title: Applying Central Composite Design and Response Surface Methodology to Optimize Growth and Biomass Production of Haemophilus influenzae Type b.
abstract: BACKGROUND
  Haemophilus influenzae type b (Hib) is the leading cause of bacterial meningitis, otitis media, pneumonia, cellulitis, bacteremia, and septic arthritis in infants and young children. The Hib capsule contains the major virulence factor, and is composed of polyribosyl ribitol phosphate (PRP) that can induce immune system response. Vaccines consisting of Hib capsular polysaccharide (PRP) conjugated to a carrier protein are effective in the prevention of the infections. However, due to costly processes in PRP production, these vaccines are too expensive.
  
  
  OBJECTIVE
  To enhance biomass, in this research we focused on optimizing Hib growth with respect to physical factors such as pH, temperature, and agitation by using a response surface methodology (RSM).
  
  
  METHODS
  We employed a central composite design (CCD) and a response surface methodology to determine the optimum cultivation conditions for growth and biomass production of H. influenzae type b. The treatment factors investigated were initial pH, agitation, and temperature, using shaking flasks. After Hib cultivation and determination of dry biomass, analysis of experimental data was performed by the RSM-CCD.
  
  
  RESULTS
  The model showed that temperature and pH had an interactive effect on Hib biomass production. The dry biomass produced in shaking flasks was about 5470 mg/L, which was under an initial pH of 8.5, at 250 rpm and 35° C.
  
  
  CONCLUSIONS
  We found CCD and RSM very effective in optimizing Hib culture conditions, and Hib biomass production was greatly influenced by pH and incubation temperature. Therefore, optimization of the growth factors to maximize Hib production can lead to 1) an increase in bacterial biomass and PRP productions, 2) lower vaccine prices, 3) vaccination of more susceptible populations, and 4) lower risk of Hib infections.
authors.affiliation: Department of Biochemistry, Faculty of Biological Sciences, Tarbiat-Modares University, Tehran, IR Iran.
authors.forename: Seyed Bahman
authors.initials: SB
authors.lastname: Momen
authors.affiliation: Department of Bacteriology, Institute Pasteur of Iran, Tehran, IR Iran.
authors.forename: Seyed Davar
authors.initials: SD
authors.lastname: Siadat
authors.affiliation: Department of Microbiology, Faculty of Basic Sciences, Arak Branch, Islamic Azad University, Arak, IR Iran.
authors.forename: Neda
authors.initials: N
authors.lastname: Akbari
authors.affiliation: Department of Biochemistry, Faculty of Biological Sciences, Tarbiat-Modares University, Tehran, IR Iran.
authors.forename: Bijan
authors.initials: B
authors.lastname: Ranjbar
authors.affiliation: Department of Biochemistry, Faculty of Biological Sciences, Tarbiat-Modares University, Tehran, IR Iran.
authors.forename: Khosro
authors.initials: K
authors.lastname: Khajeh
keywords: Biomass; Growth Optimization; Haemophilus influenzae type b (Hib); Polyribosyl Ribitol Phosphate (PRP); Response Surface Methodology (RSM)