Abstract
Sub-acute mastitis (SAM) is a prevalent disease among lactating women, being one of the main reasons for early weaning. Although the etiology and diagnosis of acute mastitis (AM) is well established, little is known about the underlying mechanisms causing SAM. We collected human milk samples from healthy and SAM-suffering mothers, during the course of mastitis and after symptoms disappeared. Total (DNA-based) and active (RNA-based) microbiota were analysed by 16S rRNA gene sequencing and qPCR. Furthermore, mammary epithelial cell lines were exposed to milk pellets, and levels of the pro-inflammatory interleukin IL8 were measured. Bacterial load was significantly higher in the mastitis samples and decreased after clinical symptoms disappeared. Bacterial diversity was lower in SAM milk samples, and differences in bacterial composition and activity were also found. Contrary to AM, the same bacterial species were found in samples from healthy and SAM mothers, although at different proportions, indicating a dysbiotic ecological shift. Finally, mammary epithelial cell exposure to SAM milk pellets showed an over-production of IL8. Our work therefore supports that SAM has a bacterial origin, with increased bacterial loads, reduced diversity and altered composition, which partly recovered after treatment, suggesting a polymicrobial and variable etiology.
Highlights
Sub-acute mastitis (SAM) is a prevalent disease among lactating women, being one of the main reasons for early weaning
Mean total load in the mastitis samples during the course of the symptoms reached 3,137,000 cells/ml (SEM = 956,632), which was significantly higher as compared to controls at the same time point
After the symptoms had disappeared, mean total load in the mastitis group decreased to 1,430,000 cells/ml (SEM = 259,037), the values were still significantly higher as compared to controls at time 0, and bacterial load did not fully return to healthy levels at this time point
Summary
Sub-acute mastitis (SAM) is a prevalent disease among lactating women, being one of the main reasons for early weaning. Most human milk microbiota studies based on molecular techniques focus on the total bacterial DNA composition, not considering that part of its DNA may correspond to dead or inactive bacteria, as well as free bacterial DNA For this reason, RNA-based sequencing of human samples is being used to clarify the elusive aetiology of some diseases with complex microbial origin[25,26]. The aim of the current work was to describe the human milk bacterial composition in mothers suffering SAM, taking into account total and active bacteria (as inferred by DNA and RNA 16S rRNA gene sequencing, respectively), and loads, in order to better characterize the etiology of the disease and find potential bacterial biomarkers. Bacterial pellets from human milk were exposed to a mammary epithelial cell line, in order to investigate their potential role in inflammatory processes
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