Streptococcus pneumoniae utilizes a novel dynamin independent pathway for entry and persistence in brain endothelium.

Adoption of an endocytosis route promoting safe intracellular trafficking is a pre-requisite for development of invasive diseases by Streptococcus pneumoniae (SPN). We aim to explore the contribution of various endocytic routes in internalization and survival of SPN in blood brain barrier (BBB), a key event in development of pneumococcal meningitis. Pneumococcal entry and survival in brain endothelial cells were evaluated following treatment with combinations of inhibitors to block multiple endocytosis pathways leaving a single entry port open. Entry of SPN into brain endothelium through a novel dynamin independent pathway dictates a separate downstream trafficking itinerary. This allows SPN to evade lysosomal degradation, potentially promoting safe transit across BBB, leading to development of meningitis.

Histochemical Staining of Collagen and Identification of Its Subtypes by Picrosirius Red Dye in Mouse Reproductive Tissues.

Collagen is one of the foremost components of tissue extracellular matrix (ECM). It provides strength, elasticity and architecture to the tissue enabling it to bear the wear and tear from external factors like physical stress as well as internal stress factors like inflammation or other pathological conditions. During normal pregnancy or pregnancy related pathological conditions like preterm premature rupture of membranes (PPROM), collagen of the fetal membrane undergoes dynamic remodeling defining biochemical properties of the fetal membrane. The protocol in this article describes the histochemical method to stain total collagen by Picrosirius red stain which is a simple, quick and reliable method. This protocol can be used on paraformaldehyde (PFA) and formaldehyde fixed paraffin embedded tissue sections. We further describe the staining and distribution of collagen in different mouse reproductive tissues and also demonstrate how this technique in combination with polarization microscopy is useful to detect the distribution of different subtypes of collagen.

Membrane Vesicles of Group B Streptococcus Disrupt Feto-Maternal Barrier Leading to Preterm Birth.

Infection of the genitourinary tract with Group B Streptococcus (GBS), an opportunistic gram positive pathogen, is associated with premature rupture of amniotic membrane and preterm birth. In this work, we demonstrate that GBS produces membrane vesicles (MVs) in a serotype independent manner. These MVs are loaded with virulence factors including extracellular matrix degrading proteases and pore forming toxins. Mice chorio-decidual membranes challenged with MVs ex vivo resulted in extensive collagen degradation leading to loss of stiffness and mechanical weakening. MVs when instilled vaginally are capable of anterograde transport in mouse reproductive tract. Intra-amniotic injections of GBS MVs in mice led to upregulation of pro-inflammatory cytokines and inflammation mimicking features of chorio-amnionitis; it also led to apoptosis in the chorio-decidual tissue. Instillation of MVs in the amniotic sac also resulted in intrauterine fetal death and preterm delivery. Our findings suggest that GBS MVs can independently orchestrate events at the feto-maternal interface causing chorio-amnionitis and membrane damage leading to preterm birth or fetal death.