Transcription profiling of Bos taurus mammary gland infected with E. coli reveals infection induces distinct local and systemic transcriptome responses in the mammary gland (Super Series)
ABSTRACT: This SuperSeries is composed of the following subset Series:; GSE15019: Transcriptome analysis of bovine mammary gland tissue treated with E. coli for 6 hours; GSE15020: Transcriptome analysis of bovine mammary gland tissue treated with E. coli for 24 hours; GSE15022: Transcriptome analysis of bovine mammary gland tissue of an udder quarter adjacent to an E.coli treated one for 24 hrs Experiment Overall Design: Refer to individual Series
Project description:By comparison of the transcriptome profiles of udder quarters neighboring to infected quarters and healthy udder tissue we analyse gene expression in the late stage of infection with E. coli 1303. Differentially expressed genes and transcription factors regulating coexpressed genes identified by SOTA clustering were used to identify key genes that define systemic reactions to infection with E. coli 1303. Experiment Overall Design: 10 samples, two conditions: udder quarters neighboring to one which was treated with E. coli for 24h vs healthy control * five replicates
Project description:By comparison of the transcriptome profiles of infected and healthy udder tissue we analyse gene expression in the late stage of infection with E. coli 1303. Differentially expressed genes and transcription factors regulating coexpressed genes identified by SOTA clustering were used to identify key genes that define late stage of infection with E. coli 1303. Experiment Overall Design: 10 samples, two conditions: E. coli treated for 24h vs healthy control * five replicates
Project description:By comparison of the transcriptome profiles of infected and healthy udder tissue we analyse gene expression in the early stage of infection with E. coli 1303. Differentially expressed genes and transcription factors regulating coexpressed genes identified by SOTA clustering were used to identify key genes that define early stage of infection with E. coli 1303. Experiment Overall Design: 10 samples, two conditions: E. coli treated for 6h vs healthy control * five replicates
Project description:Liver plays a profound role in the acute phase response (APR) observed in the early phase of acute bovine mastitis caused by Escherichia coli (E. coli). To gain an insight into the genes and pathways involved in hepatic APR of dairy cows we performed a global gene expression analysis of liver tissue sampled at different time points before and after intra-mammary (IM) exposure to E. coli lipopolysaccharide (LPS) treatment. Experiment Overall Design: Eight healthy, high yielding Holstein-Friesian dairy cows in their first lactation (9 to 12 weeks after calving) were chosen for this study. At time 0 the right front quarter was infused with 200 μg E. coli LPS dissolved in 10 ml 0.9% NaCl solution, the left front quarter serving as control was infused with 10 ml 0.9% NaCl solution. Liver biopsies were taken at –22, 3, 6, 9, 12 and 48 hours relative to LPS infusion in 4 cows, and also at –22, 9 and 48 hours in the remaining 4 cows. RNA from liver biopsies was isolated and biotin labeled cRNA was loaded onto the Affymetric GeneChip Bovine Genome Array. A control study using cows infused with 0.9% NaCl showed that there was no effect of taking the biopsy, neither in the clinical measurement nor in the expression of a selected subset of genes. Therefore, only samples taken from the LPS treated cows were measured for the gene expression using microarrays.
Project description:Previous studies have investigated the peptidomic changes occurring in cow milk during mastitis; however, these focused mainly on clinical mastitis, either spontaneous (Mansor et al., 2013) or induced by experimental infection (Thomas et al., 2016). Mansor and coworkers were the first to use mass spectrometry to demonstrate that several peptides found increased in milk from cows with clinical S. aureus or E. coli mastitis were mainly derived from aS1- and b-casein. In that study, 48 peptides were significantly different between the milks of healthy and mastitic cows. Non-mastitic samples were confirmed to be non-mastitic by having SCC <100,000 cells/mL (Mansor et al., 2013). Thomas and coworkers expanded the peptidomic repertoire in a study evaluating the kinetics of experimental S. uberis infection, and found signature peptides with potential as mastitis markers (Thomas et al., 2016). Only one study evaluated the milk peptidome in subclinical mastitis (Guerrero et al., 2015) demonstrating that even subclinical infections can cause significant increases in the total number of released peptides when compared to uninfected milk. However, neither the IMI agents nor the somatic cell counts were reported. With the aim of understanding high abundance protein and peptidomic changes due to subclinical CNS mastitis, to identify signature peptides with potential for subclinical mastitis detection, and to compare the proteomic and peptidomic findings with those reported in clinical mastitis, we investigated the influence of CNS IMI on high abundance milk proteins by SDS-PAGE and densitometric analysis, followed by a detailed characterization of the milk peptidome by means of high-performance liquid chromatography/tandem mass spectrometry and bioinformatic analysis.
Project description:Bovine mammary gland provide the largest amount of milk for dairy industry to date. Insight in functional adaptation of this organ is critical in order to improve efficiency of milk synthesis and milk quality. In the present experiment microarray analysis in combination with bioinformatics tools was performed in mammary tissue from 8 Holstein cows during the entire lactation cycle. The mammary biopsy was performed at -30, -15, 1, 15, 30, 60, 120, 240, and 300 days (d) relative to parturition. A dye-swap reference design (reference = mixture of RNA from several bovine tissues) was used.
Project description:MicroRNAs (miRNAs) are small noncoding RNAs that participate in regulation of gene expression. Their role during mammary gland development is still largely unknown. In the present study, we performed a microarray analysis to identify miRNAs associated with high mammogenic potential of bovine mammary gland. We identified 54 miRNAs differing significantly between mammary tissue of dairy (Holstein-Friesian, HF) and beef (Limousine, LM) post-pubertal heifers. Fifty two miRNAs had higher expression in the mammary tissue of LM heifers. Enrichment analyses for targeted genes revealed that the major differences between miRNA expression in the mammary gland of HF vs. LM were associated with regulation of signalling pathways crucial for mammary gland development, such as: TGF-beta, insulin, WNT and inflammatory pathways. Moreover, a number of genes potentially targeted by differentially expressed miRNAs was associated with mammary stem cells’ activity. These data indicate that in dairy cattle high developmental potential of the mammary gland, leading to high milk productivity, not only depends on central neuro-endocrine regulation but also on specific miRNA expression pattern. miRNA profiling of Holstein Freisian (dairy breed) and Limousne heifers (beef breed) mammay glands. Two-condition experiment, LM (test) vs. HF (reference). Total RNA was isolated from quarters of 4 LM and 4 HFmammary glands.
Project description:This investigation reports a differential proteomic analysis of the secretome of six S. aureus strains belonging to two genotypes with opposite within-herd prevalence, GTB (high) and GTS (low), corresponding to sequence types (ST) 8 and 398, respectively. Total proteins released in the growth medium were subjected to high-resolution tandem mass spectrometry and differential analysis with Proteome Discoverer by label-free approach. Here, we reported both the characterization of secretome proteins and a panel of differential proteins specific of S. Aureus depending on high or low within-herd prevalence. GTB/ST8 (High) released more immunoglobulin-binding proteins, complement and antimicrobial peptide inhibitors, enterotoxins, and metabolic enzymes, while GTS/ST398 (Low) released more leukocidins, hemolysins, lipases, and peptidases. Furthermore, GTB/ST8 (High) released the von Willebrand factor protein, staphylokinase, and clumping factor B, while GTS/ST398 (Low) released the staphylococcal coagulase and clumping factor A. In conclusion, our results provide an in-depth characterization of secretome proteins of the three S. aureus GTB/ST8 and three GTS/ST398 strains with high and low within-herd prevalence. We describe the role of extracellular proteins in S. Aureus pathogenesis.
Project description:The benefit of treatment in mild to moderate cases of E. coli mastitis in dairy cows remains a topic of discussion. We investigated the effect of intramammary treatment with Cefapirin + Prednisolone compared to Cefapirin only on gene expression profiles in experimentally induced E. coli mastitis. Five midlactating Holstein Friesian cows were challenged with 100 CFU in 3 quarters and treated with Cefapirin only in one quarter and the combination of Cefaprirn and Prednisolne in another quarter at 4, 12, 24 and 36h post challenge. After 24h (n=2) or 48h (n=3) post challenge cows were sacrificed and mammary gland tissue was collected from each quarter. From each cow total RNA of one non challenged quarter, one challenged not treated quarter, on Cefapirin treated and one Cefapirin + Prednisolone treated quarter was subjected to microarray analysis. Data were analyzed separately for the two sample time points
Project description:The objective of this study was to determine the effect of the DGAT1 K232A polymorphism on the global mRNA expression pattern of genes in the mammary gland tissue of grazing dairy cows in order to get more insight into the effects of this polymorphism on the physiology of the mammary glandgland of grazing dairy cows. Microarray analysis was used to identify genes whose expression was affected by the DGAT1 polymorphism in the mammary gland biopsies of 9 A232A cows, 13 K232A cows, and 4 K232K cows. The Microarray Analysis of Variance (MAANOVA) and Factor Analysis for Multiple Testing method (FAMT) were used to associate the expression of the genes present on Affymettrix Bovine Genome Arrays with the DGAT1 polymorphism. The data was also analysed at the level of functional modules by gene set enrichment analysis. In this experimental setting, DGAT1 polymorphism did not modify milk yield and composition significantly, although changes occurred in the yields of C14:0, C16:1cis-9, and some long chain fatty acids in milk. The DGAT1 polymorphism resulted in 30 differentially expressed genes related to cell growth, proliferation and development, signalling, remodelling and immune system. At the functional level, the pathways most affected by DGAT1 polymorphism were related to lipid biosynthesis, which likely reflected counter mechanisms of mammary tissue to respond to changes in milk FA composition, signalling, as well as immune system responses. A total of 28 Holstein-Friesian dairy cows in mid-lactation (DIM; 153 ± 32.8 days), milk yield (25.7 ± 3.08 kg/d) and fat content (4.3 ± 0.12%) were used in the study. Two consecutive milk samples (a.m. and p.m. milking) were obtained and pooled. One aliquot was stored at 4°C until analysis of fat, protein and lactose percentage, and another aliquot was frozen at -20°C until analysis for FA composition by gas chromatography. Specific details regarding the analysis of FA in milk are presented in Mach et al. (2011). Approximately 750 to 1,000 mg of mammary tissue from each cow was obtained by surgical biopsy. One part of the tissue was used for isolation of DNA and the other for extraction of the RNA. Genotyping of the DGAT1 polymorphism was performed using a TaqMan allelic discrimination method in an Applied Biosystems 7500 Real-Time PCR System (Applied Biosystems) as described by Schennink et al. (2007). The genotype at the DGAT1 locus was designated KK, KA, or AA for homozygous Lysine, heterozygous Lysine/Alanine, or homozygous Alanine, respectively. Microarray analysis was used to identify genes whose expression was affected by the DGAT1 polymorphism in the mammary gland biopsies of 11 A232A cows, 13 K232A cows, and 4 K232K cows. Total RNA from mammary gland tissue (50-100 mg) was isolated using TRIzol reagent (Invitrogen, Breda, The Netherlands), following the manufacturer’s instructions. The RNA purity and concentrations were determined using a NanoDrop ND-1000 spectrophotometer (Isogen, Maarssen, the Netherlands), and the RNA quality was assessed using the BioAnalyzer 2100 (Agilent Technologies, Amsterdam, the Netherlands). The RNA of each biopsy was amplified, biotin-labeled, and hybridized to single-dye Affymetrix GeneChip® Bovine Genome Array (#900493) by ServiceXS (Leiden, the Netherlands)