Fig 1.
Parasite histories and culture.
The small pictures are from Biorender.com. (https://app.biorender.com).
Fig 2.
Growth of MAK65 and MAK98 strains in rodents.
A. Parasitaemias after infection of three or four mice with 1,000 strain MAK65 parasites. "d" indicates death and "k" indicates killing because of clear symptoms. The y axis scale is the log10 of the parasitaeimia. B. Parasitaemias after infection of 3 mice with 1,000 strain MAK98 parasites. C. Growth of both strains in rats before harvest for RNA purification. The percentage of PAD1 positive cells at the time of harvest is indicated. D. The transcriptomes of population 65 (rat A) and 98 (rat A) parasites (panel C) were compared and the log2 ratio (65A/98A, S1 Table, sheet 1) is on the y-axis. The x-axis shows published stumpy/long slender results for trypanosomes grown in mice [22]. The correlation coefficients are from linear regression analysis using the log-transformed rations, and were generated in Microsoft Excel.
Fig 3.
Cumulative cell counts for all cultures that were used for DNA preparation. Full details of these cultures with linear-scale plots are in S2 Fig panels C-I.
Fig 4.
Copy number distributions for different trypanosome strains and cultures.
After genome sequencing all reads were allowed to align 20 times. For a set of unique genes, containing one representative each for each set of repeated genes, reads per million reads were calculated. Copy numbers for each gene were then calculated based on the assumption that most genes are present once in every haploid genome (or twice per diploid genome). The numbers of genes with copy numbers between 1 and 4 are plotted here for the strains and cultures indicated. A. T. rhodesiense, TREU927, EATRO1125 and Lister 427. B. MAK65 from rats and culture C. MAK98 from rats and culture.
Fig 5.
Gene copy numbers of standard strains and a T. rhodesiense isolate, plotted across the genome.
Copy numbers for each unique open reading frame were determined as in Fig 3. The unique genes were then placed in order along the chromosomes (S2 Table sheet 3) and their copy numbers plotted with one bar for each unique gene. The approximate chromosome boundaries are indicated on the plot; columns exceeding 8 copies are truncated at the top of each panel. Red bars indicate departures from diploidy. The nature of the sample is indicated on each panel. A: TREU927 (genome strain); B: Cultured EATRO1125 bloodstream forms expressing the tet repressor; C: Lister 427 2T1 bloodstream forms expressing the tet repressor and T7 polymerase (427_2T1) [45]; D: Lister 427 bloodstream forms expressing the tet repressor (427_1313) [31]; E: Lister 427 procyclic forms (427_PC2060) expressing the tet repressor; F, G: T. b. rhodesiense from humans, passaged 3 times in rodents [16].
Fig 6.
Gene copy numbers for original and cultured MAK65 and MAK98 A-F: Copy numbers plotted across the genome, details as in Fig 4.
G: The average read densities for DNA from rat-grown MAK98 were divided by those for MAK65 and plotted across the genome (panel D / panel A). The results are on a log scale. H: The average read densities for RNA from rat-grown MAK98 were divided by those for MAK65 and plotted across the genome. The results are on a log scale. Many differences are likely to be due to the differentiation state (Fig 1C and 1D) but MAK98 shows clear RNA over-representation from chromosome 5.
Fig 7.
Copy numbers of selected genes that appeared to be influenced by culture.
The gene products and numbers are shown above each graph; the y-axis shows the haploid gene copy number taken from S2 Table, sheet 1. Columns with green underlay are for non-culture-selected parasites, and those with blue underlay were culture-selected. Colours indicate the origin. The four T. b. rhodesiense isolates (Tbr) are from independent patients but have been assigned the same colour because they were all taken from a single clinic. The same colours for underlay and text are used in S2 Table. Panels A-I are different genes as indicated.