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. 2022 Sep 16;13:5445. doi: 10.1038/s41467-022-33108-z

Fig. 4. Development and fly infection phenotypes of carp3 and flam8 KO.

Fig. 4

a Infection rates and (b) parasite densities in posterior midgut (PM), anterior midgut (AM) or proventriculus (PV or cardia) of tsetse flies infected with T. brucei AnTat 1.1 WT, carp3 KO1 or in situ CARP3 rescue (resc1) cells (Institute of Tropical Medicine Antwerp fly colony). Flies were dissected 21 days p.i. n (flies) = 15 (WT), 14 (KO1), 10 (resc1). Parasite densities were scored only in flies with positive midgut infection. Density scoring (parasites per field): ‘4’ > 1000, ‘3’ 100-1000, ‘2’ 10-100’, ‘1’ 1-10, ‘0’ no parasites. Note that in contrast to Fig. 1d no L-glutathione was added to the bloodmeal, resulting in lower midgut infection rates but increased sensitivity of phenotype detection. c Infection rates and (d) parasite densities in MG, PV or salivary glands (SG) of tsetse flies infected with T. brucei AnTat 1.1E ‘Paris’ (parent), flam8 KO (subclones KO1, KO2, KO3) or in situ FLAM8 rescue cells (Institut Pasteur Paris fly colony). All cell lines express a red fluorescent triple marker92. Flies were dissected 28 days p.i.; Mean ± SD of n (replicates, flies) = 6, 122 (WT); 4, 88 (KO1); 4, 109 (KO2); 3, 58 (KO3); 2, 40 (resc). Density scoring as in (b). Absolute transmission efficiencies differ between CARP3 and FLAM8 experiments due to different fly colonies and parasite strains. e RBP6-induced development in carp3 KO and parental RBP6 line. Procyclic, epimastigote and metacyclic forms were classified according to morphology, kinetoplast position relative to nucleus (DAPI staining) and expression of stage-specific marker proteins (see Supplementary Fig. 6). >100 cells were analyzed at each time point and replicate. Mean ± SD of n = 3. f Quantification of different developmental stages (procyclic, long mesocyclic trypomastigote, epimastigote) in PM or AM/PV of tsetse flies infected with T. brucei AnTat 1.1E ‘Paris’ (parent), flam8 KO (subclones KO1, KO2, KO3) or FLAM8 rescue cell lines. Flies were dissected 28 days p.i.; n (flies, trypanosomes) = 3, 689 (parent); 4, 522 (KO1); 2, 272 (KO2); 3, 304 (KO3); 2, 302 (rescue). Source data to (a–f) are provided as Source Data file.