For LYVE-1 blocking experiments mice were injected intra-peritoneally with 0.5 mg anti mLYVE-1 mAb2125 (R&D) or control rat IgG (isotype control or polyclonal) (R&D) 24 hours prior to infection. Data represent mean+/-SD) and representative fluorescence microscopy of Rabbit Polyclonal to Dynamin-1 (phospho-Ser774) adherent GAS (30 min incubation) in the presence of control mAb or LYVE-1 blocking mAb. Scale bars (20 m).(TIF) ppat.1005137.s003.tif (1.4M) GUID:?D748BB82-5329-473E-A667-A77D4445BC43 S4 Fig: Acapsular GAS adhere to human and mouse derived LECs independently of LYVE-1. Adhesion of M18capsule GAS to HDLECs (A), LYVE-1 lentivirus-transfected HDLECs (B) and MDLECs (C). Left to right; quantitative culture (n = 4; Data represent mean+/-SD) and representative fluorescence microscopy of adherent GAS (30 min incubation) in the presence of control mAb or LYVE-1 blocking mAb. Scale bars (20 m).(TIF) ppat.1005137.s004.tif (4.5M) GUID:?E94F9E19-73D7-4E37-A0AE-74E75A5BA08B S5 TMP 195 Fig: LYVE-1 functional blockade reduces M89 GAS dissemination to draining lymph nodes. Dissemination of M89 GAS in murine soft-tissue contamination following LYVE-1 mAb blockade (n = 8/group). Numbers of GAS at site of contamination (A) and ipsilateral lymph node (B), were determined by quantitative TMP 195 culture three hours post contamination. Lines depict median values in each case.(TIF) ppat.1005137.s005.tif (143K) GUID:?42D3EED4-ED78-46CE-9DC4-30114C889F0A S6 Fig: Prolonged LYVE-1 functional blockade re-routes M18 GAS to the blood circulation. Dissemination of M18 GAS at 24h after onset of murine soft-tissue contamination following LYVE-1 mAb blockade TMP 195 or control (n = 8/group). Numbers of GAS at site of contamination (A), ipsilateral draining LN (B), spleen (C) blood (D) and liver (E) were determined by quantitative culture 24 hours post contamination. Lines depict median values in each case (Mann Whitney U;* = p 0.05, ** = p 0.01).(TIF) ppat.1005137.s006.tif (319K) GUID:?7A738654-AF8C-4DC4-BEA8-8659050589D5 Data Availability StatementAll relevant data are within the paper and its TMP 195 Supporting Information files. Abstract The host lymphatic network represents an important conduit for pathogen dissemination. Indeed, the lethal human pathogen group A streptococcus has a predilection to induce pathology in the lymphatic system and draining lymph nodes, however the underlying basis and subsequent consequences for disease outcome are currently unknown. Here we report that this hyaluronan capsule of group A streptococci is usually a crucial virulence determinant for lymphatic tropism impeded bacterial dissemination to local draining lymph nodes and, in the case of a hyper-encapsulated M18 strain, redirected streptococcal entry into the blood circulation, suggesting a pivotal role in the manifestation of streptococcal infections. Our results reveal a novel function for bacterial capsular polysaccharide in directing lymphatic tropism, with potential implications for disease pathology. Author Summary Pathogens are known to invade the host not only via the systemic circulation but also via the lymphatic network, however the mechanisms underlying the latter route and the consequences for disease outcome have not been well studied. The important human pathogen, group A streptococcus, is responsible for a number of clinical syndromes affecting both the lymphatic vessels and draining lymph nodes, such as lymphangitis and lymphadenitis. How such pathologies are orchestrated, and their significance in the development of serious contamination are currently unknown. In this study, we show that this hyaluronan capsule secreted by group A streptococcus is critical for bacterial spread to draining lymph nodes, and we demonstrate that this occurs as a result of a specific conversation with the lymphatic vessel endothelial receptor-1. Genetic deletion or functional blockade of this receptor prevented streptococcal transit to draining lymph nodes in a TMP 195 murine model of contamination, which in turn enhanced bacterial spread into the blood circulation. Together these results define a novel interaction between the group A streptococcal capsule and the lymphatic endothelial receptor-1 as a critical axis in the establishment of lymphatic tropism for this pathogen, with clear implications for disease severity in the host. Introduction Lymphatic dissemination of intracellular bacteria and viruses is usually a well characterized mechanism of pathogenic invasion in the host, which occurs independently of transit through blood [1C3]. In contrast, exploitation of lymphatics by extracellular bacterial pathogens has received scant attention, despite clear clinical evidence that such pathogens can induce pathology within the lymphatic system [4,5]. Group A streptococcus (GAS) is usually one such important, exclusively human, extracellular pathogen. Pathology in the host is initiated by breach of mucosal surfaces and subsequent tissue destruction, resulting in a diverse disease spectrum spanning the superficial (pharyngitis, pyoderma) to the systemic (necrotizing fasciitis, toxic shock syndrome) as well as subsequent.