Metagenome of gut microbes has been implicated in metabolism, immunity, and health maintenance of its host. However, in most of previous studies, the microbiota was sampled from feces instead of gastrointestinal (GI) tract. In this study, we compared the microbial populations from feces at four different developmental stages and contents of four intestinal segments at maturity to examine the dynamic shift of microbiota in pigs and investigated whether adult porcine fecal samples could be used to represent samples of the GI tract. Analysis results revealed that the ratio of Firmicutes to Bacteroidetes from the feces of the older pigs (2-, 3-, 6- month) were 10 times higher compared to those from piglets (1-month). As the pigs matured, so did it seem that the composition of microbiome became more stable in feces. In adult pigs, there were significant differences in microbial profiles between the contents of the small intestine and large intestine. The dominant genera in the small intestine belonged to aerobe or facultative anaerobe categories, whereas the main genera in the large intestine were all anaerobes. Compared to the GI tract, the composition of microbiome was quite different in feces. The microbial profile in large intestine was more similar to feces than those in the small intestine, with the similarity of 0.75 and 0.38 on average, respectively. Microbial functions, predicted by metagenome profiles, showed the enrichment associated with metabolism pathway and metabolic disease in large intestine and feces while higher abundance of infectious disease, immune function disease, and cancer in small intestine. Fecal microbes also showed enriched function in metabolic pathways compared to microbes from pooled gut contents. Our study extended the understanding of dynamic shift of gut microbes during pig growth and also characterized the profiles of bacterial communities across GI tracts of mature pigs.

Significance: Uncoupling protein 1 (UCP1) is responsible for brown adipose tissue-mediated thermogenesis and plays a critical role in protecting against cold and regulating energy homeostasis. Modern pigs lack functional UCP1, which makes them susceptible to cold and prone to fat deposition and results in neonatal mortality and decreased production efficiency. In the current study, a CRISPR/Cas9-mediated homologous recombination-independent approach was established, and mouse adiponectin-UCP1 was efficiently inserted into the porcine endogenous UCP1 locus. The resultant UCP1 KI pigs showed an improved ability to maintain body temperature, decreased fat deposition, and increased carcass lean percentage. UCP1 KI pigs are a potentially valuable resource for the pig industry that can improve pig welfare and reduce economic losses.

Pigs lack functional uncoupling protein 1 (UCP1) making them susceptible to cold. Nevertheless, several pig breeds are known to be cold resistant. The molecular mechanism(s) enabling such adaptation are currently unknown. Here, we show that this resistance is not dependent on shivering, but rather depends on UCP3 and white adipose tissue (WAT) browning. In two cold-resistant breeds (Tibetan and Min), but not a cold-sensitive breed (Bama), WAT browning was induced after cold exposure. Beige adipocytes from Tibetan pigs exhibited greater oxidative capacity than those from Bama pigs. Notably, UCP3 expression was significantly increased only in cold-resistant breeds, and knockdown of UCP3 expression in Tibetan adipocytes phenocopied Bama adipocytes in culture. Moreover, the eight dominant pig breeds found across China can be classified into cold-sensitive and cold-resistant breeds based on the UCP3 cDNA sequence. This study indicates that UCP3 has contributed to the evolution of cold resistance in the pig and overturns the orthodoxy that UCP1 is the only thermogenic uncoupling protein.

Pig shows multiple superior characteristics in anatomy, physiology, and genome that have made this species to be more suitable models for human diseases, especially for neurodegenerative diseases, because they have similar cerebral convolutions compared with human neocortex. Recently, CRISPR/Cas9 system shows enormous potential for engineering the pig genome. In this study, we expect to generate human Parkinson’s disease pig model using CRISPR/Cas9 system by simultaneously targeting three distinct genomic loci, parkin/DJ-1/PINK1, in Bama miniature pigs. By co-injection of Cas9 mRNA and multiplexing single guide RNAs (sgRNAs) targeting parkin, DJ-1, and PINK1 genes, respectively, into in vivo derived pronuclear embryos, we simultaneously targeted three distinct genomic loci. The gene modified piglets remain healthy and display normal behavior at the age of 10 months. In addition, despite the high number of sgRNAs were employed in the present study, our trio-based whole-genome sequencing analysis suggested that the incidence of off-target events is low. Our results demonstrate that the simplicity, efficiency, and power of the CRISPR/Cas9 system to allow for the modification of multiple genes in pigs and yield results of high medical value.

Anatomic and physiological similarities to the human make swine an excellent large animal model for human health and disease. Cloning from a modified somatic cell, which can be determined in cells prior to making the animal, is the only method available for the production of targeted modifications in swine. Since some strains of swine are similar in size to humans, technologies that have been developed for swine can be readily adapted to humans and vice versa. Here the importance of swine as a biomedical model, current technologies to produce genetically enhanced swine, current biomedical models, and how the completion of the swine genome will promote swine as a biomedical model are discussed. The completion of the swine genome will enhance the continued use and development of swine as models of human health, syndromes and conditions.

Large animals (non-human primates, livestock and dogs) are playing important roles in biomedical research, and large livestock animals serve as important sources of meat and milk. The recently developed programmable DNA nucleases have revolutionized the generation of gene-modified large animals that are used for biological and biomedical research. In this review, we briefly introduce the recent advances in nuclease-meditated gene editing tools, and we outline these editing tools’ applications in human disease modeling, regenerative medicine and agriculture. Additionally, we provide perspectives regarding the challenges and prospects of the new genome editing technology.