A genetically attenuated malaria vaccine candidate based on P. falciparum b9/slarp gene-deficient sporozoites
- Radboud University Nijmegen Medical Center, Netherlands
- Leiden University Medical Center, Netherlands
- Ghent University and University Hospital, Belgium
- Université Pierre et Marie Curie-Paris 6, France
- INSERM, U1135, Paris, France
- Assistance Publique—Hôpitaux de Paris, Groupe hospitalier Pitié-Salpêtrière, France
- Sanaria Inc., United States
Figures
Figure 1 with 1 supplement
Generation and genotype analyses of P. berghei mutant PbΔb9Δslarp.
(A) Generation of mutant PbΔb9Δslarp. For PbΔb9Δslarp, the DNA-construct pL1740 was generated containing the positive/negative selectable marker cassette hdhfr/yfcy. This construct was subsequently …
Figure 1—figure supplement 1
Generation and genotype analyses of P. berghei mutant PbΔslarp-a.
(A) Generation of mutant PbΔslarp-a. For PbΔslarp-a, the DNA-construct pL1740 was generated containing the positive/negative selectable marker cassette hdhfr/yfcy. This construct was subsequently …
Figure 2 with 2 supplements
Phenotypes of P. falciparum PfΔslarp and PfΔb9Δslarp parasites.
(A) Gametocyte, oocyst, and sporozoite production. Gametocyte numbers (stage II and IV–V) per 1000 erythrocytes at day 8 and day 14 after the start of gametocyte cultures. Exflagellation (Exfl) of …
Figure 2—figure supplement 1
Consecutive gene deletion of slarp and b9 in P. falciparum.
Schematic representation of the genomic loci of (A) slarp (PF11_0480; PF3D7_1147000) on chromosome 11 (Chr. 11) and (B) b9 (PFC_0750w; PF3D7_0317100) on chromosome 3 (Chr. 3) of wild-type (wt; …
Figure 2—figure supplement 2
Genotype analysis of the generated PfΔslarp and PfΔb9Δslarp parasites.
(A) Long range PCR analysis of genomic DNA from WT, PfΔslarp and PfΔb9Δslarp asexual parasites confirms the slarp gene deletion and consecutive gene deletions of both slarp and b9 respectively and …
Figure 3
Development of P. falciparum PfΔslarp and PfΔb9Δslarp parasites in human primary hepatocytes.
(A) In vitro invasion of P. falciparum wt, PfΔslarp-a, PfΔslarp-b, PfΔb9Δslarp-F7, and PfΔb9Δslarp-G9 sporozoites in primary human hepatocytes. Invasion is represented as the mean ratio ± standard …
Tables
Table 1
Protection of mice after immunization with P. berghei PbΔb9 or PbΔb9Δslarp sporozoites
| Mouse strain | Pb mutant | Day of challenge* | Immunization regimes no. protected/no challenged | ||
|---|---|---|---|---|---|
| BALB/c | 10k† | 5k | 1k | ||
| PbΔb9 | 10 | 10/10‡ | 18/20 | 8/10 | |
| PbΔb9Δslarp | 10 | 20/20 | 10/10 | 20/20 | |
| C57Bl6 | 50/20/20k§ | 10/10/10k | 1/1/1k | ||
| PbΔb9 | 10 | 4/4 | nd | nd | |
| 90 | 5/5 | ||||
| 180 | 9/9# | ||||
| 365 | 5/11 | ||||
| PbΔb9Δslarp | 10 | Nd | 10/10 | 6/10 | |
| 180 | 6/6 | nd | nd | ||
-
*
Number of days post last immunization; 104 wild-type sporozoites were injected by IV route.
-
†
Immunization dose: number of sporozoites x1000.
-
‡
Protected/total # of immunized mice (%); protection was 0/15 in naive control BALB/c and 0/10 in C57BL/6 mice.
-
§
Immunization dose with 7 day intervals between immunizations.
-
#
Immunization dose 50/10/20k with 7 day intervals between immunizations. nd = not done.
Table 2
Breakthrough blood-stage infections after intravenous injection of PbΔslarp and PbΔb9Δslarp sporozoites
| Mouse strain | Mutant | Infection* Spz x 103 | Breakthrough blood infection/total # mice | Pre-patent period‡ (days) |
|---|---|---|---|---|
| BALB/c | WT† | 10 | 5/5 | 4–5 |
| PbΔslarp | 50 | 0/5 | ||
| PbΔslarp | 25 | 0/10 | ||
| PbΔb9Δslarp | 25 | 0/10 | ||
| C57BL/6 | WT† | 10 | 5/5 | 4–5 |
| PbΔslarp | 500 | 0/5 | ||
| PbΔslarp | 200 | 0/10 | ||
| PbΔb9Δslarp | 200 | 0/10 | ||
| PbΔb9Δslarp | 150 | 0/5 |
-
*
Inoculation dose of sporozoites administered IV.
-
†
P. berghei ANKA strain: line cl15cy1.
-
‡
Day with parasitemia of 0.5–2%.
Additional files
-
Supplementary file 1
Oocyst and sporozoite production and sporozoite characteristics (motility, traversal, hepatocyte invasion) of the P. berghei mutants PbΔslarp and PbΔb9Δslarp.
- https://doi.org/10.7554/eLife.03582.011
-
Supplementary file 2
Primer sequences.
- https://doi.org/10.7554/eLife.03582.012
