Borrelia burgdorferi
metodutveckling och tillämpning avseende odling och resistensstudier mot komplement, särskilt interaktion med faktor H
The genus Borrelia is a widespread, pathogenic pest and the causative of among others borreliosis or Lyme disease. The vector for the bacteria is the hard tick, Ixodes ricinus and the infection is transferred through a bite. Untreated, Borrelia may cause arthritis, heart damage or neuroborreliosis. Infection is made possible through different strategies for avoiding the body?s complement system. One such strategy involves the binding of factor H to specific bacterial membrane proteins and, thus, no complement activation and lysis. Another defence mechanism, phagocytosis, acts in cooperation with the complement and is subsequently retarded. The present study includes optimizing of Borrelia culturing, characterisation of different Borrelia strains in regard to sensitivity against the complement including culture counting and the analysis of free C3a and the Terminal Complement Complex (TCC). Further, tagging with FITC in order to study morphology as well as phagocytosis with the aid of microscopy and FACS was performed. The culturing experiments showed Borrelia to be rather easy to culture, although a strict sterile technique has to be applied. Concerning sensitivity to complement, the B.afzelii strains showed to be resistant to complement action, while most of the B. garninii are sensitive. Analysis of C3a and TCC after incubation with normal human blood serum as well as human whole blood, showed that complement activation demands rather or very high cell densities. Tagging with FITC followed by microscopic analysis resulted in good illustrations of the bacterial morphology. The FACS analysis resulted in findings of phagocytosis in both monocytes and granulocytes and, further, the different stages of phagocytosis were visualised through nuclear staining followed by microscopy. The genus Borrelia is a widespread, pathogenic pest and the causative of among others borreliosis or Lyme disease. The vector for the bacteria is the hard tick, Ixodes ricinus and the infection is transferred through a bite. Untreated, Borrelia may cause arthritis, heart damage or neuroborreliosis. Infection is made possible through different strategies for avoiding the body?s complement system. One such strategy involves the binding of factor H to specific bacterial membrane proteins and, thus, no complement activation and lysis. Another defence mechanism, phagocytosis, acts in cooperation with the complement and is subsequently retarded. The present study includes optimizing of Borrelia culturing, characterisation of different Borrelia strains in regard to sensitivity against the complement including culture counting and the analysis of free C3a and the Terminal Complement Complex (TCC). Further, tagging with FITC in order to study morphology as well as phagocytosis with the aid of microscopy and FACS was performed. The culturing experiments showed Borrelia to be rather easy to culture, although a strict sterile technique has to be applied. Concerning sensitivity to complement, the B.afzelii strains showed to be resistant to complement action, while most of the B. garninii are sensitive. Analysis of C3a and TCC after incubation with normal human blood serum as well as human whole blood, showed that complement activation demands rather or very high cell densities. Tagging with FITC followed by microscopic analysis resulted in good illustrations of the bacterial morphology. The FACS analysis resulted in findings of phagocytosis in both monocytes and granulocytes and, further, the different stages of phagocytosis were visualised through nuclear staining followed by microscopy.