The effect of biofilm materials on level of expression of Legionella pneumophila virulence factors : a longitudinal study

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The effect of biofilm materials on level of expression of Legionella pneumophila virulence factors : a longitudinal study

 

Author: Poon, Kin Chiu
Title: The effect of biofilm materials on level of expression of Legionella pneumophila virulence factors : a longitudinal study
Degree: M.Sc.
Year: 2014
Subject: Biofilms.
Legionella pneumophila.
Hong Kong Polytechnic University -- Dissertations
Department: Dept. of Health Technology and Informatics
Pages: xii, 75 leaves : illustrations ; 30 cm
Language: English
InnoPac Record: http://library.polyu.edu.hk/record=b2759007
URI: http://theses.lib.polyu.edu.hk/handle/200/7589
Abstract: Introduction: Legionella is the source of Legionnaires' disease, which can cause severe pneumonia. It is believed that biofilm is the reservoirs of Legionella in the environment. Biofilm acts as shelter and supports the growth of Legionella; however, the effect of biofilm materials on the Legionella virulence was not investigated. Therefore, biofilm might affect the virulence in Legionella and cause Legionella to become more invasive to human body. Since biofilm can easily be found in man-made water systems as well as in the environment, the findings might be helpful in preventing Legionnaires' disease. Therefore, the effect of biofilm materials on the level of expression of Legionella pneumophila virulence factors was investigated by using a longitudinal study approach. Aims: In this study, the effect of biofilm materials on the three target Legionella virulence genes, the global regulator letA, transporter protein in Dot/Icm type IV secretion system icmS and flagella subunit flaA, were investigated. Gene expression levels were investigated by quantitative real time RT-PCR with reference to housing keeping gene gyrB.
Method: Legionella pneumophila subsp. pneumophila (ATCC 33152) was used in this study. Filtered materials prepared from biofilm microorganisms from water tap were added to the AYE broth. The L. pneumophila suspension was added to the broth and incubated 18 days. Bacterial counts and real time RT-PCR were performed to analyze the growth patterns and gene expression levels respectively. Results were analyzed statistically in order to find out if there was significant difference in L. pneumophila growth pattern and gene expression in the presence and absence of biofilm materials. Results: After 18 days incubation period, the growth pattern and gene expression of Legionella were investigated. From the experiment, the first nine days were the growth phase or exponential phase of the L. pneumophila; while after nine days, Legionella started to enter the stationary phase or post-exponential phase. letA was less down-regulated in the presence of biofilm material group when compared with the absence of biofilm material group. However, the difference of expression was less than two-fold. icmS expression elevated at approximately seven-fold in the presence of biofilm material in day 3 when compared with the absence of biofilm materials. However, after day 3, the fold change of icmS had no significant difference between both groups. flaA showed down-regulation in both group throughout the first six days incubation period. However, after day 9, flaA expression was up-regulated. After 12 days, flaA expression was down-regulated again. After the statistical analysis by Wilcoxon Signed Rank test, growth of Legionella was not significantly different in the presence or absence of biofilm. Meanwhile, the biofilm materials also had no significant effects on the expression of three target virulence genes. Conclusion: Biofilm materials seemed not affecting the expression of the three virulence genes. However, there was difference in the expression level of icmS in first three days between two groups, but the final difference was not significant. Further studies on the relationships between biofilm and Legionella virulence factors should be done in order to investigate the regulatory effects of biofilm and biofilm materials on Legionella virulence expression, especially the icm genes.

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