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Subclinical mastitis (SCM) is a major constraint in dairy production and is driven by complex host-pathogen interactions. Although transcriptional responses associated with SCM have been widely investigated, the epigenetic mechanisms that stably regulate these programs remain less well characterized, particularly in crossbred cattle populations. This study aimed to characterize DNA methylation-based regulatory networks by integrating whole-genome methylation and transcriptome data from milk somatic cells of Vrindavani (Bos taurus × Bos indicus) cattle. Whole-genome methylation (n = 6) and corresponding transcriptome profiling (n = 6) were performed on milk somatic cells from SCM-affected and healthy control cows.
Differential methylation analysis (q-value < 0.05) identified 62,940 differentially methylated cytosines (DMCs), 7,706 differentially methylated regions (DMRs), and 6,203 differentially methylated genes (DMGs), with a predominant bias toward hypomethylation in SCM. Integrative analysis using stringent thresholds for both methylation (≥ 10%) and expression change (|log2 fold change| ≥ 1; P of GMM < 0.001) identified 1,407 differentially methylated and expressed genes (DMEGs). Functional enrichment analysis revealed 47 KEGG pathways and 30 Gene Ontology biological process terms (FDR < 0.05), primarily associated with immune signaling and inflammatory responses. In contrast, a subset of DMEGs showed methylation-associated repression of lactation- and metabolism-related genes. Selected genes were experimentally validated by qPCR, including upregulation of the inflammatory mediator S100A8 and downregulation of CSN3 (κ-casein), a key milk protein gene.
These findings provide an integrated view of the DNA methylation and transcriptional landscape of SCM in milk somatic cells and demonstrate that epigenetic remodeling is associated with coordinated activation of immune pathways alongside repression of lactation-associated functions. The results contribute to understanding the molecular basis of subclinical mastitis and may inform future efforts toward biomarker development and epigenetically informed strategies for improving disease resilience in dairy cattle.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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