Gut Virome Research Articles

Comparison of gut viral communities in children under 5 years old and newborns

ObjectivesThe gut virome of humans is mainly composed of bacteriophages and their role in shaping the gut microbiome and influencing human health is increasingly recognized. However, little is known about the dynamic changes of the gut virome in children and its role in growth and development. In this study, we collected fecal samples from newborns and children under 5 years old from the same area during the same time period to investigate the gut viral community using viral metagenomic technique.MethodsWe used viral metagenomics to compare the gut bacteriophage composition between newborns and children under 5 years of age. We collected fecal samples from 45 newborns who were born at the Affiliated Hospital of Jiangsu University and 45 healthy children who were examined at the same hospital. The two groups were classified as the newborn group and the children group.ResultsOur sequencing analysis showed that the number of seqeunce reads of the children group were more than that of the newborn group. The results of alpha diversity and beta diversity both indicated that the diversity of the children group was significantly higher than that of the newborn group and the children group is different from the newborn group. The abundance of gut virome in the children group was also higher than that in the newborn group. The analysis of the genetic characteristics of the viruses showed that the phage genome was scattered and clustered with specificity.ConclusionOur findings indicate that the gut bacteriophage communities undergo changes over time, presenting diversity and dynamic characteristics. We characterized the composition of gut virome in children and newborns in this region. However, further research is needed to investigate the function of bacteriophages in the ecology of the gastrointestinal tract.

Metagenomic Sequencing Identified Specific Bacteriophage Signature Discriminating between Healthy and Diarrheal Neonatal Piglets

Neonatal diarrhea is one of the most severe diseases in human beings and pigs, leading to high mortality and growth faltering. Gut microbiome-related studies mostly focus on the relationship between bacteria and neonatal diarrhea onset, and no research study has investigated the role of the gut virome in neonatal diarrhea. Here, using metagenomic sequencing, we characterized the fecal viral community of diarrheal and healthy neonatal piglets. We found that the viral community of diarrheal piglets showed higher individual heterogeneity and elevated abundance of Myoviridae. By predicting the bacterial host of the identified viral genomes, phages infecting Proteobacteria, especially E. coli, were the dominant taxa in neonatal diarrheal piglets. Consistent with this, the antibiotic resistance gene of E. coli origin was also enriched in neonatal diarrheal piglets. Finally, we established a random forest model to accurately discriminate between neonatal diarrheal piglets and healthy controls and identified genus E. coli- and genus listeria-infecting bacteriophages, including psa and C5 viruses, as key biomarkers. In conclusion, we provide the first glance of viral community and function characteristics in diarrheal and healthy neonatal piglets. These findings expand our understanding of the relationship among phages, bacteria and diarrhea, and may facilitate the development of therapeutics for the prevention and treatment of neonatal diarrhea.

Bacteriophages of the Order Crassvirales: What Do We Currently Know about This Keystone Component of the Human Gut Virome?

The order Crassvirales comprises dsDNA bacteriophages infecting bacteria in the phylum Bacteroidetes that are found in a variety of environments but are especially prevalent in the mammalian gut. This review summarises available information on the genomics, diversity, taxonomy, and ecology of this largely uncultured viral taxon. With experimental data available from a handful of cultured representatives, the review highlights key properties of virion morphology, infection, gene expression and replication processes, and phage-host dynamics.

Strain-Level Dynamics Reveal Regulatory Roles in Atopic Eczema by Gut Bacterial Phages.

The vast population of bacterial phages or viruses (virome) plays pivotal roles in the ecology of human microbial flora and health conditions. Obstacles, including poor viral sequence inference, strain-sensitive virus-host relationship, and the high diversity among individuals, hinder the in-depth understanding of the human virome. We conducted longitudinal studies of the virome based on constructing a high-quality personal reference metagenome (PRM). By applying long-read sequencing for representative samples, we could build a PRM of high continuity that allows accurate annotation and abundance estimation of viruses and bacterial species in all samples of the same individual by aligning short sequencing reads to the PRM. We applied this approach to a series of fecal samples collected for 6 months from a 2-year-old boy who had experienced a 2-month flare-up of atopic eczema (dermatitis) in this period. We identified 31 viral strains in the patient's gut microbiota and deciphered their strain-level relationship to their bacterial hosts. Among them, a lytic crAssphage developed into a dozen substrains and coordinated downregulation in the catabolism of aromatic amino acids (AAAs) in their host bacteria which govern the production of immune-active AAA derivates. The metabolic alterations confirmed based on metabolomic assays cooccurred with symptom remission. Our PRM-based analysis provides an easy approach for deciphering the dynamics of the strain-level human gut virome in the context of entire microbiota. Close temporal correlations among virome alteration, microbial metabolism, and disease remission suggest a potential mechanism for how bacterial phages in microbiota are intimately related to human health. IMPORTANCE The vast populations of viruses or bacteriophages in human gut flora remain mysterious. However, poor annotation and abundance estimation remain obstacles to strain-level analysis and clarification of their roles in microbiome ecology and metabolism associated with human health and diseases. We demonstrate that a personal reference metagenome (PRM)-based approach provides strain-level resolution for analyzing the gut microbiota-associated virome. When applying such an approach to longitudinal samples collected from a 2-year-old boy who has experienced a 2-month flare-up of atopic eczema, we observed thriving substrains of a lytic crAssphage, showing temporal correlation with downregulated catabolism of aromatic amino acids, lower production of immune-active metabolites, and remission of the disease. The PRM-based approach is practical and powerful for strain-centric analysis of the human gut virome, and the underlying mechanism of how strain-level virome dynamics affect disease deserves further investigation.

Viral Metagenomic Analysis of the Fecal Samples in Domestic Dogs (Canis lupus familiaris).

Canine diarrhea is a common intestinal illness that is usually caused by viruses, bacteria, and parasites, and canine diarrhea may induce morbidity and mortality of domestic dogs if treated improperly. Recently, viral metagenomics was applied to investigate the signatures of the enteric virome in mammals. In this research, the characteristics of the gut virome in healthy dogs and dogs with diarrhea were analyzed and compared using viral metagenomics. The alpha diversity analysis indicated that the richness and diversity of the gut virome in the dogs with diarrhea were much higher than the healthy dogs, while the beta diversity analysis revealed that the gut virome of the two groups was quite different. At the family level, the predominant viruses in the canine gut virome were certified to be Microviridae, Parvoviridae, Siphoviridae, Inoviridae, Podoviridae, Myoviridae, and others. At the genus level, the predominant viruses in the canine gut virome were certified to be Protoparvovirus, Inovirus, Chlamydiamicrovirus, Lambdavirus, Dependoparvovirus, Lightbulbvirus, Kostyavirus, Punavirus, Lederbergvirus, Fibrovirus, Peduovirus, and others. However, the viral communities between the two groups differed significantly. The unique viral taxa identified in the healthy dogs group were Chlamydiamicrovirus and Lightbulbvirus, while the unique viral taxa identified in the dogs with diarrhea group were Inovirus, Protoparvovirus, Lambdavirus, Dependoparvovirus, Kostyavirus, Punavirus, and other viruses. Phylogenetic analysis based on the near-complete genome sequences showed that the CPV strains collected in this study together with other CPV Chinese isolates clustered into a separate branch, while the identified CAV-2 strain D5-8081 and AAV-5 strain AAV-D5 were both the first near-complete genome sequences in China. Moreover, the predicted bacterial hosts of phages were certified to be Campylobacter, Escherichia, Salmonella, Pseudomonas, Acinetobacter, Moraxella, Mediterraneibacter, and other commensal microbiota. In conclusion, the enteric virome of the healthy dogs group and the dogs with diarrhea group was investigated and compared using viral metagenomics, and the viral communities might influence canine health and disease by interacting with the commensal gut microbiome.

Diversity and potential function of pig gut DNA viruses

Viruses are ubiquitous in the gut of animals and play an important role in the ecology of the gut microbiome. The potential effects of these substances on the growth and development of the body are not fully known. Little is known about the effects of breeding environment on pig gut virome. Here, there are 3584 viral operational taxonomic units (vOTUs) longer than 5 kb identified by virus-enriched metagenome sequencing from 25 pig fecal samples. Only a small minority of vOTUs (11.16%) can be classified at the family level, and ∼50% of the genes could be annotated, supporting the concept of pig gut as reservoirs of substantial undescribed viral genetic diversity. The composition of pig gut virome in the six regions may be related to geography. There are only 20 viral clusters (VCs) shared among pig gut virome in six regions of Shanxi Province. These viruses rarely carry antibiotic resistance genes (ARGs). At the same time, they possess abundant auxiliary metabolic genes (AMGs) potentially involved in carbon, sulfur metabolism and cofactor biosynthesis, etc. This study has revealed the unique characteristics and potential function of pig gut DNA virome and established a foundation for the recognition of the viral roles in gut environment.

Interpersonal variability of the human gut virome confounds disease signal detection in IBD

Viruses are increasingly recognised as important components of the human microbiome, fulfilling numerous ecological roles including bacterial predation, immune stimulation, genetic diversification, horizontal gene transfer, microbial interactions, and augmentation of metabolic functions. However, our current view of the human gut virome is tainted by previous sequencing requirements that necessitated the amplification of starting nucleic acids. In this study, we performed an original longitudinal analysis of 40 healthy control, 19 Crohn’s disease, and 20 ulcerative colitis viromes over three time points without an amplification bias, which revealed and highlighted the interpersonal individuality of the human gut virome. In contrast to a 16 S rRNA gene analysis of matched samples, we show that α- and β-diversity metrics of unamplified viromes are not as efficient at discerning controls from patients with inflammatory bowel disease. Additionally, we explored the intrinsic properties of unamplified gut viromes and show there is considerable interpersonal variability in viral taxa, infrequent longitudinal persistence of intrapersonal viruses, and vast fluctuations in the abundance of temporal viruses. Together, these properties of unamplified faecal viromes confound the ability to discern disease associations but significantly advance toward an unbiased and accurate representation of the human gut virome.

Multiomic spatial analysis reveals a distinct mucosa-associated virome

ABSTRACT The human gut virome has been increasingly explored in recent years. However, nearly all virome-sequencing efforts rely solely on fecal samples and few studies leverage multiomic approaches to investigate phage–host relationships. Here, we combine metagenomics, metaviromics, and metatranscriptomics to study virome-bacteriome interactions at the colonic mucosal-luminal interface in a cohort of three individuals with inflammatory bowel disease; non-IBD controls were not included in this study. We show that the mucosal viral population is distinct from the stool virome and houses abundant crAss-like phages that are undetectable by fecal sampling. Through viral protein prediction and metatranscriptomic analysis, we explore viral gene transcription, prophage activation, and the relationship between the presence of integrase and temperate phages in IBD subjects. We also show the impact of deep sequencing on virus recovery and offer guidelines for selecting optimal sequencing depths in future metaviromic studies. Systems biology approaches such as those presented in this report will enhance our understanding of the human virome and its interactions with our microbiome and our health.

The Human Gut Virome and Its Relationship with Nontransmissible Chronic Diseases.

The human gastrointestinal tract contains large communities of microorganisms that are in constant interaction with the host, playing an essential role in the regulation of several metabolic processes. Among the gut microbial communities, the gut bacteriome has been most widely studied in recent decades. However, in recent years, there has been increasing interest in studying the influences that other microbial groups can exert on the host. Among them, the gut virome is attracting great interest because viruses can interact with the host immune system and metabolic functions; this is also the case for phages, which interact with the bacterial microbiota. The antecedents of virome-rectification-based therapies among various diseases were also investigated. In the near future, stool metagenomic investigation should include the identification of bacteria and phages, as well as their correlation networks, to better understand gut microbiota activity in metabolic disease progression.

Gut virome-colonising Orthohepadnavirus genus is associated with ulcerative colitis pathogenesis and induces intestinal inflammation in vivo

ObjectivesUlcerative colitis (UC) is a chronic inflammatory disorder of unknown aetiology. Gut virome dysbiosis is fundamental in UC progression, although its role in the early phases of the disease is...

Identifying the Gut Virome of Diaphorina citri from Florida Groves.

Asian citrus psyllid (Diaphorina citri) transmits the bacterial pathogen Candidatus Liberibacter asiaticus (CLas), the putative causative agent of citrus Huanglongbing disease (HLB). Insect-specific viruses can act against insects as their natural enemies, and recently, several D. citri-associated viruses were discovered. The insect gut plays an important role as not only a pool for diverse microbes but also as a physical barrier to prevent the spread of pathogens such as CLas. However, there is little evidence of the presence of D. citri-associated viruses in the gut and of the interaction between them and CLas. Here, we dissected psyllid guts collected from five growing regions in Florida, and the gut virome was analyzed by high throughput sequencing. Four insect viruses, including D. citri-associated C virus (DcACV), D. citri densovirus (DcDV), D. citri reovirus (DcRV), and D. citri flavi-like virus (DcFLV), were identified, and their presence in the gut, including an additional D. citri cimodo-like virus (DcCLV), were confirmed with PCR-based assays. Microscopic analysis showed that DcFLV infection leads to morphological abnormalities in the nuclear structure in the infected psyllid gut cells. The complex and diverse composition of microbiota in the psyllid gut suggests a possible interaction and dynamics between CLas and the D. citri-associated viruses. Our study identified various D. citri-associated viruses that localized in the psyllid gut and provided more information that helps to evaluate the potential vectors for manipulating CLas in the psyllid gut.

Abstract TP238: Acute Ischemic Stroke Induces Gut Virome And Bacteriome Dysbiosis

We currently evaluated the effect of focal cerebral ischemia on gut virome and bacteriome in adult mice. Adult male C57BL/6 mice were subjected to transient middle cerebral artery occlusion or sham surgery. Virome and bacteriome were analyzed using shotgun metagenomics in the fecal samples collected from each mouse before and at 24h of reperfusion. Bioinformatics tools including VIBRANT (v1.2.1), DIAMOND Blastp (v0.9.14.115), VConTACT2 (v0.9.5), Cytoscape (v3.7.2), mash (v2.0), MUMmer (v3.1), Samtools (v1.11), DESeq2 (v1.28.1), MetaWRAP, and CRISPR Recognition Tool (v1.2) were used to assess viral networks, viral auxiliary metabolic genes, and viral protein network changes. Bacteriome was analyzed by kneaddata v0.7.2, MetaPhlAn version 2.7.7, and humann2 v2.8.1. Focal ischemia induced significant differences in fecal viral and bacterial taxa at the strain levels compared with sham. Furthermore, viral protein networks altered significantly after stroke. In particular, the clusters of Clostridia-like phages and Erysipelatoclostridiaceae phages showed a differential association between stroke and sham. In addition, we identified a significant reduction in the phages and bacteria of Lactobacillus after stroke. These studies indicate a possible viral-bacterial correlative change in the gut after stroke.

Longitudinal comparison of the developing gut virome in infants and their mothers

The human gut virome and its early life development are poorly understood. Prior studies have captured single-point assessments with the evolution of the infant virome remaining largely unexplored. We performed viral metagenomic sequencing on stool samples collected longitudinally from a cohort of 53 infants from age 2weeks to 3 years (80.7 billion reads), and from their mothers (9.8 billion reads) to examine and compare viromes. The asymptomatic infant virome consisted of bacteriophages, nonhuman dietary/environmental viruses, and human-host viruses, predominantly picornaviruses. In contrast, human-host viruses were largely absent from the maternal virome. Previously undescribed, sequence-divergent vertebrate viruses were detected in the maternal but not infant virome. As infants aged, the phage component evolved to resemble the maternal virome, but by age 3, the human-host component remained dissimilar from the maternal virome. Thus, early life virome development is determined predominantly by dietary, infectious, and environmental factors rather than direct maternal acquisition.

DOP53 Community types of the human gut virome are associated with endoscopic outcome in UC patients

Abstract Background Inflammatory bowel diseases (IBD) are a group of chronic inflammatory diseases of the gut. The pathophysiology is unknown; however, it is thought to result from an inappropriate immune reaction to the commensal gut microbiota. Community-typing is a common practice in bacteriome analysis allowing for the stratification of individuals based on their gut microbiome (eg., ‘enterotyping’). A viral counterpart of these enterotypes might allow stratification of individuals based on their gut virome. The aim of the present study is to investigate the existence of such viral community types and assess the impact of therapeutic outcome (and other covariates) on the gut virome in IBD patients. Methods Fecal samples were selected from 181 patients undergoing immunomodulatory therapy, and a baseline (pre-intervention) and a primary endpoint (post-intervention) have been collected for each patient. Viral metagenomics and deep sequencing were performed following viral enrichment with the NetoVIR protocol. Redundancy analysis and Dirichlet Multinomial Mixtures were applied to determine covariates of the virome composition and to condense the gut virota into viral community types, respectively. Results IBD patients were stratified based on unsupervised machine learning into two viral community types. Community type CA showed a low α-diversity and a high relative abundance of Caudoviricetes [non-CrAss] phages and was associated to the dysbiotic Bact2-enterotype. Community type CrM showed a high α-diversity and a high relative abundance of Caudoviricetes [CrAss] and Malgrandaviricetes phages. The gut virome variation was explained by several factors: patients’ individuality (75.8%), disease location (1.4%), age (0.5%) and faecal moisture (0.3%), with diagnosis not showing a non-redundant effect. Despite our expectations the choice of biological therapy did not show an association with the virome variation. During post-interventional analysis, endoscopic outcome (0.5%) was associated to gut virome variation as well. Remitting UC, but not CD, patients revealed a high percentage of community type CrM, a high Shannon diversity and a low phage lysogenic potential. Conversely, non-remitting UC, but not CD, patients revealed a high percentage of community type CA, a low Shannon diversity and a high lysogenic potential. Conclusion These findings suggest that viral community-typing allows for stratification of IBD patients based on their gut virome composition and might be a valuable tool to better understand IBD subtypes or as a potential future biomarker.

OP32 The gut virome-colonizing Orthohepadnavirus genus is associated with ulcerative colitis pathogenesis and induces intestinal inflammation in vivo

Abstract Background Ulcerative colitis (UC) is a chronic inflammatory disorder with an unknown etiology1. Over recent years, a growing body of evidence has been pinpointing gut virome dysbiosis as a fundamental component in its progression2, although its role during the early phases of chronic inflammation is far from being fully defined. Therefore, we sought to investigate the role of a virome-associated protein encoded by the Orthohepadnavirus genus, the Hepatitis B virus X protein (HBx)3, during UC aetiopathogenesis. Methods HBx positivity of UC patient-derived blood and gut mucosa was assessed by RT-PCR and Sanger sequencing correlated with clinical characteristics by multivariate analysis. Transcriptomics was performed on HBx-overexpressing endoscopic biopsies from healthy donors. C57BL/6 mice underwent intramucosal injections of liposome-conjugated HBx-encoding plasmids or the control. Multidimensional flow cytometry analysis was performed on colonic samples from HBx-treated and control animals. Transepithelial electrical resistance measurement, proliferation assay, ChIP-Seq, and RNA-Seq were performed on in vitro models of the gut barrier. Results HBx was detected in about 45% of patients with UC (Figure 1) and found to induce colonic inflammation in mice (Figure 2A-2D), while its silencing reverted the colitis phenotype in vivo (Figure 2E-2H). HBx acts as a transcriptional regulator in epithelial cells (Figure 3), provoking barrier leakage and altering inflammatory response (Figures 3 and 4). Conclusion This study paves the way for the understanding of the aetiopathogenesis of UC and provides a brand-new standpoint that looks at the virome as a target for tailored treatments, possibly leading to an entirely new approach to therapeutic intervention.

Evaluation of computational phage detection tools for metagenomic datasets.

As new computational tools for detecting phage in metagenomes are being rapidly developed, a critical need has emerged to develop systematic benchmarks. In this study, we surveyed 19 metagenomic phage detection tools, 9 of which could be installed and run at scale. Those 9 tools were assessed on several benchmark challenges. Fragmented reference genomes are used to assess the effects of fragment length, low viral content, phage taxonomy, robustness to eukaryotic contamination, and computational resource usage. Simulated metagenomes are used to assess the effects of sequencing and assembly quality on the tool performances. Finally, real human gut metagenomes and viromes are used to assess the differences and similarities in the phage communities predicted by the tools. We find that the various tools yield strikingly different results. Generally, tools that use a homology approach (VirSorter, MARVEL, viralVerify, VIBRANT, and VirSorter2) demonstrate low false positive rates and robustness to eukaryotic contamination. Conversely, tools that use a sequence composition approach (VirFinder, DeepVirFinder, Seeker), and MetaPhinder, have higher sensitivity, including to phages with less representation in reference databases. These differences led to widely differing predicted phage communities in human gut metagenomes, with nearly 80% of contigs being marked as phage by at least one tool and a maximum overlap of 38.8% between any two tools. While the results were more consistent among the tools on viromes, the differences in results were still significant, with a maximum overlap of 60.65%. Discussion: Importantly, the benchmark datasets developed in this study are publicly available and reusable to enable the future comparability of new tools developed.

Disturbed gut virome with potent interferonogenic property in systemic lupus erythematosus

Accumulating evidence suggests an essential role of disturbed gut microbiota in the etiopathogenesis of systemic lupus erythematosus (SLE), but it remains unclear as to gut virome. In this study, fecal virus-like particles (VLPs) isolated from 76 non-treated SLE patients and 75 healthy controls were subjected to gut virome profiling. The proportion of bacteriophages was significantly elevated in the SLE gut, and the altered viral taxa were correlated with clinical parameters. Gut virome and bacteriome were closely associated with each other in SLE patients. The combination of gut viral and bacterial markers displayed better performance in distinguishing SLE patients from healthy controls. Further, VLPs from non-treated SLE patients promoted interferon-α production in an epithelial cell line and human immune cells. Intriguingly, the interferon-stimulatory capacity diminished in VLPs from post-treated SLE patients. Our findings may shed novel insights into SLE pathogenesis. Further in-depth understanding of gut virome might help develop future biomarkers and therapeutics for SLE patients.

Alterations of the gut virome in patients with systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease that has been linked to the dysbiosis of the gut microbiome and virome. However, the potential characterization of the gut virome in SLE patients needs to be explored more extensively. Herein, we analyzed the gut viral community of 16 SLE patients and 31 healthy controls using both bulk and virus-like particle (VLP)-based metagenomic sequencing of their fecal samples. A total of 15,999 non-redundant viral operational taxonomic units (vOTUs) were identified from the metagenomic assembled contigs and used for gut virome profiling. SLE patients exhibited a significant decrease in gut viral diversity in the bulk metagenome dataset, but this change was not significant in the VLP metagenome dataset. Also, considerable alterations of the overall gut virome composition and remarkable changes in the viral family compositions were observed in SLE patients compared with healthy controls, as observed in both two technologies. We identified 408 vOTUs (177 SLE-enriched and 231 control-enriched) with significantly different relative abundances between patients and controls in the bulk virome, and 18 vOTUs (17 SLE-enriched in 1 control-enriched) in the VLP virome. The SLE-enriched vOTUs included numerous Siphoviridae, Microviridae, and crAss-like viruses and were frequently predicted to infect Bacteroides, Parabacteroides, and Ruminococcus_E, while the control-enriched contained numerous members of Siphoviridae and Myoviridae and were predicted to infect Prevotella and Lachnospirales_CAG-274. We explored the correlations between gut viruses and bacteria and found that some Lachnospirales_CAG-274 and Hungatella_A phages may play key roles in the virus-bacterium network. Furthermore, we explored the gut viral signatures for disease discrimination and achieved an area under the receiver operator characteristic curve (AUC) of above 0.95, suggesting the potential of the gut virome in the prediction of SLE. Our findings demonstrated the alterations in viral diversity and taxonomic composition of the gut virome of SLE patients. Further research into the etiology of SLE and the gut viral community will open up new avenues for treating and preventing SLE and other autoimmune diseases.

Gut bacteriome, mycobiome and virome alterations in rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic destructive autoimmune disease of the joints which causes significant pain, functional disability, and mortality. Although aberrant immune cell activation induced by the imbalance between T helper Th1/Th17 and Treg cells is implicated in the RA development, its etiopathogenesis remains unclear. The presence of mucosal inflammation and systemic IgA-isotype-autoantibodies (anti-citrullinated peptide antibodies and rheumatoid factor) in pre-clinical RA supports the mucosal origin hypothesis involving altered microbiota in disease development. The gut microbiota comprises diverse bacteria, fungal and viral components, which are critical in developing host immunity. Alterations in microbial abundance are known to exacerbate or attenuate immune responses in the gut microenvironment subsequently affecting the joints. Further, these changes can provide biomarkers for disease activity and outcome in RA. Most of the research till date has been focused on describing gut bacterial components in RA. Studies on gut mycobiome and virome components in RA are relatively new and burgeoning field. Given the paucity of mycobiome or virome specific studies in RA, this review, discusses the recent findings on alterations in gut bacterial, fungal, and viral components as well as their role in regulating the spectrum of immune-pathogenic events occurring in RA which might be explored in future as a potential therapeutic target. Further, we provide an overview on inter-kingdom interactions between bacteria, fungi, and viruses in RA. The current understanding on gut microbiota modulation for managing RA is also summarised.

The Emerging Role of the Gut Virome in Health and Inflammatory Bowel Disease: Challenges, Covariates and a Viral Imbalance.

Virome research is a rapidly growing area in the microbiome field that is increasingly associated with human diseases, such as inflammatory bowel disease (IBD). Although substantial progress has been made, major methodological challenges limit our understanding of the virota. In this review, we describe challenges that must be considered to accurately report the virome composition and the current knowledge on the virome in health and IBD. First, the description of the virome shows strong methodological biases related to wetlab (e.g., VLP enrichment) and bioinformatics approaches (viral identification and classification). Second, IBD patients show consistent viral imbalances characterized by a high relative abundance of phages belonging to the Caudovirales and a low relative abundance of phages belonging to the Microviridae. Simultaneously, a sporadic contraction of CrAss-like phages and a potential expansion of the lysogenic potential of the intestinal virome are observed. Finally, despite numerous studies that have conducted diversity analysis, it is difficult to draw firm conclusions due to methodological biases. Overall, we present the many methodological and environmental factors that influence the virome, its current consensus in health and IBD, and a contributing hypothesis called the "positive inflammatory feedback loop" that may play a role in the pathophysiology of IBD.