STUDY OF B. PERTUSSIS ADAPTATION TO INTRACELLULAR SURVIVAL USING SEMI-QUANTITATIVE RT-PCR

VALDEZ, HUGO; LAMBERTI YANINA; ALVAREZ HAYES, JIMENA; GORGOJO JUAN PABLO; RODRIGUEZ, MARÍA EUGENIA

Buenos Aires

Congreso; First French-Argentine Immunology Congress (FAIC). LVIII reunion Annual de la Sociedad Argentina de Inmunología; 2010

Sociedad Argentina de Inmunología

Bordetella pertussis is the etiologic agent of whooping cough. Despite the high coverage of vaccination, the circulation of B. pertussis throughout the world continues largely unabated. The mechanisms that allow this pathogen to evade immune clearance are not yet clear. Our group recently showed that the encounter of B. pertussis with human macrophages in the absence of opsonic antibodies leads to the intracellular survival of a significant number of bacteria which, after a lag period, are able to replicate inside early endosomes. This ability to replicate seems to require bacterial adaptation to the intracellular environment. The goal of this study is to gain a first insight into the adaptive genetic mechanisms utilized by B. pertussis to survive, and eventually grow, inside macrophages. In this work, we report the study of two groups of genes usually implicated in pathogen adaptation to the phagosomal environment: genes involved in bacterial virulence and eventual host cell intoxication, and genes involved in iron homeostasis, one of the main stressors intracellular pathogens have to face. Using semi-quantitative RT-PCR we determined the mRNA expression of selected genes are indicative of iron stress response and bacterial virulence phase. The study of mRNA levels of B. pertussis inside the human macrophage show that genes that are repressed by iron as fumC, irp1-3, and sodA, are 20 to 30 times up regulated compared with infecting bacteria. Regarding virulence genes, cyaA was found down regulated 0,5 times while bipA was up regulated in bacteria inside the cell also 0,5 times, suggesting a possible intermediate virulence state in intracellular bacteria. Accordingly, no vrg-6 expression was observed. These early results suggest that B. pertussis adopts a particular phenotype that allows intracellular life.