Optimization of culture conditions to improve Helicobacter pylori growth in Ham's F-12 medium by response surface methodology

Abstract

Helicobacter pylori is a gastroduodenal pathogen that colonizes the human stomach and is the causal agent of gastric diseases. From the clinical and epidemiological point of view, enhancing and improving the growth of this bacterium in liquid media is an important goal to achieve in order to allow the performance of accurate physiological studies. The aim of this work was to optimize three culture conditions that influence the growth of H. pylori in the defined medium Ham´s F-12 supplemented with 5% fetal bovine serum by using response surface methodology as a statistical technique to obtain the optimal conditions. The factors studied in this experimental design (Box-Behnken design) were the pH of the medium, the shaking speed (rpm) and the percentage of atmospheric oxygen, in a total of 17 experiments. The biomass specific growth rate (μ) was the response measured. The model was validated for pH and shaking speed. The percentage of atmospheric oxygen did not influence the growth for the range of values studied. At the optimal values found for pH and shaking speed, 8 and 130 rpm, respectively, a specific growth rate value of 0.164 h-1, corresponding to a maximal concentration of approximately 1.5x108 CFU/ml, was reached after 8 hours. The experimental design strategy allowed, for the first time, the optimization of H. pylori growth in a semi-synthetic medium, which may be important to improve physiological and metabolic studies of this “fastidious” bacterium.