Integrated analysis of muscle transcriptome, miRNA, and proteome of Chinese indigenous breed Ningxiang pig in three developmental stages.

The Ningxiang pig, a distinguished local breed in China, is recognized for its good meat quality traits. This study examines the proteomics of Ningxiang pigs at three developmental stages and delves into the upstream transcriptomics of these proteomics. Such an analysis facilitates a deeper understanding of the molecular interplay between proteins and transcriptomes in the Ningxiang pig muscle, influencing muscle growth and development. In this research, we analyzed the muscles of Ningxiang pigs at three developmental stages: 30 days in weaned piglets, 90 days in nursery pigs, and 210 days in late fattening pigs. There a total of 16 differentially co-expressed miRNAs (ssc-miRNA-1, ssc-miRNA-378, ssc-miRNA-143, ssc-miRNA-30e, etc.), 74 differentially co-expressed mRNA (PLIN3, CPT2, IGF2 and HSP90AB1, etc.) have been identified in the three stages. 572 differentially abundant proteins (DAPs) (APOC3, NDUFA2, HSPD1, ATP5E, PDHA1, etc.) were readily identified by comparing different time periods. According to the KEGG enrich pathway results that DAPs most enriched in growth and development pathways, immune mechanism pathways and maintaining functions of physical. Through short time-series expression miner (STEM) association analysis, a total of 571 negative miRNA-mRNA interaction pairs and 2 negative miRNA-mRNA-protein (Chr05_11955-Pig.17268.1-ATP5F1B, ssc-miR-194a-3p-Pig.15802.1-ACY1) interaction pairs were found. Our study provides a theoretical basis on molecular mechanism for the study of IMF deposition, muscle growth and immunity in Ningxiang pig breed.

Genes influencing deposition of melanin in breast muscle of the Xuefeng black bone chicken based on bioinformatic analysis.

The Xuefeng black bone chicken (XFBC) represents an important poultry genetic resource. However, the darkness in breast muscle is heterogeneous. The molecular genetic mechanisms underlying melanogenesis of breast muscle in XFBC remains unclear. This study used RNA-seq to compare the difference in transcriptome between hyperpigmentation and hypopigmentation of breast muscle. Six cDNA libraries were constructed for hyperpigmentation and hypopigmentation groups in XFBC. We identified 395 differently expressed genes (DEGs) between hyperpigmentation and hypopigmentation group (P < 0.05, |log2FC|≥1). Gene ontology (GO) enrichment and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated several differentially enriched biological functions and pathways involved in melanogenesis of the breast muscle. Gene set enrichment analysis (GSEA) GO analysis identified two significant gene sets, including the pathways of pigment metabolic process and transmembrane receptor protein tyrosine kinase activity. GSEA-KEGG analysis identified the process of tyrosine metabolism and several genes related with melanogenesis in breast muscle of the XFBC. The protein-protein interaction network was constructed and eight genes were clustered in the module. We identified nine hub genes, including TYR, TYRP1, DCT, GPR143, MLANA, SLC24A5, GPNMB, MLPH, and EDNRB2. Taken together, the DEGs and hub genes identified in the study provide a solid basis for the study of the genetic regulatory mechanisms involved the melanogenesis in the breast muscle of the XFBC.

Effects of melatonin on intestinal function and bacterial compositions in sucking piglets.

Melatonin has been reported to affect intestinal function by targeting microbiome, morphological structure, barrier integrity and nutrient absorptive system. While the effect of melatonin on intestinal development in newborn infants is obscure, thus, this study firstly attempted to investigate the hypothesis that melatonin treatment improves intestinal development in sucking piglets. 14 healthy newborn piglets received 10 ml melatonin solution (1 mg/ml) or drinking water (n = 7) for 21 days. The results showed that oral administration of melatonin increased liver relative weight (p < 0.05) but failed to affect growth performance in sucking piglets (p > 0.05). Immunostaining jejunal samples from melatonin group showed high expressions of nnos and claudin1, indicating that melatonin improved intestinal neural development and barrier integrity. Also, melatonin promoted intestinal absorptive function evidenced by the increased serum proline concentration in melatonin-treated piglets compared with the control (p < 0.05). Gut microbiota compositions were tested by 16S rDNA sequencing and the results showed that melatonin increased the relative abundance of Actinobacteria compared with the control (p < 0.05) at the phylum level. However, Selenomonadales was markedly reduced compared with the control at the order level (p < 0.05). Gut and faecal volatile fatty acids were tested to evaluate the microbiota metabolism, but no difference was noticed in volatile fatty acid concentrations (p > 0.05). Melatonin improved intestinal development by affecting neural development, barrier integrity, nutrient absorption and microbiota in sucking piglets.

Long-read assembly of the Chinese indigenous Ningxiang pig genome and identification of genetic variations in fat metabolism among different breeds.

Advances in long-read sequencing technology and genome assembly provide an opportunity to improve the pig genome and reveal the full range of structural variations (SVs) between local Chinese and European pigs. To date, little is known about the genomes of some unique Chinese indigenous breeds, such as the Ningxiang pig. Here, we report the sequencing and assembly of a highly contiguous Ningxiang pig genome (NX) via an integration of PacBio single-molecule real-time sequencing, Illumina next-generation sequencing, BioNano optical mapping and Hi-C (chromosome conformation capture) approaches. The assembled genome comprises 2.44 Gb with a contig N50 of 26.1 Mb and 418 contigs in total. These contigs are organized into 121 scaffolds with a scaffold N50 of 139.0 Mb. More than 99.1% of the assembled sequence could be localized to 19 pseudochromosomes and is annotated with 20,914 protein-coding genes and 34.04% repetitive sequences. Comparisons between the NX and European Duroc assemblies revealed many SVs in genes involved in the immune system, nervous system, lipid metabolism and environmental adaptation. The genetic variants include 47 Chinese domestic pig-specific SVs and the associated 74 genes may contribute to the differences in domestic traits compared to European pigs. Moreover, single nucleotide polymorphisms (SNPs) identified from whole genome resequencing data of 73 Chinese pigs, representing 17 geographically isolated breeds, showed their specific genetic variations, population structure and evolutionary patterns. Finally, we explore transcriptional regulation in the first intron of the MYL4 gene, as the genomic SV (281-bp deletion) in Ningxiang pig promotes its subcutaneous fat compared to European pig breeds. This work identifies a set of Asian-specific SVs and SNPs, which will be important resources for modern pig breeding and genetic conservation.

Modulation of intestinal morphology and microbiota by dietary Macleaya cordata extract supplementation in Xuefeng Black-boned Chicken.

Antibiotics are commonly overused to prevent livestock from diseases and to increase production performance. As potential substitutes of antibiotics, plant extracts have attracted the attention of researchers. It was known to all that addition of Macleaya cordata extract (MCE) to the food could advance immunity, intestinal health and animal performance. Thus, it was conducted to investigate the influence of MCE (0, 100, 150 and 200 mg/kg, with six replicate pens/treatment and 24 hens/pen) on intestinal morphology and microbial diversity in different intestinal segments in Xuefeng black-boned chicken in this study. The results showed that MCE supplement (100, 150 and 200 mg/kg) significantly diminished (P < 0.05) the crypt depth of the jejunum as compared to basal diet group. The 100 mg/kg group displayed a marked increase (P < 0.05), compared with 0 and 200 mg/kg group, in ileum microbial diversity as represented by the Shannon's index. In the cecum, treatment of MCE significantly decreased (P < 0.01) the Firmicutes, but Deferribacteres in 200 mg/kg MCE group were significantly raised (P < 0.05). In conclusion, we found that MCE improved intestinal morphology and reduced the crypt depth in jejunum. Together, addition of 200 mg/kg MCE modulated intestinal microbiota, increased beneficial bacteria such as Lactobacillus. Adding 100 mg/kg MCE to diet increased bacterial community diversity and relative abundance in jejunum and ileum, but had no effect on cecum microbial diversity.

Eugenol Alleviates Dextran Sulfate Sodium-Induced Colitis Independent of Intestinal Microbiota in Mice.

The present study investigated the effect of eugenol (EUG) on dextran sulfate sodium (DSS)-induced colitis and explored the underlying mechanisms. C57BL/6 mice were intragastrically administered normal saline or EUG (20 mg/kg body weight) for 17 days, and colitis was induced by using 3% DSS from day 7. The results showed that EUG increased the body weight and reduced the disease activity index score and colon pathological scores in DSS-treated mice (P < 0.05). Further, EUG preserved the proinflammatory cytokines (interleukin (IL)-6, -12, -21, and -23), lowered (P < 0.05) colonic malondialdehyde (MDA), uncoupling protein 2 (UCP2) expression and p65 phosphorylation, and activated (P < 0.05) colonic kelch-like ECH-associated protein 1 and nuclear factor (erythroid-derived 2)-like 2 expressions but did not affect the intestinal microbiota in DSS-treated mice. Furthermore, EUG ameliorated colitis in antibiotic-treated mice, while fecal microbiota transplantation from EUG preadministered mice failed to ameliorate colitis. In conclusion, EUG could alleviate colitis by attenuating colonic inflammation and oxidative stress independent of intestinal microbiota.

Chloroquine Improves Deoxynivalenol-Induced Inflammatory Response and Intestinal Mucosal Damage in Piglets.

We investigated the effects of rapamycin (RAPA) and chloroquine (CQ) in supporting growth performance and the intestinal mucosal barrier in response to deoxynivalenol (DON) in piglets. A total of 32 healthy weaned piglets (bodyweight 7.10 ± 0.58 kg) were divided into four groups and treated daily with RAPA (1 mg/kg BW), CQ (10 mg/kg BW), or a control volume of normal saline (two groups) until the end of the experiment. After feeding a basal diet for seven days, three groups were then switched to mildewed feed containing 1 mg kg/DON for a further seven days. In contrast to the control group, DON-treated piglets showed decreased average daily gain (ADG) and daily feed intake (ADFI), as well as negatively affected intestinal morphology as indicated by villus height, crypt depth, and tight junction protein expression. A group treated with RAPA and DON showed increased intestinal autophagy, aggravated inflammatory responses, and damage to the intestinal mucosa and permeability, leading to reduced growth performance. Meanwhile, a group treated with CQ and DON showed indices comparable to the non-DON control group, with alleviated inflammatory cytokines and healthy intestinal morphology and structure. They also showed better growth performance compared to DON treatment alone. These findings have important implications for mediating autophagy against DON in vivo, as well as the potential for CQ in improving growth performance and maintaining intestinal barrier integrity in weanling piglets.

Chloroquine Downregulation of Intestinal Autophagy to Alleviate Biological Stress in Early-Weaned Piglets.

Early weaning stress impairs the development of gastrointestinal barrier function, causing immune system dysfunctions, reduction in feed intake, and growth retardation. Autophagy was hypothesized to be a key underlying cellular process in these dysfunctions. We conjectured that rapamycin (RAPA) and chloroquine (CQ), as two autophagy-modifying agents, regulate the autophagy process and may produce deleterious or beneficial effects on intestinal health and growth. To explore the effect of autophagy on early weaning stress in piglets, 18 early-weaned piglets were assigned to three treatments (each treatment of six piglets) and treated with an equal volume of RAPA, CQ, or saline. The degree of autophagy and serum concentrations of immunoglobulins and cytokines, as well as intestinal morphology and tight junction protein expression, were evaluated. Compared with the control treatment, RAPA-treated piglets exhibited activated autophagy and had decreased final body weight (BW) and average daily gain (ADG) (p < 0.05), impaired intestinal morphology and tight junction function, and higher inflammatory responses. The CQ-treated piglets showed higher final BW, ADG, jejuna and ileal villus height, and lower autophagy and inflammation, compared with control piglets (p < 0.05). Throughout the experiment, CQ treatment was beneficial to alleviate early weaning stress and intestinal and immune system dysfunction.