Background: Antimicrobial resistance (AMR) has been regarded as a major threat to global health. Pigs are considered an important source of antimicrobial resistance genes (ARGs). However, there is still a lack of large-scale quantitative data on the distribution of ARGs in the pig production industry. The bacterial species integrated ARGs in the gut microbiome have not been clarified. Results: In the present study, we used deep metagenomic sequencing data of 451 samples from 425 pigs including wild boars, Tibetan pigs, and commercial or cross-bred experimental pigs under different rearing modes, to comprehensively survey the diversity and distribution of ARGs and detect the bacteria integrated in these ARGs. We identified a total of 1295 open reading frames (ORFs) recognized as antimicrobial resistance protein-coding genes. The ORFs were clustered into 349 unique types of ARGs, and these could be further classified into 69 drug resistance classes. Tetracycline resistance was most enriched in pig feces. Pigs raised on commercial farms had a significantly higher AMR level than pigs under semi-free ranging conditions or wild boars. We tracked the changes in the composition of ARGs at different growth stages and gut locations. There were 30 drug resistance classes showing significantly different abundances in pigs between 25 and 240 days of age. The richness of ARGs and 41 drug resistance classes were significantly different between cecum lumen and feces in pigs from commercial farms, but not in wild boars. We identified 24 bacterial species that existed in almost all tested samples (core bacteria) and were integrated 128 ARGs in their genomes. However, only nine ARGs of these 128 ARGs were core ARGs, suggesting that most of the ARGs in these bacterial species might be acquired rather than constitutive. We selected three subsets of ARGs as indicators for evaluating the pollution level of ARGs in samples with high accuracy (r = 0.73~0.89). Conclusions: This study provides a primary overview of ARG profiles in various farms under different rearing modes, and the data serve as a reference for optimizing the use of antimicrobials and evaluating the risk of pollution by ARGs in pig farms.

Prophages, viruses integrated into bacterial or archaeal genomes, can carry cargo that confers beneficial phenotypes to the host. The porcine gut microbiota constitutes a complex, dynamic, and interconnected ecosystem, yet the distribution of prophages and their unique functional characteristics within this microbial community remains poorly understood. In this study, we identified 10,742 prophage genomes through systematic screening of 7,524 prokaryotic genomes from porcine gut sources, representing both bacterial and archaeal lineages, with the distribution of integrated prophages exhibiting pronounced heterogeneity across host species. Additionally, 1.70% (183/10,742) of prophages exhibited a broad host range infectivity, while 5.07% (545/10,742) of integrated prophages enhanced prokaryotic adaptive immune capabilities by augmenting or directly providing host defense mechanisms. Notably, within tripartite phage-phage-host interactions network analysis, we observed that these prophages (n = 15) exhibit preferential acquisition of exogenous invasive phage sequences through CRISPR spacer integration mechanisms. Functional annotation revealed that prophage-encoded integrases and tail tube proteins may be critical determinants of phage host specificity. In addition, key auxiliary metabolic genes are encoded in the prophage of the pig intestinal tract, such as those promoting the synthesis of host microbiota-derived vitamin B12, encoded antibiotic resistance genes, and virulence factors that provide the host with a survival advantage. Furthermore, comparative analysis with existing viral and phage sequences uncovered a substantial reservoir of high-quality novel prophage sequences. Our findings systematically investigated the diversity of prophages in the pig gut, further characterizing their host range, functional attributes, and interactions with both host bacteria and other phages, through large-scale analysis of porcine gut microbiota genomes. This work offers new insights into the ecological roles of prophages and provides valuable genomic resources for studying prophages in this ecosystem.

Phages are ubiquitous and diverse, playing a key role in maintaining microbial ecosystem balance. However, their diversity, potential applications, and their interactions with hosts and other phages remain largely unexplored. To address this, we collected 59,652,008 putative viral genomes from our laboratory, 45 public viral datasets, and an integrated public viral genome database (IGN), covering seven habitats. We obtained 741,692 phage genomes with completeness ≥50% (PGD50), and most (93.83%, 695,938/741,692) of these phage genomes were classified into the Caudoviricetes class. We found that 158,522 species-level viral clusters that contained 28.96% (214,814/741,692) phage genomes without any known phage genomes in the IGN, indicating substantial novelty. Global phylogenetic trees for five iterations based on complete phage genomes significantly expanded the known diversity of the virosphere. Genome analysis revealed phage potential divergence with habitat types and highlighted the utilization of alternative genetic codes. Furthermore, 3D structural similarity searches demonstrated significant potential for annotating previously uncharacterized viral proteins. Analysis of CRISPR spacer inferred potential hosts of phages and competitive networks among phages, highlighting virulent phages as promising candidates for phage therapy against pathogenic bacteria. Intriguingly, diverse CRISPR-Cas systems were detected within phage genomes themselves, suggesting their enormous potential as novel gene editing tools. Collectively, this study provides a comprehensive phage genome resource, foundational for future research into phage–host and phage–phage interactions, phage therapy development, and the mining of next-generation genetic tools.

Gut microbiome research has historically focused on bacterial communities. In contrast, the roles of viruses, especially jumbo phages, remain poorly understood. Jumbo phages are of major interest because their large genomes encode unique functions that can influence host metabolism and ecosystem dynamics. This study bridges this gap by identifying 1,545 jumbo phage genomes from 450 pig gut metagenomes. Using CRISPR spacer analysis, we predicted archaeal or bacterial hosts and reconstructed competitive phage networks within this ecosystem. Phylogenetic divergence combined with orthologous protein comparisons supported establishing 14 novel jumbo phage families. Functionally, 10 of these novel families encode auxiliary metabolic genes (AMGs) that enhance host metabolism alongside anti-defense systems including DNA methyltransferases, HNH endonucleases, and glycosyltransferases. Ecological interactions were further elucidated through co-abundance networks ( n = 857 pairs) and CRISPR spacer matching ( n = 425 pairs), revealing relationships between novel jumbo phages and other jumbo phages. Collectively, this work expands genomic resources for pig gut viromes and delivers new insights into jumbo phages’ functional capabilities, host associations, and global prevalence.

Summary: We have generated a large heterogenous stock population by intercrossing eight divergent pig breeds for multiple generations. We have analyzed the composition of the intestinal microbiota at different ages and anatomical locations in > 1,000 6 ^th - and 7 ^th - generation animals. We show that, under conditions of exacerbated genetic yet controlled environmental variability, microbiota composition and abundance of specific taxa (including Christensenellaceae ) are heritable in this monogastric omnivore. We fine-map a QTL with major effect on the abundance of Erysipelotrichaceae to chromosome 1q and show that it is caused by a common 2.3-Kb deletion inactivating the ABO acetyl-galactosaminyl-transferase gene. We show that this deletion is a trans-species polymorphism that is ≥3.5 million years old and under balancing selection. We demonstrate that it acts by decreasing the concentrations of N-acetyl-galactosamine in the cecum thereby reducing the abundance of Erysipelotrichaceae strains that have the capacity to import and catabolize N-acetyl-galactosamine.

Background: Jumbo phages are phages with comparatively large genome sizes. Jumbo phages have been identified in various microbial communities. However, their diversity, genome structure, potential function, and their interactions with hosts and other phages are largely unknown due to insufficient genomic data. Results: We collected 59,652,008 putative viral genomes from seven habitats by using 38 public metagenome datasets, an integrated public viral genome database (IGN), and pig gut viral genome databases. We obtained 10,754 jumbo phage genomes with sizes ranging from 200 to 831 kb. Most (94.64%) of these jumbo phage genomes were classified into Caudoviricetes, and the results have expanded the known diversity of Caudoviricetes. We found 2,389 species-like operational genome clusters that contained 3,727 (34.69%) genomes without any known viral genomes in the IGN, suggesting potential novel species-like genomes. Genome analysis suggested the potential coevolution of jumbo phages with habitat types and highlighted the utilization of alternative genetic codes and their corresponding suppressor tRNAs for recoding stop codons. CRISPR spacer analysis revealed potential bacterial or archaeal hosts of jumbo phages and uncovered competitive networks among jumbo phages. Habitat type had an important effect on the variation in phage auxiliary metabolic genes. Conclusions: This study provides an important resource and new knowledge for future studies on the interaction between jumbo phages and their bacterial or archaeal hosts.