| Popis: |
Streptococcus pneumoniae or pneumococcus is included among major human pathogens and is responsible for a number of diseases including life-threatening conditions such as pneumonia, meningitis and sepsis. Though pneumococcal vaccines are available, they provide limited coverage against infections as pneumococcus shows extensive variation, which also allows escape from vaccines and antibiotic resistance. It is armed with several virulence factors including capsule, surface proteins, enzymes and toxins, which are variably expressed and altogether determine pneumococcal virulence. The aim of this project was to study pneumococcal genetic variation and its effect on virulence, with a focus on pneumococcal capsule, which is considered the major determinant of virulence and is involved in interaction with host immune system. It is the target for current vaccines and at least 93 pneumococcal serotypes are known, which differ in pathogenicity. To study the effect of capsule on the pneumococcal virulence, capsule-switch mutants were constructed in three genetic backgrounds; TIGR4 (serotype 4, virulent), 403 (serotype 4, avirulent) and D39 (serotype 2, virulent) and were studied for variation in their in vivo and in vitro characteristics. These mutants were compared with their parent strains and other mutants for effects of capsule switching on their growth, formation of capsular polysaccharide, capsular thickness, chain formation and virulence in murine models of infection using MF1 mice. Significant differences were observed in behaviour of parent and mutant strains. To develop a broader insight into pneumococcal virulence, avirulent derivative of strain TIGR4, 403 was genome sequenced and compared with TIGR4 for genetic mutations. To study differences in gene expression both the strains were also compared using microarrays. Genome analysis revealed only few mutations in strain 403 but microarray experiments showed 288 genes to be expressed differently in strain 403. Strain 403 was also tested as live attenuated vaccine to see if it could provide protection against the same and different serotypes, as it can be used as a vehicle for delivery of different polysaccharides to the host body along with the whole set of pneumococcal antigenome. Vaccine trials of 403 were not very fruitful as it failed to provide any protection through intranasal route though partial protection was observed in mice vaccinated intraperitoneally with significant differences in levels of bacteraemia, survival, weight and temperature losses on challenging with homologous strain. |
