regarding phage-host ranges and their genomic drivers while also exploring the 'host environment' as a driver for switching phage replication mode.
Assuntos
Bacteriófagos/genética , Bacteroidetes/virologia , Cromossomos Bacterianos , Genoma Viral , Especificidade de Hospedeiro/genética , Podoviridae/genética , Bacteriófagos/classificação , Bacteriófagos/patogenicidade , Bacteroidetes/genética , Genômica , Lisogenia , Podoviridae/classificação , Podoviridae/patogenicidade , Água do Mar/microbiologiaRESUMO
Viruses impact microbial systems through killing hosts, horizontal gene transfer, and altering cellular metabolism, consequently impacting nutrient cycles. A virus-infected cell, a "virocell," is distinct from its uninfected sister cell as the virus commandeers cellular machinery to produce viruses rather than replicate cells. Problematically, virocell responses to the nutrient-limited conditions that abound in nature are poorly understood. Here we used a systems biology approach to investigate virocell metabolic reprogramming under nutrient limitation. Using transcriptomics, proteomics, lipidomics, and endo- and exo-metabolomics, we assessed how low phosphate (low-P) conditions impacted virocells of a marine Pseudoalteromonas host when independently infected by two unrelated phages (HP1 and HS2). With the combined stresses of infection and nutrient limitation, a set of nested responses were observed. First, low-P imposed common cellular responses on all cells (virocells and uninfected cells), including activating the canonical P-stress response, and decreasing transcription, translation, and extracellular organic matter consumption. Second, low-P imposed infection-specific responses (for both virocells), including enhancing nitrogen assimilation and fatty acid degradation, and decreasing extracellular lipid relative abundance. Third, low-P suggested virocell-specific strategies. Specifically, HS2-virocells regulated gene expression by increasing transcription and ribosomal protein production, whereas HP1-virocells accumulated host proteins, decreased extracellular peptide relative abundance, and invested in broader energy and resource acquisition. These results suggest that although environmental conditions shape metabolism in common ways regardless of infection, virocell-specific strategies exist to support viral replication during nutrient limitation, and a framework now exists for identifying metabolic strategies of nutrient-limited virocells in nature.
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Bacteriófagos , Bacteriófagos/genética , Bacteriófagos/fisiologia , Proteômica , Fosfatos/metabolismo , Metabolômica , Biologia de Sistemas , Transcriptoma , Reprogramação MetabólicaRESUMO
BACKGROUND: Microbiomes are now recognized as the main drivers of ecosystem function ranging from the oceans and soils to humans and bioreactors. However, a grand challenge in microbiome science is to characterize and quantify the chemical currencies of organic matter (i.e., metabolites) that microbes respond to and alter. Critical to this has been the development of Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS), which has drastically increased molecular characterization of complex organic matter samples, but challenges users with hundreds of millions of data points where readily available, user-friendly, and customizable software tools are lacking. RESULTS: Here, we build on years of analytical experience with diverse sample types to develop MetaboDirect, an open-source, command-line-based pipeline for the analysis (e.g., chemodiversity analysis, multivariate statistics), visualization (e.g., Van Krevelen diagrams, elemental and molecular class composition plots), and presentation of direct injection high-resolution FT-ICR MS data sets after molecular formula assignment has been performed. When compared to other available FT-ICR MS software, MetaboDirect is superior in that it requires a single line of code to launch a fully automated framework for the generation and visualization of a wide range of plots, with minimal coding experience required. Among the tools evaluated, MetaboDirect is also uniquely able to automatically generate biochemical transformation networks (ab initio) based on mass differences (mass difference network-based approach) that provide an experimental assessment of metabolite connections within a given sample or a complex metabolic system, thereby providing important information about the nature of the samples and the set of microbial reactions or pathways that gave rise to them. Finally, for more experienced users, MetaboDirect allows users to customize plots, outputs, and analyses. CONCLUSION: Application of MetaboDirect to FT-ICR MS-based metabolomic data sets from a marine phage-bacterial infection experiment and a Sphagnum leachate microbiome incubation experiment showcase the exploration capabilities of the pipeline that will enable the research community to evaluate and interpret their data in greater depth and in less time. It will further advance our knowledge of how microbial communities influence and are influenced by the chemical makeup of the surrounding system. The source code and User's guide of MetaboDirect are freely available through ( https://github.com/Coayala/MetaboDirect ) and ( https://metabodirect.readthedocs.io/en/latest/ ), respectively. Video Abstract.
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Ecossistema , Metabolômica , Espectrometria de Massas/métodos , Metabolômica/métodos , Software , SoloRESUMO
The fate of oceanic carbon and nutrients depends on interactions between viruses, prokaryotes, and unicellular eukaryotes (protists) in a highly interconnected planktonic food web. To date, few controlled mechanistic studies of these interactions exist, and where they do, they are largely pairwise, focusing either on viral infection (i.e., virocells) or protist predation. Here we studied population-level responses of Synechococcus cyanobacterial virocells (i.e., cyanovirocells) to the protist Oxyrrhis marina using transcriptomics, endo- and exo-metabolomics, photosynthetic efficiency measurements, and microscopy. Protist presence had no measurable impact on Synechococcus transcripts or endometabolites. The cyanovirocells alone had a smaller intracellular transcriptional and metabolic response than cyanovirocells co-cultured with protists, displaying known patterns of virus-mediated metabolic reprogramming while releasing diverse exometabolites during infection. When protists were added, several exometabolites disappeared, suggesting microbial consumption. In addition, the intracellular cyanovirocell impact was largest, with 4.5- and 10-fold more host transcripts and endometabolites, respectively, responding to protists, especially those involved in resource and energy production. Physiologically, photosynthetic efficiency also increased, and together with the transcriptomics and metabolomics findings suggest that cyanovirocell metabolic demand is highest when protists are present. These data illustrate cyanovirocell responses to protist presence that are not yet considered when linking microbial physiology to global-scale biogeochemical processes.
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Viruses that infect microbial hosts have traditionally been studied in laboratory settings with a focus on either obligate lysis or persistent lysogeny. In the environment, these infection archetypes are part of a continuum that spans antagonistic to beneficial modes. In this Review, we advance a framework to accommodate the context-dependent nature of virus-microorganism interactions in ecological communities by synthesizing knowledge from decades of virology research, eco-evolutionary theory and recent technological advances. We discuss that nuanced outcomes, rather than the extremes of the continuum, are particularly likely in natural communities given variability in abiotic factors, the availability of suboptimal hosts and the relevance of multitrophic partnerships. We revisit the 'rules of life' in terms of how long-term infections shape the fate of viruses and microbial cells, populations and ecosystems.
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Bacteriófagos/crescimento & desenvolvimento , Bacteriófagos/genética , Evolução Biológica , Genes Virais , Interações Hospedeiro-Patógeno/genéticaRESUMO
Ecosystems are controlled by 'bottom-up' (resources) and 'top-down' (predation) forces. Viral infection is now recognized as a ubiquitous top-down control of microbial growth across ecosystems but, at the same time, cell death by viral predation influences, and is influenced by, resource availability. In this Review, we discuss recent advances in understanding the biogeochemical impact of viruses, focusing on how metabolic reprogramming of host cells during lytic viral infection alters the flow of energy and nutrients in aquatic ecosystems. Our synthesis revealed several emerging themes. First, viral infection transforms host metabolism, in part through virus-encoded metabolic genes; the functions performed by these genes appear to alleviate energetic and biosynthetic bottlenecks to viral production. Second, viral infection depends on the physiological state of the host cell and on environmental conditions, which are challenging to replicate in the laboratory. Last, metabolic reprogramming of infected cells and viral lysis alter nutrient cycling and carbon export in the oceans, although the net impacts remain uncertain. This Review highlights the need for understanding viral infection dynamics in realistic physiological and environmental contexts to better predict their biogeochemical consequences.
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Organismos Aquáticos/virologia , Interações entre Hospedeiro e Microrganismos , Metabolismo , Água do Mar/microbiologia , Replicação Viral , Vírus/crescimento & desenvolvimento , Ecossistema , Oceanos e MaresRESUMO
Ocean viruses are abundant and infect 20-40% of surface microbes. Infected cells, termed virocells, are thus a predominant microbial state. Yet, virocells and their ecosystem impacts are understudied, thus precluding their incorporation into ecosystem models. Here we investigated how unrelated bacterial viruses (phages) reprogram one host into contrasting virocells with different potential ecosystem footprints. We independently infected the marine Pseudoalteromonas bacterium with siphovirus PSA-HS2 and podovirus PSA-HP1. Time-resolved multi-omics unveiled drastically different metabolic reprogramming and resource requirements by each virocell, which were related to phage-host genomic complementarity and viral fitness. Namely, HS2 was more complementary to the host in nucleotides and amino acids, and fitter during infection than HP1. Functionally, HS2 virocells hardly differed from uninfected cells, with minimal host metabolism impacts. HS2 virocells repressed energy-consuming metabolisms, including motility and translation. Contrastingly, HP1 virocells substantially differed from uninfected cells. They repressed host transcription, responded to infection continuously, and drastically reprogrammed resource acquisition, central carbon and energy metabolisms. Ecologically, this work suggests that one cell, infected versus uninfected, can have immensely different metabolisms that affect the ecosystem differently. Finally, we relate phage-host genome complementarity, virocell metabolic reprogramming, and viral fitness in a conceptual model to guide incorporating viruses into ecosystem models.
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Bacteriófagos/fisiologia , Pseudoalteromonas/virologia , Bacteriófagos/genética , Ecologia , Ecossistema , Microbiologia Ambiental , Vírus/genéticaRESUMO
We report here the genome sequences and morphological characterizations of phages p000v and p000y, which infect the bacterial pathogen Shiga-toxigenic Escherichia coli O157:H7 and which are potential candidates for phage therapy against such pathogens.
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Phage-host interactions are critical to ecology, evolution, and biotechnology. Central to those is infection efficiency, which remains poorly understood, particularly in nature. Here we apply genome-wide transcriptomics and proteomics to investigate infection efficiency in nature's own experiment: two nearly identical (genetically and physiologically) Bacteroidetes bacterial strains (host18 and host38) that are genetically intractable, but environmentally important, where phage infection efficiency varies. On host18, specialist phage phi18:3 infects efficiently, whereas generalist phi38:1 infects inefficiently. On host38, only phi38:1 infects, and efficiently. Overall, phi18:3 globally repressed host18's transcriptome and proteome, expressed genes that likely evaded host restriction/modification (R/M) defenses and controlled its metabolism, and synchronized phage transcription with translation. In contrast, phi38:1 failed to repress host18's transcriptome and proteome, did not evade host R/M defenses or express genes for metabolism control, did not synchronize transcripts with proteins and its protein abundances were likely targeted by host proteases. However, on host38, phi38:1 globally repressed host transcriptome and proteome, synchronized phage transcription with translation, and infected host38 efficiently. Together these findings reveal multiple infection inefficiencies. While this contrasts the single mechanisms often revealed in laboratory mutant studies, it likely better reflects the phage-host interaction dynamics that occur in nature.
Assuntos
Bacteriófagos/genética , Bacteriófagos/fisiologia , Bacteroidetes/virologia , Proteoma/genética , Transcriptoma , Bacteroidetes/fisiologia , Flavobacteriaceae/fisiologia , Flavobacteriaceae/virologia , Genômica , Metabolômica , Mutação , Biossíntese de Proteínas , Proteômica , Análise de Sequência de RNA , Transcrição GênicaRESUMO
Hemolyticâ»uremic syndrome is a life-threating disease most often associated with Shiga toxin-producing microorganisms like Escherichia coli (STEC), including E. coli O157:H7. Shiga toxin is encoded by resident prophages present within this bacterium, and both its production and release depend on the induction of Shiga toxin-encoding prophages. Consequently, treatment of STEC infections tend to be largely supportive rather than antibacterial, in part due to concerns about exacerbating such prophage induction. Here we explore STEC O157:H7 prophage induction in vitro as it pertains to phage therapy-the application of bacteriophages as antibacterial agents to treat bacterial infections-to curtail prophage induction events, while also reducing STEC O157:H7 presence. We observed that cultures treated with strictly lytic phages, despite being lysed, produce substantially fewer Shiga toxin-encoding temperate-phage virions than untreated STEC controls. We therefore suggest that phage therapy could have utility as a prophylactic treatment of individuals suspected of having been recently exposed to STEC, especially if prophage induction and by extension Shiga toxin production is not exacerbated.
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Viruses that infect bacteria (phages) can influence bacterial community dynamics, bacterial genome evolution and ecosystem biogeochemistry. These influences differ depending on whether phages establish lytic, chronic or lysogenic infections. Although the first two produce virion progeny, with lytic infections resulting in cell destruction, phages undergoing lysogenic infections replicate with cells without producing virions. The impacts of lysogeny are numerous and well-studied at the cellular level, but ecosystem-level consequences remain underexplored compared to those of lytic infections. Here, we review lysogeny from molecular mechanisms to ecological patterns to emerging approaches of investigation. Our goal is to highlight both its diversity and importance in complex communities. Altogether, using a combined viral ecology toolkit that is applied across broad model systems and environments will help us understand more of the diverse lifestyles and ecological impacts of lysogens in nature.
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Bactérias/virologia , Bacteriófagos/fisiologia , Microbiologia Ambiental , Lisogenia/fisiologia , Fenômenos Fisiológicos Virais , Bactérias/genéticaRESUMO
Bacteria impact humans, industry and nature, but do so under viral constraints. Problematically, knowledge of viral infection efficiencies and outcomes derives from few model systems that over-represent efficient lytic infections and under-represent virus-host natural diversity. Here we sought to understand infection efficiency regulation in an emerging environmental Bacteroidetes-virus model system with markedly different outcomes on two genetically and physiologically nearly identical host strains. For this, we quantified bacterial virus (phage) and host DNA, transcripts and phage particles throughout both infections. While phage transcriptomes were similar, transcriptional differences between hosts suggested host-derived regulation of infection efficiency. Specifically, the alternative host overexpressed DNA degradation genes and underexpressed translation genes, which seemingly targeted phage DNA particle production, as experiments revealed they were both significantly delayed (by >30 min) and reduced (by >50%) in the inefficient infection. This suggests phage failure to repress early alternative host expression and stress response allowed the host to respond against infection by delaying phage DNA replication and protein translation. Given that this phage type is ubiquitous and abundant in the global oceans and that variable viral infection efficiencies are central to dynamic ecosystems, these data provide a critically needed foundation for understanding and modeling viral infections in nature.
Assuntos
Bacteriófagos/isolamento & purificação , Bacteroidetes/virologia , Água do Mar/virologia , Bacteriófagos/classificação , Bacteriófagos/genética , Bacteriófagos/fisiologia , Bacteroidetes/fisiologia , Oceanos e Mares , Proteínas Virais/genética , Proteínas Virais/metabolismoRESUMO
This corrects the article DOI: 10.1038/ismej.2016.81.