ABSTRACT Rapid progress in sequencing technology has made it possible to study the genome and transcriptional maps of single cells. However, to fully grasp the intricacies of multicellular organisms, methods that enable high‐throughput measurements while retaining spatial information about the tissue context or subcellular localization of the analyzed nucleic acids are essential. Over the past few years, as transcriptome research has advanced, the limitations of traditional transcriptomic approaches have become increasingly evident. In response, innovative sequencing techniques, such as spatial transcriptome sequencing, have emerged to better accommodate diverse research contexts. This review offers a comprehensive examination of the evolution and limitations of spatial transcriptomics. We summarize its applications in livestock and poultry research and explore its potential future developments. By providing insights into the current state and future directions of spatial transcriptomics, this review highlights its importance in advancing our understanding of complex biological systems.

This study investigates the interplay between host genetics and gut microbiota in regulating fat deposition and reproductive outcomes in sows. Integrating multiomics data from 348 Yorkshire sows-including whole-genome sequencing, 16S rRNA sequencing, and metabolomic profiling-we identified 37 microbial taxa significantly associated with thick backfat (TBP) through microbiome-wide association studies (MWAS). Microbial genome-wide association analysis (mGWAS) revealed heritable genera, including Ezakiella and Corynebacterium, and implicated host genes such as MED17 and VSTM5 in microbial modulation. Concurrently, genome-wide association studies (GWAS) identified MC4R and MEDAG as candidate genes directly influencing TBP. Metabolomic analysis highlighted acetate, a short-chain fatty acid, as a key mediator in host adipogenesis. Functional validation in experimental models demonstrated that exogenous acetate supplementation alters lipid metabolism and enhances reproductive performance. These findings elucidate a mechanistic axis wherein host genetics shape gut microbial composition, which in turn modulates fat metabolism via acetate production, ultimately affecting pregnancy outcomes. This integrated approach provides insights into host-microbe co-regulation of metabolic traits and highlights potential targets for improving reproductive efficiency in livestock.

The growth and development of skeletal muscle are closely associated with the proliferation, differentiation, and fusion of myoblasts. MicroRNAs (miRNAs) are a class of non-coding RNAs approximately 22 nucleotides in length that regulate skeletal muscle formation. miR-205-5p has been implicated in various physiological processes and diseases; however, its role in muscle development remains unclear. Therefore, this study used C2C12 myoblasts as a model to investigate the regulatory effects of miR-205-5p on myoblast proliferation, differentiation, and muscle atrophy, and to preliminarily elucidate its role in maintaining skeletal muscle homeostasis. The results showed that overexpression of miR-205-5p significantly inhibited the proliferation of C2C12 myoblasts while promoting their differentiation (P0.05), whereas interference with miR-205-5p yielded opposite results. In both the muscle atrophy model established after 8 days of induced differentiation and the dexamethasone-induced muscle atrophy model, overexpression of miR-205-5p alleviated the occurrence of muscle atrophy, whereas suppression of miR-205-5p expression exacerbated muscle atrophy (P0.05). In conclusion, miR-205-5p contributes to the maintenance of skeletal muscle homeostasis during muscle development by inhibiting proliferation, promoting differentiation, and attenuating muscle atrophy. This study provides a new molecular basis for elucidating the regulatory mechanisms of microRNAs in muscle development and for muscle atrophy intervention research.

The improvement of forage quality through proper fermentation is well documented. Grape pomace (GP), an economically valuable by-product, remains underutilized despite its potential benefits. This study explores the effects of fermented GP (FGP) supplementation on weaned piglets, focusing on immune and antioxidant capacity, intestinal flora homeostasis, growth performance, and diarrhea reduction. We first analyzed and compared the nutritional composition of GP before and after fermentation. Subsequently, a diet supplemented with 5

Background Understanding the genetic basis of hybridization trait between Chinese indigenous pig and their commercial relatives may contribute to germplasm innovation. The Qinchuan black (QCB) pig is a new composite breed under development generated from Chinese indigenous pig breeds (i.e., Guanzhong black pig, GZB) and intensive pig breeds (i.e., Yorkshire, also called Large white pig, LW). Investigated gene functions inherited from parent help us to understand the genetic basis of excellent phenotype similar with their parents, in which parents correspond to grandparents for hybrid population in our study. Result Here, we performed population structure and admixture analyses to elucidate the genetic position of QCB among Asian and European pig breeds. Ancestral track analysis revealed the LW contributed more genomic components in QCB genome, particularly on chromosome 7. This asymmetric inheritance may be influenced by linkage disequilibrium blocks. The offspring genome exhibited high genetic diversity, as did its maternal genome (GZB). Both genomes were characterized by a reduced number and shorter length of Run of Homozygosity (ROH). Finally, we used HKA test to investigated key gene in QCB that were inherited from its parents and that regulate important economic traits. Our results identified a significant association between an NPC1L1 haplotype inherited from the LW genome and body weight in four-month-old QCB. Conclusion Our research provides a novel strategy for utilizing previously identified functional loci or genes to guide the breeding of domestic pig for improvement economic traits in crossbreeding systems.

The domestic pig is a key agricultural species and biomedical model, yet its reference genome has remained incomplete. Using PacBio HiFi, Oxford Nanopore reads, and Hi-C sequencing, we assembled the first telomere-to-telomere (T2T), gap-free pig genome (T2T-Sscrofa). Importantly, this assembly derives from the same individual pig that provided the original reference genome, thereby completing the long-standing foundation of pig genomics. This assembly resolves 274.8 Mb of previously unassembled sequence, including centromeres, segmental duplications, and ribosomal DNA arrays, and identifies 255 new protein-coding genes. Comparative analyses reveal a Robertsonian translocation in the western European wild boar and extensive structural variation across global pig populations. T2T-Sscrofa provides a comprehensive genomic foundation for agricultural, evolutionary, and biomedical research. ### Competing Interest Statement The authors have declared no competing interest.

BACKGROUND:Traditional genomic analysis relies on a single reference genome, which struggles to effectively characterize the genetic diversity among populations. This is due to the substantial genetic differences between the genome of the studied species and the reference genome, potentially introducing reference bias. RESULTS:In this study, we focused on Guanzhong Black pigs (GZB), Danish Large White pigs (DLW), and their hybrid offspring, Qinchuan Black pigs (QCB). We provided two high-quality parental genomes at the chromosomal level and constructed a parental genomic reference panel to detect SNPs (single nucleotide polymorphisms), INDELs (insertions and deletions), and SVs (structural variations). Compared with the single-reference method, the integrated parental genomic strategy identified 5.48% more SNPs and 67.84% more INDELs. The uniformity of variant distribution and genome functional annotation remained consistent before and after integration, while the ratio of non-reference/non-reference genotypes was also improved. In population genetic structure analysis, principal component analysis (PCA) of the three variant types (SNPs, INDELs, and SVs) exhibited good clustering effects, and ADMIXTURE analysis demonstrated consistent stratification. Selection signal analysis based on the integrated parental genomic strategy successfully identified more differentiated windows and positively selected genes. By leveraging multiple variant types and employing two selection signal methods, we jointly identified several novel intramuscular fat candidate genes (MSMO1, SMC6, CCDC158, KIT, CCNC, etc.), which could not be identified by the single-reference method alone. Functional validation of the gene MSMO1 revealed its role in promoting intramuscular adipocyte proliferation and inhibiting adipogenic differentiation. CONCLUSIONS:This study is the first to construct a parental genomic reference panel specifically for pig hybrid populations, which significantly reduces reference bias and exhibits superior performance in downstream analyses. This strategy offers new possibilities for genomic selection breeding of livestock and establishes a methodological foundation for precisely dissecting complex traits in hybrid populations.

The number of mummies (MUM) in pigs is a major factor affecting sow reproductive performance. Reducing the incidence of MUM can effectively improve sow utilization efficiency. However, the complex mechanisms by which the host genome, gut microbiome, and metabolome interact to influence sow MUM remain unclear. Based on the current research landscape, this study systematically reveals the regulatory mechanisms of the host genome-gut microbiome-metabolome interaction network on sow MUM. By conducting a multi-omics analysis on the intestinal contents of Yorkshire sows during late gestation across different parities, we constructed a dynamic atlas of the gut microbiota and identified 385 core microbial taxa. Through multi-model MWAS and meta-analysis, we screened six key microbial taxa significantly associated with MUM, including Bacteroidales_RF16_group, Prevotellaceae_Ga6A1_group, Comamonas, Paraprevotella, Dorea, and Gallicola. An mGWAS analysis further identified Bacteroidales_RF16_group as regulated by host genetics, as well as candidate genes such as EGF, ENPEP, and CASP6, and important SNP loci such as rs345237235 and rs3475666995. The study found that the abundance of Proteobacteria in the sow's gut increased progressively from the first parity, providing a theoretical basis for pathogen suppression mechanisms. By integrating fecal metabolomics data, we constructed a four-dimensional regulatory network of host gene-gut microbiota-metabolite-host phenotype. This study innovatively combines quantitative genetics with multi-omics approaches, not only providing a theoretical foundation for understanding host-microbiota interaction mechanisms but also offering critical scientific guidance for reducing sow MUM incidence and improving reproductive efficiency.

The genetic basis of the phenotypic diversity of pigs is regulated by variants across the genome, especially the trait of early puberty, which is a crucial trait for enhancing the reproductive ability of pigs and the economy of the pig industry. However, the genetic basis of the early puberty trait in pigs remains largely unknown. Here, we report a comprehensive genomic variation map for pigs based on the resequencing of 493 accessions representing 59 different pig breeds or populations, which included 5,211,469 single-nucleotide polymorphisms (SNPs) and 487,725 small insertion/deletion structure variants (InDels). This sets included 45,640 high-quality structural variants (SVs). Our results suggested that Hanjiang black (HJB) pigs cluster with Jianghai-type pigs at the genetic level and that the genome characteristics of some HJB individuals exhibit a certain degree of European pig features. Using introgression and signature selection analysis, we identified several candidate genes associated with bone development and early puberty traits, such as TBX5, PAPPA2, IGFBP3, and MKRN3. Additionally, the GWAS and differential expression analysis results suggested that the PAPPA2 gene is associated with early puberty in pigs. This study revealed that past introgression events could impact the agronomical traits of pigs and contribute raw material of genetics and breeding in pig. Moreover, our results suggest that the PAPPA2 gene is a candidate gene associated with early sexual maturity in pigs and the genomic analysis provided important reference value for studying economic traits for pigs.

BACKGROUND:Pigs are not only a key source of animal protein worldwide, but also serve as important models in biological research. With the rapid development of short- and long-read sequencing technologies, genetic studies in pigs have advanced considerably. Although extensive research has been conducted on single-nucleotide polymorphisms (SNPs) and small insertions/deletions (indels), which has provided important insights into pig domestication, evolution, and trait formation, structural variants (SVs) remain underexplored due to technical limitations in sequencing resolution, challenges in variant detection, and insufficient population-scale sampling. RESULTS:In this study, we constructed the Pig Structural Variant Reference Panel (PSVRP) by integrating 21 long-read and 1,193 short-read whole-genome resequencing datasets from globally diverse pig populations. A total of 319,058 high-confidence SVs were identified, comprising 196,620 insertions and 122,438 deletions. Phylogenetic and ADMIXTURE analyses revealed clear divergence between Asian and European pigs, consistent with results derived from SNPs and indels data. Selection scans highlighted candidate genes associated with key traits, such as EPAS1 and NOVA1 for high-altitude adaptation, and PLAG1 and MIB1 for body size regulation. CONCLUSIONS:The PSVRP provides a high-resolution, population-scale pig SVs genotyping resource. This comprehensive panel deepens our understanding of genetic variation, facilitates the discovery of functional variants underlying adaptive and economic traits, and offers new insights for precision pig breeding.

BACKGROUND:Growth traits are economically important traits in pig breeding. However, the genetic mechanism of growth traits is still unclear. Qinchuan Black (QCB) pigs are crossbred and produced by hybridizing Guanzhong Black (GZB) pigs and Large White (LW) pigs, its characteristics include fast growth and excellent meat quality. In this study, whole genome and transcriptome analyses revealed the candidate genes associated with growth traits in QCB pigs based on imputed low-coverage whole-genome resequencing data. RESULTS:In total, we used 197 low-depth whole-genome resequencing data with an average depth of 3.5X, and then the data were imputed to resequencing data using SWIM reference panel, the imputation accuracy parameters, allele frequency r2 and concordance rate were 0.86 and 95.83%, respectively. We used two methods to investigate the candidate genes affecting the growth traits of QCB pigs, a total of 371 PSGs were identified, which related to muscle tissue development, tissue development and system development. A total of 30,489,782 SNPs were retained. A GWAS of ten growth traits by using fixed and random model circulating probability unification (FarmCPU) model, was performed in QCB pigs. We discovered seven genome wide significant SNPs and eight genome wide suggestive significant SNPs associated with body weight at 2 months (2-BW), body length at 2 months (2-BL), body height at 2 months (2-BH) and body height at 4 months (4-BH), and eighteen potential candidate genes were discovered. Transcriptomic data revealed that 18 differentially expression genes related to muscle and growth and development. Additionally, whole genome and transcriptome analyses found six genes (TENM3, CTNND2, RIMS1, PCDH7, ADGRL3 and CTNNA3) may affect the growth traits in Qinchuan Black pigs. CONCLUSION:Our study shows that more candidate genes associated with pig growth traits can be identified by whole genome and transcriptome analyses. We found that six genes may be new key candidate genes affecting pig growth traits. In conclusion, this study elucidated the molecular genetic mechanisms of growth traits and identified new molecular breeding targets, offering a robust scientific basis for advancing breeding strategies and genetic investigations within this breed.

The Bamei pigs (BM), an indigenous breed in Northwest China, is renowned for its superior meat quality. To uncover the genetic basis of its traits, we analyzed whole-genome sequencing data from 61 BM. Our results revealed that BM have a good genetic conservation status and distinct genomic divergence from Western breeds. We identified MALSU1 as a new candidate gene associated with intramuscular fat (IMF) by integrating selection signature analysis with public databases, such as PigGTEx, PigBiobank, and PigQTL. Overexpression and interference experiments of MALSU1 demonstrated that it regulates IMF by inhibiting the proliferation and promoting the differentiation of porcine intramuscular adipocyte primary cells. RNA-seq results further revealed that MALSU1 regulates IMF by inhibiting lipid metabolism and promoting lipid synthesis. Interestingly, a missense mutation (p.Arg10Leu) in the coding region of the MALSU1 gene was identified, which could promote the proliferation of intramuscular preadipocytes, suggesting an important role in IMF deposition.

Sophoridine (SRP) is a natural quinolizidine alkaloid found in many traditional Chinese herbs, though its effect on adipose tissue is unclear. We improved serum lipid levels by administering SRP by gavage in high-fat diet (HFD)-fed C57BL/6 mice. After 11 weeks, SRP supplementation significantly reduced body weight gain and improved glucose homeostasis, while reducing subcutaneous fat and liver weight. SRP also inhibited cell proliferation and differentiation of 3T3-L1 cells. Proteomics analysis revealed that SRP inhibits adipocyte differentiation by interacting with Src, thereby suppressing vascular endothelial growth factor receptor 2 (VEGFR2) expression and PI3K/AKT phosphorylation. This study provides an empirical basis for the treatment of obesity with small molecules.

BackgroundTeat number is one of the most important indicators to evaluate the lactation performance of sows, and increasing the teat number has become an important method to improve the economic efficiency of farms. Therefore, it is particularly important to deeply analyze the genetic mechanism of teat number traits in pigs. In this study, we detected Single Nucleotide Ploymorphism (SNP), Insertion-Deletion (InDel) and Structural variant (SV) by high-coverage whole-genome resequencing data, and selected teat number at birth and functional teat number as two types of teat number traits for genome-wide association study (GWAS) to reveal candidate genes associated with pig teat number traits.ResultsIn this study, we used whole genome resequencing data from 560 Yorkshire sows to detect SNPs, InDels and SVs, and performed GWAS for the traits of born teat number and functional teat number, and detected a total of 85 significant variants and screened 214 candidate genes, including HEG1, XYLT1, SULF1, MUC13, VRTN, RAP1A and NPVF. Among them, HEG1 and XYLT1 were the new candidate genes in this study. The co-screening and population validation of multiple traits suggested that HEG1 may have a critical effect on the born teat number.ConclusionOur study shows that more candidate genes associated with pig teat number traits can be identified by GWAS with different variant types. Through large population validation, we found that HEG1 may be a new key candidate gene affecting pig teat number traits. In conclusion, the results of this study provide new information for exploring the genetic mechanisms affecting pig teat number traits and genetic improvement of pigs.

Genomic prediction has been widely applied to the pig industry and has greatly accelerated the progress of genetic improvement in pigs. With the development of sequencing technology and price reduction, more and more genotype imputation panels of pig have been investigated, providing an effective and economical method to further study the genetic variation of pig economic traits. In this study, the imputation from 80 k Single Nucleotide Polymorphism chip data of 832 Large White pigs to whole-genome sequencing genotypes was performed by Swine Imputation Server, Pig Haplotypes Reference Panel (PHARP), Animal Genotype Imputation Database and 1k-pig-genomes four thousand-pig imputation panels. Then, linkage disequilibrium (LD) pruning and genome-wide association study (GWAS) preselected markers strategies were utilised to compare the genomic prediction accuracy of the different imputation data for reproductive traits, respectively. Our results showed that the PHARP panel exhibited the best genomic prediction accuracy among the four imputation panels. Meanwhile, the genomic prediction accuracy of the imputation data can be further improved by utilising the LD pruning and GWAS preselected marker strategies. In conclusion, our study provides insights into imputation data for pig genetic breeding.

The development of a comprehensive pig graph pangenome assembly encompassing 27 genomes represents the most extensive collection of pig genomic data to date. Analysis of this pangenome reveals the critical role of structural variations in driving adaptation and defining breed-specific traits. Notably, the study identifies BTF3 as a key candidate gene governing intramuscular fat deposition and meat quality in pigs. These findings underscore the power of pangenome approaches in uncovering novel genomic features underlying economically important agricultural traits. Collectively, these results demonstrate the value of leveraging large-scale, multi-genome analyses for advancing our understanding of livestock genomes and accelerating genetic improvement.

The quantity and distribution of fat deposits are crucial factors that impact the quality of pork. Recent studies have indicated that the utilization of natural ingredients plays a significant role in decreasing subcutaneous and visceral fat deposits, as well as enhancing intramuscular fat. Moreover, natural products possess several advantages including being environmentally friendly, safe, free of additives, and leaving no residue. Phenolic compounds derived from fruits, vegetables and herbs constitute of vital component of these natural ingredients. This article examines the influence of phenolic compounds on pig fat deposition, aiming to provide guidance on the utilization of such compounds to regulate fat deposition and enhance pork quality.

Organoids are in vitro 3D models that are generated using stem cells to study organ development and regeneration. Despite the extensive research on lung organoids, there is limited information on pig lung cell generation or development. Here, we identified five epithelial cell types along with their characteristic markers using scRNA-seq. Additionally, we found that NKX2.1 and FOXA2 acted as the crucial core transcription factors in porcine lung development. The presence of SOX9/SOX2 double-positive cells was identified as a key marker for lung progenitor cells. The Monocle algorithm was used to create a pseudo-temporal differentiation trajectory of epithelial cells, leading to the identification of signaling pathways related to porcine lung development. Moreover, we established the differentiation method from porcine pluripotent stem cells (pPSCs) to SOX17+FOXA2+ definitive endoderm (DE) and NKX2.1+FOXA2+CDX2- anterior foregut endoderm (AFE). The AFE is further differentiated into lung organoids while closely monitoring the differentiation process. We showed that NKX2.1 overexpression facilitated the induction of lung organoids and supported subsequent lung differentiation and maturation. This model offers valuable insights into studying the interaction patterns between cells and the signaling pathways during the development of the porcine lung. The core pathway of porcine lung cell composition and epithelial cell development was analyzed by scRNA-seq. The differentiation system of pEPSCs overexpressing NKX2.1 into definitive endoderm, anterior foregut endoderm, foregut endoderm spheroids, and porcine lung organoids was established, and the differentially expressed genes among them were identified by bulk RNA-seq.image

The feeding mode is an important factor affecting the reproductive performance of pigs. The composition and expression of the intestinal microbiota are closely related to the physiological and biochemical indicators of animals. Therefore, to explore the impact of different feeding patterns on the reproductive performance of pigs, this study collected reproductive performance data from 1607 Yorkshire pigs raised under different feeding patterns and conducted a fixed-effect variance analysis. Among them, 731 were in the artificial feeding (AM) group and 876 were in the feeding station feeding (SM) group. Additionally, 40 Yorkshire sows in the late gestation period were randomly selected from each feeding mode for intestinal microbiota analysis. The results of the analysis showed that, in the AM group, both the number of birth deformities (NBD) and the number of stillbirths (NSB) were significantly greater than they were in the SM group (p < 0.05). Additionally, the total number born (TNB) in the AM group was significantly lower than that in the SM group (p < 0.05). The results of the intestinal microbiota analysis revealed that at the phylum level, there were significant differences in nine bacterial taxa between the AM and SM groups (p < 0.05). At the genus level, the abundance of a variety of beneficial bacteria related to reproductive performance in the SM group was significantly greater than that in the AM group. Finally, fecal metabolomic analysis revealed that the contents of butyric acid, isovaleric acid, valeric acid, and isobutyric acid, which are associated with reproductive performance, in the feces of sows in the SM group were significantly higher than those in the AM group (p < 0.05). These results indicate that different feeding methods can affect the gut microbiota composition of Yorkshire pigs and further influence the reproductive performance of pigs through the gut microbiota–metabolic product pathway. The results of this study provide valuable insights for further exploring the relationships between feeding modes, intestinal microbial composition, and host phenotypes.

Abstract Background Feeding mode is an important factor affecting the reproductive performance of pigs. The composition and expression of the intestinal microbiota are closely related to the physiological and biochemical indicators of animals. Therefore, to investigate the effects of different feeding modes on the intestinal microbial community structure and reproductive performance of pigs, reproductive performance data were collected from 1607 Yorkshire pigs raised under different feeding modes for fixed-effect variance analysis. Among them, 731 were in the artificial feeding (AM) group, and 876 were in the feeding station feeding (SM) group. Additionally, 40 Yorkshire sows in the late gestation period were randomly selected from each feeding mode for intestinal microbiota analysis. Results The results of the analysis showed that, in the AM group, both the number of born deformity(NBD)and number of stillborn(NSB)were significantly greater than they were in the SM group (P < 0.05). Additionally, the TNB in the AM group was significantly lower than that in the SM group (P < 0.05). The results of the intestinal microbiota analysis revealed that at the phylum level, there were significant differences in 9 bacterial taxa between the AM and SM groups (P < 0.05). At the genus level, the abundance of a variety of beneficial bacteria related to reproductive performance in the SM group was significantly greater than that in the AM group. Differential pathway enrichment analysis revealed that the expression of the Cellular (P < 0.01) community-prokaryote pathway was significantly lower in the AM group than in the SM group (P < 0.05). However, the expression of the pathways glycan biosynthesis and metabolism, infectious diseases: viral, cardiovascular diseases, cancers: specific types, neurodegenerative diseases and metabolism of other amino acids was significantly greater in the AM group than in the SM group (P < 0.05). Conclusion These results indicate that different feeding modes can have an impact on the intestinal microbial composition of Yorkshire pigs and further affect the reproductive performance of pigs through the pathway of the intestinal microbiota. The results of this study provide valuable insights for further exploring the relationships between feeding modes, intestinal microbial composition, and host phenotypes.