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. 2024 Mar 21;16(4):723–754. doi: 10.1038/s44321-024-00057-7

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© The Author(s) 2024

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. Creative Commons Public Domain Dedication waiver http://creativecommons.org/publicdomain/zero/1.0/ applies to the data associated with this article, unless otherwise stated in a credit line to the data, but does not extend to the graphical or creative elements of illustrations, charts, or figures. This waiver removes legal barriers to the re-use and mining of research data. According to standard scholarly practice, it is recommended to provide appropriate citation and attribution whenever technically possible.

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(A) Exflagellation centers were counted in mature gametocyte cultures of PfLARC2 on day 15 post set up for three PfLARC2 clones F7 (red), C12 (green), B7 (orange), and found to be comparable to PfNF54-WT (gray). These gametocyte cultures were fed to female Anopheles stephensi mosquitoes in standard membrane feeding assays and midguts were dissected on day 8 post feed. (B) Oocyst prevalence and, (C) counts for oocyst/mosquito and (D) sporozoites/mosquito were comparable to PfNF54-WT for PfLARC2 for clone F7, therefore clone F7 was used for all further phenotypic evaluation. (E) Research grade (non-GMP) sporozoites were generated from PfLARC2 clone F7 (PfSPZ-LARC2) by feeding mature gametocytes from PfLARC2 clone F7 to aseptic mosquitoes following aseptic procedures. Robust infection of mosquitoes was confirmed by (i) numbers of oocysts/midgut, (ii) oocyst prevalence and (iii) salivary gland sporozoites/mosquito. In (A-D), data is represented as mean +/− SD, n = 3 biological replicates. Statistical analysis was carried out using two-way ANOVA using Tukey’s multiple comparison test. P values are indicated in the figure. P > 0.05 is taken as not significant. In (E), data is represented as mean +/− SD, n = 3 biological replicates. For quantification of oocyst/midgut and oocyst prevalence, at least 12 mosquitoes were dissected per strain, and for quantification of sporozoites/mosquito, at least 50 mosquitoes were dissected per strain. Source data are available online for this figure.