Microbiomes are vast communities of microorganisms and viruses that populate all natural ecosystems. Viruses have been considered to be the most variable component of microbiomes, as supported by virome surveys and examples of high genomic mosaicism. However, recent evidence suggests that the human gut virome is remarkably stable compared with that of other environments. Here, we investigate the origin, evolution and epidemiology of crAssphage, a widespread human gut virus. Through a global collaboration, we obtained DNA sequences of crAssphage from more than one-third of the world’s countries and showed that the phylogeography of crAssphage is locally clustered within countries, cities and individuals. We also found fully colinear crAssphage-like genomes in both Old-World and New-World primates, suggesting that the association of crAssphage with primates may be millions of years old. Finally, by exploiting a large cohort of more than 1,000 individuals, we tested whether crAssphage is associated with bacterial taxonomic groups of the gut microbiome, diverse human health parameters and a wide range of dietary factors. We identified strong correlations with different clades of bacteria that are related to Bacteroidetes and weak associations with several diet categories, but no significant association with health or disease. We conclude that crAssphage is a benign cosmopolitan virus that may have coevolved with the human lineage and is an integral part of the normal human gut virome.

An important viromics challenge is associating bacteriophages to hosts. To address this, we developed adsorption sequencing (AdsorpSeq), a readily implementable method to measure phages that are preferentially adsorbed to specific host cell envelopes. AdsorpSeq thus captures the key initial infection cycle step. Phages are added to cell envelopes, adsorbed phages are isolated through gel electrophoresis, after which adsorbed phage DNA is sequenced and compared with the full virome. Here, we show that AdsorpSeq allows for separation of phages based on receptor-adsorbing capabilities. Next, we applied AdsorpSeq to identify phages in a wastewater virome that adsorb to cell envelopes of nine bacteria, including important pathogens. We detected 26 adsorbed phages including common and rare members of the virome, a minority being related to previously characterized phages. We conclude that AdsorpSeq is an effective new tool for rapid characterization of environmental phage adsorption, with a proof-of-principle application to Gram-negative host cell envelopes.

The human gut contains an expanse of largely unstudied bacteriophages. Among the most common are crAss-like phages, which were predicted to infect Bacteriodetes hosts. CrAssphage, the first crAss-like phage to be discovered, contains a protein encoding a Bacteroides-associated carbohydrate-binding often N-terminal (BACON) domain tandem repeat. Because protein domain tandem repeats are often hotspots of evolution, BACON domains may provide insight into the evolution of crAss-like phages. Here, we studied the biodiversity and evolution of BACON domains in bacteriophages by analysing over 2 million viral contigs. We found a high biodiversity of BACON in seven gut phage lineages, including five known crAss-like phage lineages and two novel gut phage lineages that are distantly related to crAss-like phages. In three BACON-containing phage lineages, we found that BACON domain tandem repeats were associated with phage tail proteins, suggestive of a possible role of these repeats in host binding. In contrast, individual BACON domains that did not occur in tandem were not found in the proximity of tail proteins. In two lineages, tail-associated BACON domain tandem repeats evolved largely through horizontal transfer of separate domains. In the third lineage that includes the prototypical crAssphage, the tandem repeats arose from several sequential domain duplications, resulting in a characteristic tandem array that is distinct from bacterial BACON domains. We conclude that phage tail-associated BACON domain tandem repeats have evolved in at least two independent cases in gut bacteriophages, including in the widespread gut phage crAssphage.

Several risk factors are associated with gallstone disease after bariatric surgery, but the underlying pathophysiological mechanisms of gallstone formation are unclear. We hypothesize that gallstone formation after bariatric surgery is induced by different pathways compared with gallstone formation in the general population, since postoperative formation occurs rapidly in patients who did not develop gallstones in preceding years. To identify both pathophysiological and potentially protective mechanisms against postoperative gallstone formation, we compared the preoperative fasting metabolome, fecal microbiome, and liver and adipose tissue transcriptome obtained before or during bariatric surgery of obese patients with and without postoperative gallstones. In total, 88 patients were selected from the BARIA longitudinal cohort study. Within this group, 32 patients had postoperative gallstones within 2 years. Gut microbiota metagenomic analyses showed group differences in abundance of 41 bacterial species, particularly abundance of Lactobacillaceae and Enterobacteriaceae in patients without gallstones. Subcutaneous adipose tissue transcriptomic analyses revealed four genes that were suppressed in gallstone patients compared with patients without gallstones. These baseline gene expression and gut microbiota composition differences might relate to protective mechanisms against gallstone formation after bariatric surgery. Moreover, baseline fasting blood samples of patients with postoperative gallstones showed increased levels of several bile acids. Overall, we revealed different genes and bacteria associated with gallstones than those previously reported in the general population, supporting the hypothesis that gallstone formation after bariatric surgery follows a different trajectory. Further research is necessary to confirm the involvement of the bile acids, adipose tissue activity, and microbial species observed here.