Shengchao Ma

Bovine mastitis remains a major impediment to optimal dairy production. Somatic cell count (SCC) is commonly used as an indicator of mammary gland inflammation, while milk microbiota may also reflect mastitis-related changes. Here, we employed Oxford Nanopore full-length transcript sequencing to delineate the peripheral blood transcriptomic landscape of Xinjiang Brown cattle stratified by high (SCC ≥ 1,000,000 cells mL⁻¹) and low (SCC ≤ 200,000 cells mL⁻¹) SCCs, with the objective of identifying candidate genes underpinning mastitis resistance. We identified 226 differentially expressed genes and 441 differentially expressed transcripts. Genes in the high-SCC group were prominently enriched in immune response pathways and chemokine signalling cascades. Protein–protein interaction network analysis further delineated a core module of ten immune-related genes, including CCL4, IL1B and CXCL2. Integrative analysis with complementary second-generation sequencing data pinpointed CXCL2 as a high-priority candidate. Subsequent RT–qPCR and enzyme-linked immunosorbent assay (ELISA) validation revealed that CXCL2 expression was significantly elevated both in high-SCC individuals and in an LPS-induced bovine mammary epithelial cell inflammation model. Collectively, these findings establish CXCL2 as a putative molecular marker for mastitis resistance breeding and provide a foundational resource for deciphering the molecular mechanisms governing mammary health.

Dynamic changes in gene and transcript expression represent a key factor in regulating the cyclical development of hair follicles. In this study, based on Nanopore sequencing (ONT-seq) data of skin tissue from three developmental stages (anagen (An), catagen (Cn), and telogen (Tn)) of Jiangnan cashmere goat hair follicles, this study presents a profile of candidate DETs implicated in cycle regulation by delineating their stage-specific expression patterns and dynamic expression trends from anagen to telogen. A large proportion of the candidate DETs were significantly enriched in functions related to fat synthesis, storage, or metabolism, with significance levels of p < 0.05 or p < 0.01. These significantly enriched DETs, which were generally upregulated from An to Cn or downregulated from Cn to Tn, support a model where accelerated intradermal fat deposition promotes the progression from An to Cn, while its subsequent decrease facilitates the transition from Cn to Tn. Concurrently, our results also suggest a potential role for dynamic changes in AS patterns in regulating the hair follicle cycle. This regulatory role of AS patterns is potentially mediated through affecting genes related to lipid synthesis/metabolism or cell structure/interaction. Notably, a broader range of fat synthesis, storage, or metabolism-related transcripts showed significant differential expression (p < 0.05) in the An vs. Cn group. Ultimately, by establishing this candidate DET profile, we aim to provide fresh perspectives for deciphering the complex molecular regulation of the hair follicle cycle and to identify new targets for genetically enhancing or molecularly breeding cashmere traits in cashmere goats.

MicroRNA (miRNA) is a prominent topic in biological research, as it plays a crucial role in regulating various physiological processes. Identifying miRNAs associated with fiber traits enhances our understanding of the complex biological mechanisms involved in hair follicle development and provides potential targets for improving fine-wool sheep breeds. This study focused on 20 Ordos fine-wool sheep and identified miRNA expression profiles in skin tissues through transcriptome sequencing. Key miRNAs related to the diameter of wool fibers were preliminarily screened through cell verification experiments. The results revealed 26 differentially expressed (DE) miRNAs in tissues with varying wool fineness, with 12 DE miRNAs being upregulated and 14 DE miRNAs being downregulated. All DE miRNAs predicted 2,844 target genes. Integrating previous mRNA sequencing data, oar-miR-370-3p has 23 differentially expressed target genes. Following the overexpression of miR-370-3p in fibroblasts, the expression levels of the predicted target genes ZDHHC15 , JUNB , TGFBI , and IFI6 were upregulated compared to the control group, resulting in increased cell activity. These results provide new insights into the molecular mechanisms regulating the diameter traits of sheep wool fibers and offer important resources for precise sheep breeding research.

Reproductive and growth traits are key economic traits in sheep. This study aims to identify key single nucleotide polymorphisms (SNPs) and candidate genes associated with reproductive and growth traits in Tianmu polytocous sheep through a genome-wide association study (GWAS). The findings are expected to provide both a theoretical foundation for molecular breeding in this breed and novel insights into the genetic basis of ovine reproductive and growth performance. This study took 483 adult Tianmu polytocous ewes as the research subjects, collected their lambing records, measured their phenotypic values of growth traits (3 weight and 11 body size traits), and collected their blood samples for whole-genome resequencing to identify SNPs in the Tianmu polytocous sheep genome. The results identified a total of 9,499,019 (3× coverage) and 27,413,216 (30× coverage) high-quality SNPs in the Tianmu polytocous sheep genome. Subsequently, the association analysis between SNPs and reproductive and growth traits was conducted using a mixed linear model. A total of 92, 66, 18, 28, 6, 42, 3, 3, 6, 1, 12, 3, 22, 8, 6, and 3 SNPs were found associated with litter size at first parity, litter size at second parity, litter size at third parity, litter size at fourth parity, birth weight, weaning weight, body height, withers height, body length, head length, head width, cannon bone circumference, forelimb height, chest girth, chest depth, and withers width, respectively. Further, based on SNP annotation, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, candidate genes associated with the reproductive and growth traits were identified. Among these genes, 11 LOC, DEPTOR, GNG12, GRM7, PTH, PTH2R, WWOX, INHA, and NRG3 are candidate genes associated with litter size at first parity or litter size at third parity. These genes are involved in the G protein-coupled receptor signaling pathway, G protein-coupled receptor activity, ovarian tissue development, and hormone secretion. Additionally, TFRC and NTN1 are candidate genes associated with birth weight, while five UGT1A and CASR are candidate genes associated with weaning weight. These candidate genes are primarily involved in lipid metabolism. Finally, the following genes were identified as candidates associated with specific traits: DLG2, TMEM126A, and TMEM126B with body height; DSCAM and SCN8A with body length; BARX1 with cannon bone circumference; four LOC genes with forelimb height; EPHA4 with chest depth; and MRS2 with withers width.

To identify molecular markers associated with wool traits in fine-wool sheep, we examined genetic polymorphisms in the NOTCH2 and CD1A genes in 944 Subo Merino sheep in this study. Subsequently, we performed association analyses between mutation sites in the NOTCH2 and CD1A genes and wool traits using SAS 9.4 software, followed by linkage disequilibrium (LD) analysis of different mutation sites using Haploview 4.2 software. Additionally, bioinformatics tools were employed to predict the potential impacts of missense mutations on protein secondary and tertiary structures. Finally, quantitative PCR (qPCR) was used to assess the expression levels of the NOTCH2 and CD1A genes. Genetic analysis revealed six polymorphic sites in NOTCH2 and CD1A, all of which were missense mutations. Two SNPs in NOTCH2 (SNP1 and SNP2) showed significant associations with the coefficient of variation of fibre diameter, and SNP1 was also associated with greasy fleece weight. Four SNPs in CD1A (SNP3–SNP6) were significantly associated with fibre diameter standard deviation, and SNP3, SNP4, and SNP5 were additionally associated with crimp number. LD analysis revealed that SNP3, SNP4, and SNP5 were closely linked. Bioinformatics analysis indicated that the mutations caused alterations in the secondary and tertiary structures of the NOTCH2 and CD1A proteins. qPCR results showed that the CD1A gene was highly expressed in the fine wool fibre group compared with the ultra-fine wool fibre group. In conclusion, this study revealed a genetic association between NOTCH2 and CD1A and wool traits. The results are expected to provide a theoretical foundation for breeding wool traits in Subo Merino sheep, thereby enhancing the economic value of fine wool.

In this study, 944 Subo Merino sheep, a high-quality fine wool breed, were selected as research subjects. The SNP typing of the FAT3 gene was performed using the Fluidigm BiomarkTM HD system, and 11 missense mutation sites were identified. The analysis of population polymorphism of single-nucleotide polymorphisms was conducted. It is noteworthy that a substantial strong linkage disequilibrium was identified between SNP 5 and SNP 6 (r² > 0.8). The association between SNPs of the FAT3 gene and wool traits showed that multiple SNPs were significantly correlated with several different wool traits (p < 0.05). Furthermore, the investigation delved into the impact of the FAT3 gene on wool fiber through the utilization of quantitative polymerase chain reaction (qPCR), which yielded findings that this gene was notably expressed in fine wool fiber (FW) (p < 0.001). To predict the subcellular localization and protein transmembrane structure of FAT3, we employed the PSORT II Prediction and TMHMM online software. It was determined that the protein contains a transmembrane domain. This study provides molecular markers for the improvement of the selection and breeding of ultrafine-wool sheep and offers experimental evidence for accelerating the genetic breeding of sheep.

Xinjiang Brown cattle is an elite dual-purpose breed (raised for dairy and beef) developed in China. To elucidate its genomic architecture, we conducted whole-genome resequencing of 169 Xinjiang Brown cattle, followed by structural variation (SV) detection and a genome-wide association study (GWAS). We identified 71,668 SVs, among which deletions were the most prevalent, followed by translocations, inversions, duplications, and insertions. We further identified 1286 high-frequency SVs involving 2016 protein-coding genes. Through functional enrichment analysis of these genes, we revealed associations of genetic variation at genomic positions near genes implicated in immune response and disease resistance (NFKBIZ and PTPRT), growth and development (HDAC4 and MEF2A), and milk production (TP63, FABP4, and MEF2A). GWAS analysis of 31 body conformation traits revealed 58 SVs significantly associated with five traits (chest width, rear udder width, udder depth, rump width, and heel depth) at the genome-wide level. Additionally, nine candidate genes (CLINT1, EBF1, PAM16, GRIP1, CFAP54, SLC22A16, DOK5, ETAA1, and IPMK) were identified as potentially involved in the genetic regulation of body conformation traits. These findings provide novel insights for genetic improvement strategies and indicate that precision breeding could further enhance the production performance of this breed in the future.

Based on comprehensive proteomic analysis conducted across various stages of secondary hair follicles (SHFs), the growth and development regulatory mechanisms of SHFs in Jiangnan cashmere goats were studied. Proteomic analysis of skin tissue from the SHF anagen (An), catagen (Cn), and telogen (Tn) revealed 145 differentially expressed proteins (DEPs) between the An and Tn, 53 DEPs between the Cn and An, and 168 DEPs between the Cn and Tn. Gene Ontology (GO) annotations indicated that the DEPs were predominantly involved in keratin filament formation (KRTAP3-1, KRT1, KRT8), intermediate filament formation (KRT26, KRT35, KRT19, etc.), and lipid metabolism (FA2H, CERS6, ECH1, TECR, etc.). Furthermore, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis identified significant enrichment of DEPs in pathways related to hair follicle growth and development. Notably, these included the PPAR signaling pathway (PLIN2, PLIN4, ACSL5, etc.), the IL-17 signaling pathway (S100A7A, LOC108633164), and the estrogen signaling pathway (KRT26, KRT35, LOC102176457.). Western blotting (WB) experiments were then performed on five DEPs (KRT28, FA2H, PLIN2, FABP7, and VNN1) to validate the consistency of the WB results with the proteomic data. Overexpression and siRNA interference of PLIN2 in dermal papilla cells (DPCs) were followed by CCK8 and flow cytometry assays, revealing that PLIN2 knockdown significantly decreased DPC proliferation while inducing apoptosis, compared to controls. These findings suggest that the PLIN2 gene plays a crucial role in modulating SHF growth cycles in cashmere goats by influencing DPC proliferation. These results provide novel insights that could inform the development of breeding strategies aimed at enhancing the cashmere yield in such goats.

This study aims to elucidate the population structure and genetic diversity of Xinjiang brown cattle (XJBC) and Chinese Simmental cattle (CSC) while conducting genome-wide selective signatures analyses to identify selected genes associated with milk production traits in both breeds. Based on whole-genome resequencing technology, whole-genome single nucleotide polymorphisms (SNPs) of 83 Xinjiang brown cattle and 80 Chinese Simmental cattle were detected to resolve the genetic diversity and genetic structure of the two populations, whole-genome selective elimination analysis was performed for the two breeds of cattle using the fixation index (F_st) and nucleotide diversity (θπ ratio), and enrichment analysis was performed to explore their biological functions further. Both breeds exhibited relatively rich genetic diversity, with the Chinese Simmental cattle demonstrating higher genetic diversity than Xinjiang brown cattle. The IBS and G matrix results indicated that most individuals in the two populations were farther apart from each other. The PCA and neighbor-joining tree revealed no hybridization between the two breeds, but there was a certain degree of genetic differences among the individuals in the two breeds. Population structure analysis revealed that the optimal number of ancestors was three when K = 3. This resulted in clear genetic differentiation between the two populations, with only a few individuals having one ancestor and the majority having two or three common ancestors. A combined analysis of F_st and θπ was used to screen 112 candidate genes related to milk production traits in Xinjiang brown cattle and Chinese Simmental cattle. This study used genome-wide SNP markers to reveal the genetic diversity, population structure, and selection characteristics of two breeds. This study also screened candidate genes related to milk production traits, providing a theoretical basis for conserving genetic resources and improving genetic selection for milk production traits in Xinjiang brown cattle and Chinese Simmental cattle.

The aim of this research was to employ 16S rRNA high-throughput sequencing to thoroughly explore the interplay between milk and hindgut microbial communities and the effects of microorganisms in milk and the hindgut on the dairy quality of XJBC and CSC. In this study, 96 XJBC milk samples, 94 XJBC hindgut samples, 100 CSC milk samples, and 93 CSC hindgut samples were collected for microbial community analysis. The 16S rRNA sequencing data revealed that the microbial species richness in the milk of CSC exceeded that of XJBC, whereas the opposite was true for the hindgut microbial communities. A chi-square test was conducted using SPSS 19.0. The milk and posterior intestinal microbiota between individuals were analyzed with the Pearson chi-square test, maximum likelihood ratio, and Fisher’s exact test. Nongenetic factors substantially influenced microbial community dynamics in both milk and the hindgut. In the milk of dairy cows, a significant negative correlation was observed between one genus and milk protein production. Nine genera were significantly negatively correlated with milk fat production, whereas one genus was positively correlated. Additionally, six genera were negatively correlated with lactose production, and two genera exhibited positive correlations. Notably, Phascolarctobacterium and Turicibacter were identified as genera originating from the hindgut, which led to reduced milk quality. In the hindgut microbial community of dairy cows, seven genera were significantly negatively associated with milk fat production, whereas one genus was positively associated with milk fat production. These findings indicate that certain mammary microorganisms may migrate from the hindgut, either endogenously or exogenously, disrupting the equilibrium of the mammary microbial community in dairy cows and potentially leading to inflammation. By enhancing feeding conditions and standardizing production practices, the invasion of harmful flora into mammary tissues can be minimized, reducing the risk of inflammation and thereby preserving the health of dairy cows and enhancing milk quality.