title: Comparison of Plasmodium berghei challenge models for the evaluation of pre-erythrocytic malaria vaccines and their effect on perceived vaccine efficacy.
abstract: BACKGROUND
  The immunological mechanisms responsible for protection against malaria infection vary among Plasmodium species, host species and the developmental stage of parasite, and are poorly understood. A challenge with live parasites is the most relevant approach to testing the efficacy of experimental malaria vaccines. Nevertheless, in the mouse models of Plasmodium berghei and Plasmodium yoelii, parasites are usually delivered by intravenous injection. This route is highly artificial and particularly in the P. berghei model produces inconsistent challenge results. The initial objective of this study was to compare an optimized intravenous (IV) delivery challenge model with an optimized single infectious mosquito bite challenge model. Finding shortcomings of both approaches, an alternative approach was explored, i.e., the subcutaneous challenge.
  
  
  METHODS
  Mice were infected with P. berghei sporozoites by intravenous (tail vein) injection, single mosquito bite, or subcutaneous injection of isolated parasites into the subcutaneous pouch at the base of the hind leg. Infection was determined in blood smears 7 and 14 days later. To determine the usefulness of challenge models for vaccine testing, mice were immunized with circumsporozoite-based DNA vaccines by gene gun.
  
  
  RESULTS
  Despite modifications that allowed infection with a much smaller than reported number of parasites, the IV challenge remained insufficiently reliable and reproducible. Variations in the virulence of the inoculum, if not properly monitored by the rigorous inclusion of sporozoite titration curves in each experiment, can lead to unacceptable variations in reported vaccine efficacies. In contrast, mice with different genetic backgrounds were consistently infected by a single mosquito bite, without overwhelming vaccine-induced protective immune responses. Because of the logistical challenges associated with the mosquito bite model, the subcutaneous challenge route was optimized. This approach, too, yields reliable challenge results, albeit requiring a relatively large inoculum.
  
  
  CONCLUSIONS
  Although a single bite by P. berghei infected Anopheles mosquitoes was superior to the IV challenge route, it is laborious. However, any conclusive evaluation of a pre-erythrocytic malaria vaccine candidate should require challenge through the natural anatomic target site of the parasite, the skin. The subcutaneous injection of isolated parasites represents an attractive compromise. Similar to the mosquito bite model, it allows vaccine-induced antibodies to exert their effect and is, therefore not as prone to the artifacts of the IV challenge.
authors.affiliation: Malaria Vaccine Development, Walter Reed Army Institute of Research, Silver Spring, MD 20910, USA. wolfgang_leitner@nih.gov
authors.forename: Wolfgang W
authors.initials: WW
authors.lastname: Leitner
authors.forename: Elke S
authors.initials: ES
authors.lastname: Bergmann-Leitner
authors.forename: Evelina
authors.initials: E
authors.lastname: Angov
mesh_terms: D000818:Animals; D000852:Anopheles / Q000469:parasitology*; D019470:Biolistics; D004195:Disease Models, Animal; D005260:Female; D007271:Injections, Intradermal; D007273:Injections, Intramuscular; D007299:Insect Bites and Stings / Q000469:parasitology; D007303:Insect Vectors; D008288:Malaria / Q000209:etiology / Q000517:prevention & control*; D017780:Malaria Vaccines / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids / Q000235:genetics; D010962:Plasmodium berghei / Q000235:genetics / Q000276:immunology* / Q000472:pathogenicity; D015800:Protozoan Proteins / Q000235:genetics / Q000276:immunology; D015203:Reproducibility of Results; D012867:Skin / Q000469:parasitology; D034101:Sporozoites / Q000276:immunology; D014611:Vaccination; D019444:Vaccines, DNA / Q000235:genetics / Q000276:immunology*
chemical_list: D017780:Malaria Vaccines; D015800:Protozoan Proteins; D019444:Vaccines, DNA; C041595:circumsporozoite protein, Protozoan