Project description:Human breast milk contains a diverse community of bacteria but factors that produce variation in the breast milk microbiome are largely unknown. We evaluated if 1) maternal factors including breastfeeding practices modified the diversity and abundance of bacterial communities in breast milk and 2) if subclinical mastitis (SCM), an asymptomatic inflammatory condition occurring during lactation, induced a distinctive microbiota signature.
Project description:We report the application of miRNA next generation sequencing (NGS) for the analysis of impact of processing on miRNA in human breast milk, donated by 3 volunteers. MiRNA content of total and exosomal fraction was compared between unprocessed milk and sample subjected to either Holder (thermal) pasteurization (HoP) or elevated pressure processing (HPP). NGS reads were mapped to miRBase in order to obtain miRNA counts. Then, we analyzed differences in the miRNA abundance and function between raw and processed material. It was observed that both processing methods reduce number of miRNA reads and HoP is significantly more detrimental to miRNA than HPP.
Project description:Milk-derived extracellular vesicles (mEVs) have been proved to play a critical role in intercellular communication, mainly through the microRNAs (miRNAs) that they carry, to regulate biological functions of the target cells. Given miRNAs are evolutionarily conserved, EVs present in commercial milk may play a role in the physiology and health consumers. It is therefore essential to know the effects of technological treatments such as skimming and spray drying on the EV content of milk powders and on the cargo of bioactive molecules, in particular miRNAs, that they convey. Since goat’s milk or goat milk based formulas are considered as a healthy alternative for infants with cow’s milk sensitivities, including allergy, we undertook to analyze the EV content of skimmed and unskimmed goat's milk powders and to characterize their RNA content, in particular their miRNomes. mEVs were isolated using an optimized protocol based on Size Exclusion Chromatography (SEC) and compared regarding morphology, number and size by Transmission Electron Microscopy (TEM) and Nanoparticle Tracking Analysis (NTA). Their RNA and protein content were determined and their miRNomes established, using RNA sequencing. In this study we demonstrated that goat milk powders, skimmed or not upstream the spray drying treatment, contained many mEVs, ranging from 5.4 1011 to 2.5 1012 particles per mL of reconstituted milk, with an average size between 136.8 and 160.6 nm. We also demonstrated that mEVs carried significant amounts of RNA, including miRNAs. Using RT-qPCR, mRNAs encoding five of the major milk proteins were detected, suggesting that mEVs originated from mammary epithelial cells. We established the goat milk powder miRNome by identifying 351 miRNAs of which 233 are common to the 262 miRNAs previously profiled in raw goat milk. The 20 most abundant miRNAs (TOP 20) account for 80% of the total reads and the hierarchy of this TOP 20 miRNAs is somewhat overturned when comparing goat milk powder and raw goat milk. Surprisingly, whereas the comparison of raw from cow and goat milk confirmed the prevalence of miR-148a, miR-21-5p and miR-26a/miR-30a-5p, let-7a-5p and let-7f, which occupied ranks 1 and 2, respectively, in powders, were relegated to ranks 6 and 10 and 5 and 11 in raw goat and cow milk, respectively. Conversely to what was previously reported, we provide evidence that: i) EVs of typical morphology are present in goat milk powders; ii) mEVs survived the technological processes used to produce the powders; iii) their miRNA cargo is protected from degradation even though their miRNomes are not an exact mirror of miRNomes of EVs derived from fluid raw milk.
Project description:Human milk is highly recommended for infant during the first six month of life by World Healthy Organisation (WHO). Human milk is not only rich in macro-nutritional components, but also rich in cells and molecules. MicroRNAs are small non-coding RNAs, which enriched in human milk. These molecules are vital in enormous biological and cellular functions including immune system and in response to infections. By using deep sequencing method, 770,374,554 raw reads were generated from all samples (n=26). Then, filter analysis was done to remove 81,091,772 (10.5%), and 689,282,782 clean reads (89.5%) were considered as clean reads, which was retained for the subsequent bioinformatics analysis. Annotation and matching reads to miRBase revealed1780 mature known microRNAs identified in human milk cells and lipids derived from healthy, cold and other infection types mothers. In particular, 1680 known microRNAs were determined in infected mothers (n=14), while 1,606 known microRNAs in healthy mothers (n=12). Of these known microRNAs, 453 microRNAs were differentially expressed (p<0.05) between healthy and infected samples. The majority of the highly expressed miRNAs in all samples, in particular top 20 microRNAs, were also differentially expressed between healthy and infections. Further, 592 novel mature microRNA sequences were predicted, with only 65,878 total reads. Amongst the total reads of the novel microRNAs, top 20 novel miRNAs were found to contributed in 73.3% (total reads 48,295) of the total reads (65,878).
2017-12-22 | GSE80546 | GEO
Project description:bacterial community Raw sequence reads