ABSTRACT
The Eurasian beaver (Castor fiber) has been reintroduced successfully in Germany since the 1990s. Since wildlife is an important source of zoonotic infectious diseases, monitoring of invasive and reintroduced species is crucial with respect to the One Health approach. Three Eurasian beavers were found dead in the German federal states of Bavaria, North Rhine-Westphalia and Baden-Wuerttemberg in 2015, 2021 and 2022, respectively. During post-mortem examinations, Corynebacterium (C.) ulcerans could be isolated from the abscesses of two beavers and from the lungs of one of the animals. Identification of the bacterial isolates at the species level was carried out by spectroscopic analysis using MALDI-TOF MS, FT-IR and biochemical profiles and were verified by molecular analysis based on 16-23S internal transcribed spacer (ITS) region sequencing. Molecular characterization of the C. ulcerans isolates using whole-genome sequencing (WGS) revealed a genome size of about 2.5 Mbp and a GC content of 53.4%. Multilocus sequence typing (MLST) analysis classified all three isolates as the sequence type ST-332. A minimum spanning tree (MST) based on cgMLST allelic profiles, including 1211 core genes of the sequenced C. ulcerans isolates, showed that the beaver-derived isolates clearly group on the branch of C. ulcerans with the closest relationship to each other, in close similarity to an isolate from a dog. Antibiotic susceptibility testing revealed resistance to clindamycin and, in one strain, to erythromycin according to EUCAST, while all isolates were susceptible to the other antimicrobials tested.
ABSTRACT
BACKGROUND: Human Parvovirus B19 (PVB19) has been associated with myocarditis putative due to endothelial infection. Whether PVB19 infects endothelial cells and causes a modification of endothelial function and inflammation and, thus, disturbance of microcirculation has not been elucidated and could not be visualized so far. METHODS AND FINDINGS: To examine the PVB19-induced endothelial modification, we used green fluorescent protein (GFP) color reporter gene in the non-structural segment 1 (NS1) of PVB19. NS1-GFP-PVB19 or GFP plasmid as control were transfected in an endothelial-like cell line (ECV304). The endothelial surface expression of intercellular-adhesion molecule-1 (CD54/ICAM-1) and extracellular matrix metalloproteinase inducer (EMMPRIN/CD147) were evaluated by flow cytometry after NS-1-GFP or control-GFP transfection. To evaluate platelet adhesion on NS-1 transfected ECs, we performed a dynamic adhesion assay (flow chamber). NS-1 transfection causes endothelial activation and enhanced expression of ICAM-1 (CD54: mean ± standard deviation: NS1-GFP vs. control-GFP: 85.3 ± 11.2 vs. 61.6 ± 8.1; P<0.05) and induces endothelial expression of EMMPRIN/CD147 (CD147: mean ± SEM: NS1-GFP vs. control-GFP: 114 ± 15.3 vs. 80 ± 0.91; P<0.05) compared to control-GFP transfected cells. Dynamic adhesion assays showed that adhesion of platelets is significantly enhanced on NS1 transfected ECs when compared to control-GFP (P<0.05). The transfection of ECs was verified simultaneously through flow cytometry, immunofluorescence microscopy and polymerase chain reaction (PCR) analysis. CONCLUSIONS: GFP color reporter gene shows transfection of ECs and may help to visualize NS1-PVB19 induced endothelial activation and platelet adhesion as well as an enhanced monocyte adhesion directly, providing in vitro evidence of possible microcirculatory dysfunction in PVB19-induced myocarditis and, thus, myocardial tissue damage.
