Negative energy balance (NEB) during the perinatal period leads to metabolic and immunological disorders in dairy cows, resulting in systemic responses and inflammation. The innate immune system is crucial for the host’s protection and inflammatory response. However, systematic research is still lacking on how NEB affects the innate immune system to alter the ’host defense capability and inflammatory response. In this investigation, raw transcriptome data of adipose, blood, endometrial, hypothalamus, and liver tissues were downloaded from a public database, cleaned, aligned, quantified, and batch-corrected. The innate immune gene list was retrieved from innateDB, followed by the expression matrix of innate immune genes in various tissues for differential expression analysis, principle component analysis (PCA), and gene set enrichment analysis (GSEA). Under the effect of NEB, adipose tissue had the most differentially expressed genes, which were predominantly up-regulated, whereas blood GSEA had the most enriched biological processes, which were predominantly down-regulated. The gene sets shared by different tissues, which are predominantly involved in biological processes associated with defense responses and inflammation, were dramatically down-regulated in endometrial tissues and highly up-regulated in other tissues. Under the impact of NEB, LBP, PTX3, S100A12, and LCN2 play essential roles in metabolism and immunological control. In conclusion, NEB can downregulate the defensive response of innate immune genes in endometrial, upregulate the immune and inflammatory response of other tissues, activate the host defense response, and increase the systemic inflammatory response. The analysis of the effects of NEB on innate immune genes from the multiple tissues analysis provides new insights into the crosstalk between metabolism and immunity and also provides potential molecular targets for disease diagnosis and disease resistance breeding in dairy cows.

The heritability of litter size in sheep is low and controlled by multiple genes, but the research on its related genes is not sufficient. Here, to explore the expression pattern of multi-tissue genes in Chinese native sheep, we selected 10 tissues of the three adult ewes with the highest estimated breeding value in the early study of the prolific Xinggao sheep population. The global gene expression analysis showed that the ovary, uterus, and hypothalamus expressed the most genes. Using the Uniform Manifold Approximation and Projection (UMAP) cluster analysis, these samples were clustered into eight clusters. The functional enrichment analysis showed that the genes expressed in the spleen, uterus, and ovary were significantly enriched in the Ataxia Telangiectasia Mutated Protein (ATM) signaling pathway, and most genes in the liver, spleen, and ovary were enriched in the immune response pathway. Moreover, we focus on the expression genes of the hypothalamic–pituitary–ovarian axis (HPO) and found that 11,016 genes were co-expressed in the three tissues, and different tissues have different functions, but the oxytocin signaling pathway was widely enriched. To further explore the differences in the expression genes (DEGs) of HPO in different sheep breeds, we downloaded the transcriptome data in the public data, and the analysis of DEGs (Xinggao sheep vs. Sunite sheep in Hypothalamus, Xinggao sheep vs. Sunite sheep in Pituitary, and Xinggao sheep vs. Suffolk sheep in Ovary) revealed the neuroactive ligand–receptor interactions. In addition, the gene subsets of the transcription factors (TFs) of DEGs were identified. The results suggest that 51 TF genes and the homeobox TF may play an important role in transcriptional variation across the HPO. Altogether, our study provided the first fundamental resource to investigate the physiological functions and regulation mechanisms in sheep. This important data contributes to improving our understanding of the reproductive biology of sheep and isolating effecting molecular markers that can be used for genetic selection in sheep.

Xinggao sheep are a breed of Chinese domestic sheep that are adapted to the extremely cold climatic features of the Hinggan League in China. The economically vital reproductive trait of ewes (litter size, LS) and productive traits of lambs (birth weight, BWT; weaning weight, WWT; and average daily gain, ADG) are expressed in females and later in life after most of the selection decisions have been made. This study estimated the genetic parameters for four traits to explore the genetic mechanisms underlying the variation, and we performed genome-wide association study (GWAS) tests on a small sample size to identify novel marker trait associations (MTAs) associated with prolificacy and growth. We detected two suggestive significant single-nucleotide polymorphisms (SNPs) associated with LS and eight significant SNPs for BWT, WWT, and ADG. These candidate loci and genes also provide valuable information for further fine-mapping of QTLs and improvement of reproductive and productive traits in sheep.

Follicle-stimulating hormone (FSH) regulates ovarian follicle development through specific gene expression programs. Granulosa cells (GCs) are somatic cells surrounding the oocytes, secreting gonadotropins to regulate ovulation and promote follicular development. By analyzing the effects of different doses of FSH on the proliferation of GCs, we found that adding 10 ng/mL of FSH, as the optimal concentration, could promote the growth of GCs. Furthermore, we have successfully constructed the first CRISPR-Cas9 knockout library targeting the genes on chromosomes 2 and 3 and the X chromosomes of the sheep massively parallel coding gene, as well as an ovarian GCs knockout cell library. For the first time, we have exposed the knockout cell library to a concentration of 10 ng/mL FSH to explore the underlying mechanisms. Through this screening, we have identified 836 positive–negative screening genes that are responsive to FSH, thereby revealing the regulatory mechanisms and screening the functionality of candidate genes. Next, RNA-Seq of control (0 ng/mL), low (10 ng/mL), and high (100 ng/mL) doses of FSH revealed 1708 differentially expressed genes, and combined with 836 genes, we obtained 129 FSH dose-dependent genes with extremely significant differences. This enables us to delve deeper into investigating and identifying the mechanisms by which FSH regulates GCs. More generally, we have discovered new regulatory factors and identified reproductivity-associated major effectors. These findings provide novel research directions for further studies on sheep reproduction.

Mycoplasma ovipneumoniae poses a major threat to sheep respiratory health, contributing to significant economic losses in farming communities. In this study, we isolated a novel strain, IM-DMQ, from a Hu sheep in Inner Mongolia that exhibited pulmonary adenomatous-like lesions, which is an uncommon pathological manifestation for this pathogen. The complete genome was sequenced using a hybrid Nanopore and Illumina approach, revealing a 1,039,804 bp circular chromosome with a GC content of 29.15%, encoding 1529 genes. Functional annotation highlighted genes involved in essential metabolic processes and potential virulence mechanisms. Comparative genomic analysis demonstrated that IM-DMQ shares the closest ancestry (ANI: 98.3%) with the Chinese strain NXNK2203, while structural variations and 14 unique genes distinguished it from other global strains. Furthermore, microbial community profiling of the original lung tissue revealed a co-infection background involving multiple bacterial pathogens, offering an etiological context for the severe disease presentation. These results provide the first complete genomic resource for an M. ovipneumoniae strain from Inner Mongolia associated with unusual pulmonary pathology, offering insights into its genetic diversity and supporting the future development of targeted diagnostics and vaccines for regional disease control.

Optimizing calcium metabolism is crucial for skeletal development and overall productivity in growing ruminants. Twenty-four Sunite lambs were randomly assigned to four groups and fed 0, 0.6, 1.2, or 2.4 g/(d·head) of γ-PGA for 60 days. Growth performance, serum parameters, duodenal morphology and calcium transporter expression, bone microarchitecture, and duodenal microbiota were analyzed. Supplementation with 1.2 g/(d·head) of γ-PGA (the M group) yielded optimal results, significantly improving final body weight and size. It enhanced duodenal health, evidenced by increased villus height, crypt depth, and microvilli density. Crucially, this dose significantly upregulated the expression of key duodenal calcium transporters (TRPV5/6, CaBPD9k, PMCA, VDR, claudin-12) and altered systemic calcium-regulating hormones (elevated calcitriol, PTH, FGF23). Bone micro-CT analysis revealed changes in trabecular architecture indicative of active remodeling. 16S rRNA sequencing and weighted OTU co-expression network analysis (WOCNA) revealed that γ-PGA reshaped the duodenal microbiota and identified core microbial modules strongly associated with host phenotypes. Genera such as [Eubacterium]_ruminantium_group, Fusicatenibacter, and Prevotella emerged as central hubs. In conclusion, dietary γ-PGA at 1.2 g/(d·head) enhances calcium absorption and bone metabolism in lambs through a coordinated modulation of intestinal integrity and calcium transport, systemic endocrine responses, and the duodenal microbial community, with specific microbiota identified as potential key mediators associated with these effects.