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Chronic endometritis is associated with the imbalance of female reproductive tract microbiota and pathogenic microbial infection. This study aimed to identify the specific changes in the endometrial microbiome in patients with endometritis and to explore how Clostridium tyrobutyricum (C.t) influences the progression of endometritis in mice for further elucidating endometritis pathogenesis. For this purpose, endometrial tissues from 100 participants were collected and divided into positive, weakly positive, and negative groups based on CD138 levels, while endometrial microbiome differences were detected and analyzed using 16S rRNA gene sequencing. Staphylococcus aureus (S. aureus)-induced endometritis mouse model was established, followed by treatment with C.t, and inflammatory response, epithelial barrier, and TLR4/NF-κB pathway were evaluated. Results showed that α- and β-diversity was significantly lower in the positive group compared with the weakly positive or negative groups, where the negative group had more unique operational taxonomic units. The abundance of Proteobacteria was found to be increased, while that of Actinobacteria, Firmicutes, and Bacteroidetes was found to be reduced in the positive group, while the area under the curve value was found to be 0.664. Furthermore, C.t treatment resulted in the alleviation of S. aureus-induced inflammatory response, epithelial barrier damage, and activation of the TLR4/NF-κB pathway in mice. Clinical samples analysis revealed that the diversity and abundance of microbiota were altered in patients with endometritis having positive CD138 levels, while mechanistic investigations revealed C.t alleviated S. aureus-induced endometritis by inactivating TLR4/NF-κB pathway. The findings of this study are envisaged to provide a diagnostic and therapeutic potential of microbiota in endometritis. [ABSTRACT FROM AUTHOR]

Infection by positive-strand RNA viruses induces extensive remodeling of the host endomembrane system in favor of viral replication and movement. The integral membrane protein 6K2 of potyviruses induces the formation of membranous virus replication vesicles at the endoplasmic reticulum exit site (ERES). The intracellular trafficking of 6K2-induced vesicles along with microfilaments requires the vesicular transport pathway, actomyosin motility system, and possibly post-Golgi compartments such as endosomes as well. Recent studies have shown that endocytosis is essential for the intracellular movement of potyviruses from the site of viral genome replication/assembly site to plasmodesmata (PD) to enter neighboring cells. In this chapter, we describe a detailed protocol of how to use endomembrane trafficking pathway-specific chemical inhibitors and organelle-selective fluorescence dye to study the trafficking of potyviral proteins and potyvirus-induced vesicles and to unravel the role of endocytosis and the endocytic pathway in potyvirus infection in Nicotiana benthamiana plants., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)