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Illuminating the human virome in health and disease

DOI:10.1186/s13073-020-00766-x
Authors:
Fatemeh Adiliaghdam at Massachusetts General Hospital
Fatemeh Adiliaghdam
Kate L Jeffrey at Massachusetts General Hospital, Harvard Medical School, Boston USA
Kate L Jeffrey
  • Massachusetts General Hospital, Harvard Medical School, Boston USA
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Abstract

Abstract Although the microbiome is established as an important regulator of health and disease, the role of viruses that inhabit asymptomatic humans (collectively, the virome) is less defined. While we are still characterizing what constitutes a healthy or diseased virome, an exciting next step is to move beyond correlations and toward identification of specific viruses and their precise mechanisms of beneficial or harmful immunomodulation. Illuminating this will represent a first step toward developing virome-focused therapies.
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C O M M E N T Open Access
Illuminating the human virome in health
and disease
Fatemeh Adiliaghdam
1
and Kate L. Jeffrey
1,2*
Abstract
Although the microbiome is established as an important regulator of health and disease, the role of viruses that
inhabit asymptomatic humans (collectively, the virome) is less defined. While we are still characterizing what
constitutes a healthy or diseased virome, an exciting next step is to move beyond correlations and toward
identification of specific viruses and their precise mechanisms of beneficial or harmful immunomodulation.
Illuminating this will represent a first step toward developing virome-focused therapies.
Late to the party
In the study of microorganisms, bacteria frequently steal
the limelight. During an influenza outbreak in late 1800,
it was the bacterium Haemophilus influenzae isolated
from sputum that was first presumed to cause disease.
During the 1918 influenza pandemic, urgent efforts to
isolate this causative bacterium failed and it was not
until the 1930s that a filterable agent, a virus, Influenza
H1N1, was identified as the culprit [1]. Similarly, in the
pursuit of understanding human commensal microor-
ganisms, the last 20 years of research has focused almost
exclusively on bacteria and their regulation of our im-
mune and nervous systems. In comparison, very little is
known about eukaryotic and prokaryotic viruses that
also inhabit asymptomatic humans. Given that the name
virus was coined from the Latin word meaning slimy li-
quid or poison and that viruses are considered obligate
pathogens, a possibly “beneficial virome”is surprising to
many.
The late start for viruses in the commensal micro-
organism field is in large part due to our inability to
readily culture or detect them, as was the case during
the discovery of the influenza virus. We do not yet know
the eukaryotic cell or bacterial host of most viruses, and
there is no universal 16S ribosomal RNA equivalent, as
in bacteria, allowing for rapid taxonomic
characterization. Technologies such as metagenomics
have only recently enabled identification of viruses in
healthy human tissues. This initially involved sequencing
all DNA or RNA in a sample (human, bacterial, and
viral), and computationally aligning the massive number
of sequences to identify those that resemble known viral
genes. An improvement on this approach now involves
filtering samples to purge eukaryotic cells and bacteria
so that only virus-like particles (VLPs) remain for se-
quencing. However, since the virome consists of both
temperate bacteriophages within bacterial genomes and
free VLPs, both total and VLP sequencing will likely pro-
vide greater representation of all viruses. Nonetheless,
with the approaches taken thus far, studies have revealed
viruses are abundant in human feces, blood, skin, lung,
oral cavity, and an array of other tissues of healthy and
diseased individuals [2–5].
A moving target
The human intestinal virome established at birth is
dominated by bacteria-infecting viruses, while eukaryotic
viruses gradually emerge after birth [6]. One gram of hu-
man feces contains around 10
8
–10
9
VLPs, and explora-
tory sequencing has shown that the identifiable fraction
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* Correspondence: KJeffrey@mgh.harvard.edu
1
Department of Medicine, Division of Gastroenterology and the Center for
the Study of Inflammatory Bowel Disease, Massachusetts General Hospital,
Harvard Medical School, Boston, MA 02114, USA
2
Center for Microbiome Informatics and Therapeutics, Massachusetts Institute
of Technology, Cambridge, MA 02139, USA
Adiliaghdam and Jeffrey Genome Medicine (2020) 12:66
https://doi.org/10.1186/s13073-020-00766-x
of this virome is primarily bacteriophages, including
dsDNA Caudovirales, ssDNA Microviridae [2], and the
recently identified predominant CrAssphage [7]. Viruses
that infect eukaryotic cells within the human fecal vir-
ome have been identified to belong to families Astroviri-
dae,Anelloviridae,Picornaviridae,Caliciviridae, and
Herpesviridae, among others [2,3].
However, many hurdles in our ability to catalog the
human virome remain, making this data far from final.
(1) The vast majority of viruses share little to no hom-
ology with annotated viruses in reference databases. Vi-
ruses infecting animals, plants, fungi, and protozoans
(collectively eukaryotic viruses) number around 100 mil-
lion species while those infecting bacteria are estimated
at 10 trillion, yet a large proportion remain unannotated.
While the NCBI Virus Portal (https://www.ncbi.nlm.nih.
gov/labs/virus/vssi/#/) is reporting new annotations at
an exponential rate, virologists have struggled to distin-
guish clear classes and kingdoms for most of the viro-
sphere, with the exception of a recently published
taxonomic hierarchy [8]. Thus, reanalysis of relatively re-
cent human virome publications may already be war-
ranted. (2) Computational analysis methods vary
considerably across virome studies, as this is a nascent
field, making direct comparisons difficult. (3) Viruses
rely on the host organism for successful replication;
therefore, the discovery of viruses that specifically infect
human cells may be better achieved by analysis of less
accessible tissue and cells, rather than feces or bodily
fluids, where they are likely scarce. (4) False positives in
sequencing data remain an issue as many sequencing re-
agents, or DNA spike-ins during sequencing, are derived
from bacteriophages or bacteria carrying bacteriophages.
A consensus on laboratory techniques and computa-
tional analysis pipelines is a much-needed advance in
the virome field. (5) We still have a very limited perspec-
tive on the healthy human tissue-specific virome. We
know little about the virome in individuals from differ-
ent geographic locations, in those consuming different
diets, and in old or young individuals, and thus, it is dif-
ficult to discern cases of vertical and horizontal trans-
mission or composition changes before, during, and
after disease onset. A tighter grasp on what a healthy vir-
ome looks like—an equivalent to the Human Micro-
biome Project—would allow clearer inferences about
how the virome influences disease.
Making the leap from correlation to causation
Despite the present limitations in characterizing the hu-
man virome in health, robust fluctuations in the virome
in multiple diseases have been reported. In inflammatory
bowel disease (IBD), it was found that enteric Caudovir-
ales temperate phage expanded, although the degree that
this was due to alterations in bacteria remains
unresolved [2,3]. In colorectal cancer, virome signatures
were shown to differentiate individuals at the early ver-
sus late stages of disease. In type I diabetes, expanded
enteric bacteriophage diversity was found to precede dis-
ease and Circoviridae eukaryotic viruses were enriched
in controls. In cystic fibrosis, sputum phage communi-
ties were highly similar and eukaryotic viral communities
were found to be dominated by herpesviruses and retro-
viruses. In graft-versus-host disease (GVHD), a progres-
sive expansion of eukaryotic gut viruses was shown to
follow hematopoietic stem cell transplant, and picobir-
naviruses were associated with early post-transplant
GVHD. In HIV patients, low peripheral CD4 T cell
counts were associated with an expansion of enteric ade-
noviruses [5]. However, this cataloging of diseased vir-
omes is vastly outpacing our mechanistic understanding
as we still do not know whether these altered viromes
actually contribute to disease.
A lesson from the microbiome field, at this juncture
in virome research, would be to move beyond correla-
tions and toward a detailed analysis of how certain vi-
ruses autonomously or cooperatively educate our
physiology. Functional studies in mice have found
that enteric viruses inhabiting a healthy host provide
immune and gut homeostasis. Depletion of viruses or
virus receptors in healthy mice exacerbates intestinal
inflammation while treatment with viral ligands pro-
tects from disease [5]. However, precise mechanisms
by which individual viruses provide protection are
limited. Furthermore, how human virome composition
impacts health or disease remains ambiguous as direct
functional studies are currently lacking. However,
both prokaryotic and eukaryotic viromes possess the
ability to directly immunomodulate their human hosts
based on reports of trans-kingdom interactions of
bacteriophage and human immune cells [9]. Further-
more, given that viruses exist in complex communi-
ties comprising bacteria, fungi, and protozoans,
indirect outcomes of virome changes will almost def-
initely occur through alteration of surrounding micro-
organism communities. Another question and
appealing avenue of investigation is whether com-
mensal viruses impact the host’sabilitytofight
pathogenic viruses through tonic stimulation of anti-
viral immunity or if conversely, acute virus infection
impacts the resident virome. Finally, since complex
disease phenotypes are the result of environmental
triggers in the context of genetic susceptibility, vari-
able impact of the virome will depend on host genet-
ics and should be considered. For instance, a loss-of-
function variant of host virus receptor MDA-5
(encoded by gene IFIH1) associates with incidence of
IBD but protects from type I diabetes [5]suggesting
divergent roles for viruses in different contexts.
Adiliaghdam and Jeffrey Genome Medicine (2020) 12:66 Page 2 of 3
Translating to diagnosis and therapy
The ultimate goal of virome research is to translate find-
ings into diagnostic and therapeutic opportunities. With
accurate mapping of the virome in different human tis-
sues in healthy and disease states, we can begin to use
certain viruses as biomarkers or attempt to manipulate
virome signatures. Moving from association to causation
will confidently enable us to harness the healthy virome
or disrupt the disease-associated one. Uncoupling the
roles of eukaryotic and prokaryotic viruses on immune
state and improvements on propagation of individual
candidate viruses for functional studies will advance
these goals. Therapeutic avenues could also focus on the
beneficial or detrimental host immune responses to vi-
ruses, rather than the viruses themselves to mitigate
virome-related diseases. Viruses may also serve an im-
portant role in fecal microbiota transplants (FMTs) since
filtered feces (removing the bacterial component) have
the same efficacy in treating the Clostridium difficile pa-
tients [10]. However, there is still no knowledge of the
virome composition of FMTs and no consensus on if
this is something we should be measuring in donor and
recipient patients. Finally, benign viruses within the
healthy virome could conceivably be used for safe gene
delivery into humans.
We have just begun to reveal the complexities and
promise of the virome using computational genomics,
but application of the virome remains relatively underex-
plored. Many challenges in the virome field remain, but
let us not repeat history and let bacteria steal the lime-
light. Viruses, fungi, and other commensals within the
human microorganism ecosystem are likely equally im-
portant; we just need to overcome a few more hurdles to
realize their full potential.
Abbreviations
DNA: Deoxyribonucleic acid; RNA: Ribonucleic acid; dsDNA: Double-stranded
DNA; ssDNA: Single-stranded DNA; NCBI: National Center for Biotechnology
Information; HIV: Human immunodeficiency virus; MDA-5: Melanoma
differentiation-associated protein 5
Acknowledgements
The authors would like to thank Hajera Amatullah and Roshan Ahmed for
editorial comments.
Authors’contributions
KLJ conceived and FA and KLJ wrote the manuscript. All authors read and
approved the final manuscript.
Funding
Kenneth Rainin Foundation (Innovator and Synergy Awards to KLJ), NIH
R21AI144877 (KLJ), NIH R01DK119996 (KLJ), Harvard Catalyst | The Harvard
Clinical and Translational Science Center (National Center for Advancing
Translational Sciences, National Institutes of Health Award UL 1TR002541)
and financial contributions from Harvard University and its affiliated
academic healthcare centers (KLJ), MGH Research Scholar, class of 2020 (KLJ).
Availability of data and materials
Not applicable.
Ethics approval and consent to participate
Not applicable.
Consent for publication
Not applicable.
Competing interests
The authors declare that they have no competing interests.
Received: 15 July 2020 Accepted: 16 July 2020
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Publisher’sNote
Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations.
Adiliaghdam and Jeffrey Genome Medicine (2020) 12:66 Page 3 of 3
... However, most sequencing data on the healthy human virome are derived from biological fluids (e.g. feces, respiratory secretions, urine, saliva, mucosal or dermal swabs; reviewed in Supplementary Table S1) (11)(12)(13), which provide only a fragmentary landscape of the viral populations residing within tissues. In fact, based on numerous qPCR studies from biopsies, eukaryotic DNA viruses are expected to be more prevalent in our organs (8,(14)(15)(16)(17)(18)(19)(20)(21), in contrast to bacteriophages that predominate in most secretions (22)(23)(24)(25). ...
... In addition to the sample type, other reasons for the overall low frequency (prevalence) of human eukaryotic DNA viruses in current metagenomic studies may be ascribed to laboratory processing: the ratios of viral lineages can be distorted by different enrichment methods (e.g. virus-like particles, multiple displacement amplification, rolling circle amplification, nuclease treatment), and the true occurrence of resident viruses may be grossly underestimated by the pooling and sequencing strategies employed (13,22,26,27). ...
... The findings provide a 2 Nucleic Acids Research, 2023 comprehensive atlas of the numerous known viruses persisting within us. Importantly, we show that an understanding of the core prevalences, quantities and, most importantly, an integral analysis of different tissues are essential for discriminating virome deviations in various disease states (9,13,(31)(32)(33)(34)(35)(36). ...
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Little is known on the landscape of viruses that reside within our cells, nor on the interplay with the host imperative for their persistence. Yet, a lifetime of interactions conceivably have an imprint on our physiology and immune phenotype. In this work, we revealed the genetic make-up and unique composition of the known eukaryotic human DNA virome in nine organs (colon, liver, lung, heart, brain, kidney, skin, blood, hair) of 31 Finnish individuals. By integration of quantitative (qPCR) and qualitative (hybrid-capture sequencing) analysis, we identified the DNAs of 17 species, primarily herpes-, parvo-, papilloma- and anello-viruses (>80% prevalence), typically persisting in low copies (mean 540 copies/ million cells). We assembled in total 70 viral genomes (>90% breadth coverage), distinct in each of the individuals, and identified high sequence homology across the organs. Moreover, we detected variations in virome composition in two individuals with underlying malignant conditions. Our findings reveal unprecedented prevalences of viral DNAs in human organs and provide a fundamental ground for the investigation of disease correlates. Our results from post-mortem tissues call for investigation of the crosstalk between human DNA viruses, the host, and other microbes, as it predictably has a significant impact on our health.
View
... Indeed, the presence of many virus types infecting eukaryotic or prokaryotic cells has been unfolded via high-throughput sequencing. However, most sequencing data on the healthy human virome are derived from biological uids (e.g., feces, respiratory secretions, urine, saliva, mucosal or dermal swabs; reviewed in Supplemental Table 1) 10,12,13 , which provide only a fragmentary landscape of the viral populations residing within tissues. In fact, based on numerous qPCR studies from biopsies, eukaryotic DNA viruses are expected to be more prevalent in our organs 8,[14][15][16][17][18][19][20] , in contrast to bacteriophages that predominate in most secretions [21][22][23] . ...
... In addition to the sample type, other reasons for the overall low prevalence of human eukaryotic DNA viruses in current metagenomic studies may be ascribed to laboratory processing: the ratios of viral lineages can be distorted by different enrichment methods (e.g., virus-like particles, multiple displacement ampli cation, rolling circle ampli cation, nuclease treatment), and the true occurrence of resident viruses may be grossly underestimated by the pooling and sequencing strategies employed 13,21,24,25 . ...
... The ndings provide a comprehensive atlas of the numerous viruses persisting within us. Importantly, we show that an understanding of the core prevalences, quantities and, most importantly, an integral analysis of different tissues are essential for discriminating virome deviations in various disease states 9,13,[29][30][31][32][33][34] . ...
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Background Little is known on the landscape of viruses that have taken residence within our cells, nor on the interplay with the host imperative for their persistence. Yet, a lifetime of interactions conceivably have an imprint on our physiology and immune phenotype. Importantly, current metagenomics-insights on the healthy human virome are derived from bodily fluids, which are only a proxy of the true prevalence of human eukaryotic viruses within tissues. Most significantly, the virome’s systemic composition in an individual has thus far remained uncharted. Results In this work, we revealed the genetic make-up of the eukaryotic human DNA virome in the body and showed that each organ (colon, liver, lung, heart, brain, kidney, skin, blood, hair) has unique viral compositions. By integration of quantitative (qPCR) and qualitative (hybrid-capture sequencing) analysis, we identified the DNAs of 17 viruses, primarily herpes-, parvo-, papilloma- and anello-viruses (>80% prevalence), typically persisting in low copies (mean 540 copies/ million cells). The within-sample diversities (a-diversity) were highest in the lung, liver, colon, and kidney. We assembled in total 70 viral genomes (>90% breadth coverage), distinct in each of the individuals, and identified high sequence homology across the organs. Moreover, we detected variations in virome composition and distribution in two individuals with underlying malignant conditions. Conclusions Our findings reveal unprecedented prevalences of viral DNAs in human organs. We showed that bodily fluids, although accessible to sampling, fail to deliver a comprehensive view of the numerous intracellular viruses colonizing our tissues. Our data demonstrate how a multi-organ approach is essential for analyzing differing virome compositions in various disease states. This atlas provides a fundamental ground for the studies of correlates to disease, and for the interpretation of next-generation sequencing data in clinical virology. Ultimately, our findings call for investigation of the crosstalk between human DNA viruses, the host, and other microbes, as it predictably has a significant impact on our health.
View
... Furthermore, existing murine gut genomic databases exclude inflamed mice, and mice colonized by enteric pathogens (e.g., like Salmonella, Klebsiella, and Citrobacter), thus limiting the extension of these existing microbiome resources to pathobiome models. Despite being recognized as important contributors to human health [24], these existing curated murine gut microbial genome catalogs also lack virome sampling [25,26]. Accurate interrogation of complete microbial community functions during Salmonella infection requires comprehensive model-specific knowledge of gene content and community membership both in healthy and inflamed guts. ...
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Background The murine CBA/J mouse model widely supports immunology and enteric pathogen research. This model has illuminated Salmonella interactions with the gut microbiome since pathogen proliferation does not require disruptive pretreatment of the native microbiota, nor does it become systemic, thereby representing an analog to gastroenteritis disease progression in humans. Despite the value to broad research communities, microbiota in CBA/J mice are not represented in current murine microbiome genome catalogs. Results Here we present the first microbial and viral genomic catalog of the CBA/J murine gut microbiome. Using fecal microbial communities from untreated and Salmonella-infected, highly inflamed mice, we performed genomic reconstruction to determine the impacts on gut microbiome membership and functional potential. From high depth whole community sequencing (~ 42.4 Gbps/sample), we reconstructed 2281 bacterial and 4516 viral draft genomes. Salmonella challenge significantly altered gut membership in CBA/J mice, revealing 30 genera and 98 species that were conditionally rare and unsampled in non-inflamed mice. Additionally, inflamed communities were depleted in microbial genes that modulate host anti-inflammatory pathways and enriched in genes for respiratory energy generation. Our findings suggest decreases in butyrate concentrations during Salmonella infection corresponded to reductions in the relative abundance in members of the Alistipes. Strain-level comparison of CBA/J microbial genomes to prominent murine gut microbiome databases identified newly sampled lineages in this resource, while comparisons to human gut microbiomes extended the host relevance of dominant CBA/J inflammation-resistant strains. Conclusions This CBA/J microbiome database provides the first genomic sampling of relevant, uncultivated microorganisms within the gut from this widely used laboratory model. Using this resource, we curated a functional, strain-resolved view on how Salmonella remodels intact murine gut communities, advancing pathobiome understanding beyond inferences from prior amplicon-based approaches. Salmonella-induced inflammation suppressed Alistipes and other dominant members, while rarer commensals like Lactobacillus and Enterococcus endure. The rare and novel species sampled across this inflammation gradient advance the utility of this microbiome resource to benefit the broad research needs of the CBA/J scientific community, and those using murine models for understanding the impact of inflammation on the gut microbiome more generally. 8oDwaJeBPqYs8E78ifbGCvVideo Abstract
View
... Although one recent study suggested that aging is associated with a decline in viral richness and an increase of Crassvirales phages, the full effect of healthy aging on the gut virome and its implications on microbial ecology remains largely unexplored 4 . However, the virome's influence on the microbiome is probably profound, as viruses offer a myriad of selective advantages to bacterial hosts, which may in turn sustain gut homoeostasis in health or aggravate certain disease states 5 . and cognition in a mouse model 14 . ...
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Distinct gut microbiome ecology may be implicated in the prevention of aging-related diseases as it influences systemic immune function and resistance to infections. Yet, the viral component of the microbiome throughout different stages in life remains unexplored. Here we present a characterization of the centenarian gut virome using previously published metagenomes from 195 individuals from Japan and Sardinia. Compared with gut viromes of younger adults (>18 yr) and older individuals (>60 yr), centenarians had a more diverse virome including previously undescribed viral genera, such as viruses associated with Clostridia. A population shift towards higher lytic activity was also observed. Finally, we investigated phage-encoded auxiliary functions that influence bacterial physiology, which revealed an enrichment of genes supporting key steps in sulfate metabolic pathways. Phage and bacterial members of the centenarian microbiome displayed an increased potential for converting methionine to homocysteine, sulfate to sulfide and taurine to sulfide. A greater metabolic output of microbial hydrogen sulfide in centenarians may in turn support mucosal integrity and resistance to pathobionts.
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... Theoretically, this approach could detect all pathogens [12]. In recent years, wastewater surveillance by metagenomics has documented the dynamics of the relative abundance (RA) of bacterial pathogens in municipal wastewater [13,14], which is emerging as an early-warning tool for detecting viruses in the local community [15]. Therefore, this approach represents a promising direction for microbial risk assessment of wastewater. ...
Metagenomic sequencing combined with flow cytometry facilitated a novel microbial risk assessment framework for bacterial pathogens in municipal wastewater without cultivation
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A workflow that combined metagenomic sequencing with flow cytometry was developed. The absolute abundance of pathogens was accurately estimated in mock communities and real samples. Metagenome‐assembled genomes binned from metagenomic data set is robust in phylogenetic analysis and virulence profiling.
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... It has been suggested that the virome may be able to mold the immune system, affecting the development of asthma and respiratory diseases in childhood [5]. Specific viruses, or viral families may be able to modulate the immune responses and hence drive immune maturation, influence health and disease and be candidates for intervention strategies [6]. Clearly, infection with common cold viruses is closely linked with the development of respiratory and other allergic diseases [7,8]. ...
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Background: From early life, respiratory viruses are implicated in the development, exacerbation and persistence of respiratory conditions such as asthma. Complex dynamics between microbial communities and host immune responses, shape immune maturation and homeostasis, influencing health outcomes. We evaluated the hypothesis that the respiratory virome is linked to systemic immune responses, using peripheral blood and nasopharyngeal swab samples from preschool-age children in the PreDicta cohort. Methods: Peripheral blood mononuclear cells from 51 children (32 asthmatics, 19 healthy controls), participating in the 2-year multinational PreDicta cohort were cultured with bacterial (Bacterial-DNA, LPS) or viral (R848, Poly:IC, RV) stimuli. Supernatants were analyzed by Luminex for the presence of 22 relevant cytokines. Virome composition was obtained using untargeted high throughput sequencing of nasopharyngeal samples. The metagenomic data were used for the characterization of virome profiles and the presence of key viral families (Picornaviridae, Anelloviridae, Siphoviridae). These were correlated to cytokine secretion patterns, identified through hierarchical clustering and principal component analysis. Results: High spontaneous cytokine release was associated with increased presence of Prokaryotic virome profiles and reduced presence of Eukaryotic and Anellovirus profiles. Antibacterial responses did not correlate with specific viral families or virome profile, however, low antiviral responders had more Prokaryotic and less Eukaryotic virome profiles. Anelloviruses and Anellovirus-dominated profiles were equally distributed amongst immune response clusters. The presence of Picornaviridae and Siphoviridae was associated with low interferon-λ responses. Asthma or allergy did not modify these correlations. Conclusions: Antiviral cytokines responses at a systemic level reflect the upper airway virome composition. Individuals with low innate interferon responses have higher abundance of Picornaviruses (mostly Rhinoviruses) and bacteriophages. Bacteriophages, particularly Siphoviridae appear to be sensitive sensors of host antimicrobial capacity, while Anelloviruses are not correlated with TLR-induced immune responses.
View
... like Salmonella, Klebsiella, and Citrobacter), thus limiting the extension of these existing microbiome resources to pathobiome models. Despite being recognized as important contributors to human health [24], these existing curated murine gut microbial genome catalogs also lack virome sampling [25,26]. Accurate interrogation of complete microbial community functions during Salmonella infection requires comprehensive model-speci c knowledge of gene content and community membership both in healthy and in amed guts. ...
Exposing New Taxonomic Variation with Inflammation – A Murine Model-Specific Genome Database for Gut Microbiome Researchers
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Full-text available
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Background The murine CBA/J mouse model widely supports immunology and enteric pathogen research. This model has illuminated Salmonella interactions with the gut microbiome since pathogen proliferation does not require disruptive pretreatment of the native microbiota, nor does it become systemic, thereby representing an analog to gastroenteritis disease progression in humans. Despite the value to broad research communities, microbiota in CBA/J mice are not represented in current murine microbiome genome catalogs. Results Here we present the first microbial and viral genomic catalog of the CBA/J murine gut microbiome. Using fecal microbial communities from untreated and Salmonella-infected, highly inflamed mice, we performed genomic reconstruction to determine the impacts on gut microbiome membership and functional potential. From high depth whole community sequencing (~ 42.4 Gbps/sample), we reconstructed 2,281 bacterial and 4,516 viral draft genomes. Salmonella challenge significantly altered gut membership in CBA/J mice, revealing 30 genera and 98 species that were conditionally rare and unsampled in non-inflamed mice. Additionally, inflamed communities were depleted in microbial genes that modulate host anti-inflammatory pathways and enriched in genes for respiratory energy generation. Our findings suggest decreases in butyrate concentrations during Salmonella infection corresponded to reductions in the relative abundance in members of the Alistipes. Strain-level comparison of CBA/J microbial genomes to prominent murine gut microbiome databases identified newly sampled lineages in this resource, while comparisons to human gut microbiomes extended the host relevance of dominant CBA/J inflammation resistant strains. Conclusions This CBA/J microbiome database provides the first genomic sampling of relevant, uncultivated microorganisms within the gut from this widely used laboratory model. Using this resource, we curated a functional, strain-resolved view on how Salmonella remodels intact murine gut communities, advancing pathobiome understanding beyond inferences from prior amplicon-based approaches. Salmonella-induced inflammation suppressed Alistipes and other dominant members, while rarer commensals like Lactobacillus and Enterococcus endure. The rare and novel species sampled across this inflammation gradient advance the utility of this microbiome resource to benefit the broad research needs of the CBA/J scientific community, and those using murine models for understanding the impact of inflammation on the gut microbiome more generally.
View
... In this work, eleven taxonomic classification tools were compared 28 by analyzing their performances, in terms of sensitivity and precision, to classify reads at species 29 and family levels, using the same (viral and non-viral) datasets and evaluation metrics, as well as Viruses are the most abundant organisms on earth and can infect every living being, including 53 bacteria, plants, animals, and humans, among others. They are of great relevance since they are 54 the etiologic agents of many diseases that could cause common and severe illnesses [1,2]. They 55 also play a significant role in the dynamic of other ecosystems such as bacteria, being involved in 56 both, biogeochemical cycles, and food chains in different ecosystems. ...
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Viruses are the most abundant infectious agents on earth, and they infect living organisms such as bacteria, plants and animals, among others. They play an important role in the balance of different ecosystems by modulating microbial populations. In humans, they are responsible for some common diseases and may cause severe illnesses. Viral metagenomic studies have become essential and offer the possibility to understand and extend the knowledge of virus diversity and functionality. For these approaches, an essential step is the classification of viral sequences. In this work, 11 taxonomic classification tools were compared by analysing their performances, in terms of sensitivity and precision, to classify reads at the species and family levels using the same (viral and nonviral) datasets and evaluation metrics, as well as their processing times and memory requirements. The results showed that factors such as richness (numbers of viral species in samples), taxonomic level in the classification and read length influence tool performance. High values of viral richness in samples decreased the performances of most tools. Additionally, the classifications were better at higher taxonomic levels, such as families, compared to lower taxonomic levels, such as species, and were more evident in short reads. The results also indicated that BLAST and Kraken2 were the best tools for classifying all types of reads, while FastViromeExplorer and VirusFinder were only good when used for long reads and Centrifuge, DIAMOND, and One Codex when used for short reads. Regarding nonviral datasets (human and bacterial), all tools correctly classified them as nonviral.
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VC (virome-comparison): a novel approach to comparing viromes based on virus species specificity and virome specificity diversity
Article
The human virome, or the viral communities distributed on or in our body, is estimated to contain about 380 trillion of viruses (individuals), which has far reaching influences on our health and diseases. Obviously, the sheer numbers of viruses alone make the comparisons of two or multiple viromes extremely challenging. In fact, the theory of computation in computer science for so-termed NP-hard problems stipulates that the problem is unsolvable when the size of virome is sufficiently large even with fastest supercomputers. Practically, one has to develop heuristic and approximate algorithms to obtain practically satisfactory solutions for NP-hard problems. Here, we extend the species-specificity and specificity-diversity framework to develop a method for virome comparison (VC). The VC method consists of a pair of metrics: virus species specificity (VS) and virome specificity diversity (VSD) and corresponding pair of random search algorithms. Specifically, the VS and VS permutation (VSP) test can detect unique virus species (US) or enriched virus species (ES) in each virome (treatment), and the VSD and VSD permutation (VSDP) test can further determine holistic differences between two viromes or their subsets (assemblages of viruses). The test with four virome datasets demonstrated that the VC method is effective, efficient and robust. This article is protected by copyright. All rights reserved.
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Candida albicans-enteric viral interactions—The prostaglandin E2 connection and host immune responses
Article
Full-text available
  • Dec 2022
The human microbiome comprises trillions of microorganisms residing within different mucosal cavities and across the body surface. The gut microbiota modulates host susceptibility to viral infections in several ways, and microbial interkingdom interactions increase viral infectivity within the gut. Candida albicans, a frequently encountered fungal species in the gut, produces highly structured biofilms and eicosanoids such as prostaglandin E2 (PGE2), which aid in viral protection and replication. These biofilms encompass viruses and provide a shield from antiviral drugs or the immune system. Prostaglandin E2 is a key modulator of active inflammation with the potential to regulate interferon signalling upon microbial invasion or viral infections. In this review, we raise the perspective of gut interkingdom interactions involving C. albicans and enteric viruses, with a special focus on biofilms, PGE2 and viral replication. Ultimately, we discuss the possible implications of C. albicans-enteric virus associations on host-immune responses, particularly the interferon signalling pathway.
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The new scope of virus taxonomy: partitioning the virosphere into 15 hierarchical ranks
Article
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  • Apr 2020
Virus taxonomy emerged as a discipline in the middle of the twentieth century. Traditionally, classification by virus taxonomists has been focussed on the grouping of relatively closely related viruses. However, during the past few years, the International Committee on Taxonomy of Viruses (ICTV) has recognized that the taxonomy it develops can be usefully extended to include the basal evolutionary relationships among distantly related viruses. Consequently, the ICTV has changed its Code to allow a 15-rank classification hierarchy that closely aligns with the Linnaean taxonomic system and may accommodate the entire spectrum of genetic divergence in the virosphere. The current taxonomies of three human pathogens, Ebola virus, severe acute respiratory syndrome coronavirus and herpes simplex virus 1 are used to illustrate the impact of the expanded rank structure. This new rank hierarchy of virus taxonomy will stimulate further research on virus origins and evolution, and vice versa, and could promote crosstalk with the taxonomies of cellular organisms.
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The stepwise assembly of the neonatal virome is modulated by breastfeeding
Article
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The gut of healthy human neonates is usually devoid of viruses at birth, but quickly becomes colonized, which—in some cases—leads to gastrointestinal disorders1–4. Here we show that the assembly of the viral community in neonates takes place in distinct steps. Fluorescent staining of virus-like particles purified from infant meconium or early stool samples shows few or no particles, but by one month of life particle numbers increase to 10⁹ per gram, and these numbers seem to persist throughout life5–7. We investigated the origin of these viral populations using shotgun metagenomic sequencing of virus-enriched preparations and whole microbial communities, followed by targeted microbiological analyses. Results indicate that, early after birth, pioneer bacteria colonize the infant gut and by one month prophages induced from these bacteria provide the predominant population of virus-like particles. By four months of life, identifiable viruses that replicate in human cells become more prominent. Multiple human viruses were more abundant in stool samples from babies who were exclusively fed on formula milk compared with those fed partially or fully on breast milk, paralleling reports that breast milk can be protective against viral infections8–10. Bacteriophage populations also differed depending on whether or not the infant was breastfed. We show that the colonization of the infant gut is stepwise, first mainly by temperate bacteriophages induced from pioneer bacteria, and later by viruses that replicate in human cells; this second phase is modulated by breastfeeding.
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Bacteriophage trigger antiviral immunity and prevent clearance of bacterial infection
Article
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Phage subverts immune response Pseudomonas aeruginosa ( Pa ) is a multidrug-resistant Gramnegative bacterium commonly found in health care settings. Pa infections frequently result in considerable morbidity and mortality. Sweere et al. found that a type of temperate filamentous bacteriophage that infects and integrates into Pa is associated with chronic human wound infections. Likewise, wounds in mice colonized with phage-infected Pa were more severe and longer-lasting than those colonized by Pa alone. Immune cell uptake of phage-infected Pa resulted in phage RNA production and inappropriate antiviral immune responses, impeding bacterial clearance. Both phage vaccination and transfer of antiphage antibodies were protective against Pa infection. Science , this issue p. eaat9691
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The Intestinal Virome and Immunity
Article
Full-text available
  • Sep 2018
The composition of the human microbiome is considered a major source of interindividual variation in immunity and, by extension, susceptibility to diseases. Intestinal bacteria have been the major focus of research. However, diverse communities of viruses that infect microbes and the animal host cohabitate the gastrointestinal tract and collectively constitute the gut virome. Although viruses are typically investigated as pathogens, recent studies highlight a relationship between the host and animal viruses in the gut that is more akin to host-microbiome interactions and includes both beneficial and detrimental outcomes for the host. These viruses are likely sources of immune variation, both locally and extraintestinally. In this review, we describe the components of the gut virome, in particular mammalian viruses, and their ability to modulate host responses during homeostasis and disease.
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Efficacy of Sterile Fecal Filtrate Transfer for Treating Patients With Clostridium difficile Infection
Article
Full-text available
  • Nov 2016
Background & aims: Fecal microbiota transplantation (FMT) is a highly effective therapy for recurrent Clostridium difficile infection (CDI). However, transferring undefined living bacteria entails uncontrollable risks for infectious and metabolic or malignant diseases, particularly in immunocompromised patients. We investigated whether sterile fecal filtrates (containing bacterial debris, proteins, antimicrobial compounds, metabolic products and oligonucleotides/DNA), rather than intact microorganisms, are effective in patients with CDI. Methods: We performed a clinical case series to investigate the effects of fecal filtrate transfer (FFT) in 5 patients with symptomatic chronic-relapsing CDI at the Department of Internal Medicine I at the University Hospital Schleswig-Holstein (Kiel, Germany). Patients were followed for at least 6 months and up to 33 months. Stool was collected from 5 donors selected by the patients, and fully characterized according to FMT standards. Stool was sterile-filtered to remove small particles and bacteria; the filtrate was transferred to patients in a single administration via nasojejunal tube. Fecal samples were collected from patients before and 1 week and 6 weeks after FFT. Microbiome, virome, and proteome profiles of donors and patients were compared. Results: In all 5 patients, FFT restored normal stool habits and eliminated symptoms of CDI for a minimum period of 6 months. Proteome analyses of selected FFT filtrates revealed no obvious protein candidates associated with therapeutic efficacy. 16S rRNA gene sequencing detected diverse bacterial DNA signatures in the filtrates. Analysis of virus-like particles from a filtrate found to reduce symptoms of CDI revealed a complex signature of bacteriophages. Bacterial phylogeny and virome profile analyses of fecal samples from recipients indicated longitudinal changes in microbial and viral community structures after FFT. Conclusions: A preliminary investigation of 5 patients with CDI shows that transfer of sterile filtrates from donor stool (FFT), rather than fecal microbiota, can be sufficient to restore normal stool habits and eliminate symptoms. This finding indicates that bacterial components, metabolites or bacteriophages mediate many of the effects of FMT, and that FFT might be an alternative approach, particularly for immunocompromised patients.
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A Highly Abundant Bacteriophage Discovered in the Unknown Sequences of Human Faecal Metagenomes
Article
Full-text available
  • Jul 2014
Metagenomics, or sequencing of the genetic material from a complete microbial community, is a promising tool to discover novel microbes and viruses. Viral metagenomes typically contain many unknown sequences. Here we describe the discovery of a previously unidentified bacteriophage present in the majority of published human faecal metagenomes, which we refer to as crAssphage. Its ~97 kbp genome is six times more abundant in publicly available metagenomes than all other known phages together; it comprises up to 90% and 22% of all reads in virus-like particle (VLP)-derived metagenomes and total community metagenomes, respectively; and it totals 1.68% of all human faecal metagenomic sequencing reads in the public databases. The majority of crAssphage-encoded proteins match no known sequences in the database, which is why it was not detected before. Using a new co-occurrence profiling approach, we predict a Bacteroides host for this phage, consistent with Bacteroides-related protein homologues and a unique carbohydrate-binding domain encoded in the phage genome.
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Whole-Virome Analysis Sheds Light on Viral Dark Matter in Inflammatory Bowel Disease
Article
The human gut virome is thought to significantly impact the microbiome and human health. However, most virome analyses have been performed on a limited fraction of known viruses. Using whole-virome analysis on a published keystone inflammatory bowel disease (IBD) cohort and an in-house ulcerative colitis dataset, we shed light on the composition of the human gut virome in IBD beyond this identifiable minority. We observe IBD-specific changes to the virome and increased numbers of temperate phage sequences in individuals with Crohn's disease. Unlike prior database-dependent methods, no changes in viral richness were observed. Among IBD subjects, the changes in virome composition reflected alterations in bacterial composition. Furthermore, incorporating both bacteriome and virome composition offered greater classification power between health and disease. This approach to analyzing whole virome across cohorts highlights significant IBD signals, which may be crucial for developing future biomarkers and therapeutics.
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The virome hunters
Article
  • Oct 2018
Ambitious efforts to catalog viruses across the globe may facilitate our understanding of viral communities and ecology, boost infectious disease diagnostics and surveillance, and spur new therapeutics. Charles Schmidt investigates.
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Disease-Specific Alterations in the Enteric Virome in Inflammatory Bowel Disease
Article
Decreases in the diversity of enteric bacterial populations are observed in patients with Crohn's disease (CD) and ulcerative colitis (UC). Less is known about the virome in these diseases. We show that the enteric virome is abnormal in CD and UC patients. In-depth analysis of preparations enriched for free virions in the intestine revealed that CD and UC were associated with a significant expansion of Caudovirales bacteriophages. The viromes of CD and UC patients were disease and cohort specific. Importantly, it did not appear that expansion and diversification of the enteric virome was secondary to changes in bacterial populations. These data support a model in which changes in the virome may contribute to intestinal inflammation and bacterial dysbiosis. We conclude that the virome is a candidate for contributing to, or being a biomarker for, human inflammatory bowel disease and speculate that the enteric virome may play a role in other diseases.
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