Invasive African nontyphoidal Salmonella requires high levels of complement for cell-free antibody-dependent killing

J Immunol Methods. 2013 Jan 31;387(1-2):121-9. doi: 10.1016/j.jim.2012.10.005. Epub 2012 Oct 22.


Nontyphoidal isolates of Salmonella (NTS), particularly Salmonella Typhimurium, are a major cause of invasive bacteremia in Africa. Despite this, no vaccine against NTS is currently available for use in humans. If a NTS vaccine is to be developed in a timely manner, there is a need to develop assays to assess its in vivo efficacy. Assessment of potential efficacy of candidate vaccines in preclinical models is important for proof-of-concept and reduces attrition of vaccines in clinical trials. Serum bactericidal assays (SBA) are often used to assess the functional activity of vaccine-induced antibody responses targeted against Gram-negative bacteria with results given as the maximum dilution of serum that can effect bacterial killing. Previously we have found evidence for a protective role for antibody-induced complement-mediated killing of NTS in African children using an undiluted whole serum SBA. However, endogenous complement in diluted human sera is limiting and insufficient to effect bactericidal activity against S. Typhimurium beyond two two-fold dilutions. In the current study, we examined the requirements for SBA against NTS using baby rabbit serum (BRS) as an exogenous source of complement. We found that the amount of complement required for antibody-mediated bactericidal activity is much higher for the invasive African S. Typhimurium isolate D23580, compared with the laboratory S. Typhimurium LT2 and Salmonella Paratyphi A CVD1901. While 20% BRS was sufficient to kill LT2 and CVD1901, 75% BRS was needed to kill D23580. Our findings demonstrate that one concentration of exogenous complement is not suitable for SBA against all Salmonella isolates. To develop SBA to assess the in vivo efficacy of Salmonella vaccines, it is necessary to optimize the assay for the Salmonella isolates against which the vaccine is targeted.

MeSH terms

  • Adult
  • Africa
  • Animals
  • Antibodies, Bacterial / immunology*
  • Complement System Proteins / immunology*
  • Complement System Proteins / metabolism
  • Electrophoresis, Polyacrylamide Gel
  • Humans
  • Lipopolysaccharides / immunology
  • Lipopolysaccharides / metabolism
  • Microbial Viability / immunology
  • Rabbits
  • Salmonella / classification
  • Salmonella / immunology
  • Salmonella / metabolism
  • Salmonella Infections / immunology*
  • Salmonella Infections / metabolism
  • Salmonella Infections / microbiology
  • Salmonella Vaccines / immunology
  • Salmonella typhimurium / immunology*
  • Salmonella typhimurium / metabolism
  • Serum Bactericidal Antibody Assay / methods
  • Species Specificity


  • Antibodies, Bacterial
  • Lipopolysaccharides
  • Salmonella Vaccines
  • Complement System Proteins