Integrated analysis strategy of genome-wide functional gene mining reveals DKK2 gene underlying meat quality in Shaziling synthesized pigs.

BACKGROUND: Shaziling pig is a well-known indigenous breed in China who has superior meat quality traits. However, the genetic mechanism and genomic evidence underlying meat quality characteristics of Shaziling pigs are still unclear. To explore and investigate the germplasm characteristics of Shaziling pigs, we totally analyzed 67 individual's whole genome sequencing data for the first time (20 Shaziling pigs [S], 20 Dabasha pigs [DBS], 11 Yorkshire pigs [Y], 10 Berkshire pigs [BKX], 5 Basha pigs [BS] and 1 Warthog). RESULTS: A total of 2,538,577 SNPs with high quality were detected and 9 candidate genes which was specifically selected in S and shared in S to DBS were precisely mined and screened using an integrated analysis strategy of identity-by-descent (IBD) and selective sweep. Of them, dickkopf WNT signaling pathway inhibitor 2 (DKK2), the antagonist of Wnt signaling pathway, was the most promising candidate gene which was not only identified an association of palmitic acid and palmitoleic acid quantitative trait locus in PigQTLdb, but also specifically selected in S compared to other 48 Chinese local pigs of 12 populations and 39 foreign pigs of 4 populations. Subsequently, a mutation at 12,726-bp of DKK2 intron 1 (g.114874954 A > C) was identified associated with intramuscular fat content using method of PCR-RFLP in 21 different pig populations. We observed DKK2 specifically expressed in adipose tissues. Overexpression of DKK2 decreased the content of triglyceride, fatty acid synthase and expression of relevant genes of adipogenic and Wnt signaling pathway, while interference of DKK2 got contrary effect during adipogenesis differentiation of porcine preadipocytes and 3T3-L1 cells. CONCLUSIONS: Our findings provide an analysis strategy for mining functional genes of important economic traits and provide fundamental data and molecular evidence for improving pig meat quality traits and molecular breeding.

Leucine improves the growth performance, carcass traits, and lipid nutritional quality of pork in Shaziling pigs.

Leucine is involved in promoting fatty acid oxidation and lipolysis, mediating lipid metabolism and energy homeostasis, thus it has been widely used in livestock production. However, the effects of leucine on fat deposition and nutrition in Shaziling pigs remain unclear. A total of 72 Shaziling pigs (150 days old, weight 35.00 ± 1.00 kg) were randomly divided into 2 groups and fed with basal diet (control group) or basal diet containing 1% leucine (leucine group) for 60 days. The results showed that leucine significantly increased the average daily feed intake but decreased the ratio of feed to gain (P < 0.05), increased the loin muscle area and serum glucose content (P < 0.05) of Shaziling pigs. Besides, leucine regulated the re-distribution of fatty acids from adipose tissue to muscle as it significantly increased the contents of C18:1n-9 and C22:6n-3 (DHA) in the longissimus thoracis while decreased the contents of C22:5n-3 (DPA), C20:5n-3 (EPA), and DHA in the adipose tissue of Shaziling pigs (P < 0.05). Lipidomic analysis showed that the contents of phosphatidylethanolamines (PEs), cardiolipins (CLs), and phosphatidylglycerols (PGs) in the longissimus thoracis and the contents of lysophosphatidylethanolamines (LPEs), ceramides (Cers), phosphatidylinositols (PIs) in adipose tissue of Shaziling pigs were decreased in leucine group (P < 0.05). Collectively, this study clarified that dietary addition of 1% leucine have a better effect on growth performance and the deposition of beneficial fatty acids in the muscle of Shaziling pigs, which is conductive to the production of high quality and healthy pork. In addition, leucine altered the lipid composition of muscle and fat in Shaziling pigs. The related results provide a theoretical basis and application guidance for regulating fat deposition in Shaziling pigs, which is important for the healthy breeding of Shaziling pigs.

Genome-Wide Association Analysis Identified Variants Associated with Body Measurement and Reproduction Traits in Shaziling Pigs.

With the increasing popularity of genomic sequencing, breeders pay more attention to identifying the crucial molecular markers and quantitative trait loci for improving the body size and reproduction traits that could affect the production efficiency of pig-breeding enterprises. Nevertheless, for the Shaziling pig, a well-known indigenous breed in China, the relationship between phenotypes and their corresponding genetic architecture remains largely unknown. Herein, in the Shaziling population, a total of 190 samples were genotyped using the Geneseek Porcine 50K SNP Chip, obtaining 41857 SNPs for further analysis. For phenotypes, two body measurement traits and four reproduction traits in the first parity from the 190 Shaziling sows were measured and recorded, respectively. Subsequently, a genome-wide association study (GWAS) between the SNPs and the six phenotypes was performed. The correlation between body size and reproduction phenotypes was not statistically significant. A total of 31 SNPs were found to be associated with body length (BL), chest circumference (CC), number of healthy births (NHB), and number of stillborns (NSB). Gene annotation for those candidate SNPs identified 18 functional genes, such as GLP1R, NFYA, NANOG, COX7A2, BMPR1B, FOXP1, SLC29A1, CNTNAP4, and KIT, which exert important roles in skeletal morphogenesis, chondrogenesis, obesity, and embryonic and fetal development. These findings are helpful to better understand the genetic mechanism for body size and reproduction phenotypes, while the phenotype-associated SNPs could be used as the molecular markers for the pig breeding programs.

Profiles of muscular amino acids, fatty acids, and metabolites in Shaziling pigs of different ages and relation to meat quality.

Pork meat is closely related to physicochemical alterations during growth and development, resulting in differences in nutritional value and meat flavor. This study aimed to evaluate the composition of amino acids, fatty acids, and metabolic profiles in the longissimus thoracis muscle (LM) of Shaziling pigs aged 30, 90, 150, 210, and 300 days. The results showed that the predominant fatty acids identified in the LM of Shaziling pigs were C16:0, C16:1, C18:0, C18:1n9c, and C18:2n6c. An opposite correlation was observed for C18:2n6c and n6/n3 polyunsaturated fatty acids (P<0.05). Alanine, aspartate, glutamate, D-glutamine, and D-glutamate metabolism were the main metabolic pathways for the Shaziling pig meat flavor (P<0.05). Moreover, the correlation coefficients revealed that the contents of anserine, C16:0, C16:1, and C18:1n9c were positively correlated with intramuscular fat and/or pH24h and were negatively correlated with the values of L* (lightness) and b* (yellowness) (P<0.05). In conclusion, age greatly affected the meat quality of Shaziling pigs, and the contents of muscular anserine, C16:0, C16:1, and C18:1n9c might be promising indicators for better meat quality.

Effects of Long-Term Protein Restriction on Meat Quality and Muscle Metabolites of Shaziling Pigs.

Background: It has been demonstrated that low-protein diets can improve the meat quality of pork. This study aimed to investigate the effects of long-term protein restriction from piglets to finishing pigs for 24 weeks on meat quality and muscle metabolites of Shaziling pigs. Results: Compared to the control group, reducing dietary protein levels by 20% reduced the L* value (p < 0.05), increased the a* value (p < 0.01), and tended to decrease pressing loss (p = 0.06) of longissimus thoracis muscle (LTM). Furthermore, compared to the control group, the −20% group had significantly lower levels of muscular danazol, N,N-dimethyl-Safingol, and cer(d18:0/14:0) (p < 0.05), all of which were positively associated with the L* value and negatively associated with the a* value (p < 0.05). Therefore, danazol, N,N-dimethyl-Safingol, and cer(d18:0/14:0) might be potential biomarkers for meat color. Conclusions: These results indicated that reducing dietary crude protein by 20% for 24 weeks could improve meat quality and alter muscular metabolites of Shaziling pigs, and the improvement in meat quality might be ascribable to decreased danazol, N,N-dimethyl-Safingol and cer(d18:0/14:0).

Long-Term Protein Restriction Modulates Lipid Metabolism in White Adipose Tissues and Alters Colonic Microbiota of Shaziling Pigs.

Obesity is a matter of concern to the public. Abundant evidence has been accumulated that nutritional intervention is a promising strategy to address this health issue. The objective of this study is to investigate alterations in the lipid metabolism in white adipose tissues and the gut microbiota of Shaziling pigs challenged by long-term protein restriction. Results showed that compared with the control group, reducing the protein level by 20% (−20%) increased the mRNA abundance of FABP4 in white adipose tissues (p < 0.05). This occurred in conjunction with increases in PPARγ protein expression. Conversely, the protein expression of C/EBPα was reduced in the −20% group (p < 0.05). Moreover, the −20% group had increased/decreased phosphorylation of AMPKα/mTOR, respectively (p < 0.05). As for the colonic gut microbiota, a 20% reduction in the protein level led to increased Lachnospiraceae XPB1014 group abundance at the genus level (p < 0.01). Collectively, these results indicated that a 20% protein reduction could modulate lipid metabolism and alter the colonic microbiota of Shaziling pigs, an approach which might be translated into a treatment for obesity.