Plasmodium infections

Springer, Andrea, Fichtel, Claudia, Sebastien ́ Calvignac-Spencer, Leendertz, Fabian H. & Kappeler, Peter M., 2015, Hemoparasites in a wild primate: Infection patterns suggest interaction of Plasmodium and Babesia in a lemur species, International Journal for Parasitology: Parasites and Wildlife 4 (3), pp. 385-395 : 388-390

3.1. Plasmodium infections

Thirteen of 36 individuals tested PCR-positive for Plasmodium spp. infection. Seven individuals were tested more than once, and repeated sampling at different times of the year almost always gave identical results for these animals. Only results for one animal (age 9) switched from negative to positive during the study period. Population prevalence is thus estimated at 36%. We did not observe any parasite stages in Diff-quick ® -stained blood smears.

Only two unique sequences were amplified in this study, which showed 95 ‾ 100% sequence identity to publicly available Plasmodium sequences. Most sequences could be unambiguously assigned to Plasmodium sp. LLD-2010 which has previously been found in Verreaux's sifakas ( Duval et al., 2010). Two animals carried a different parasite (EMBL accession no.: LN869522), which unambiguously nested within the clade of malaria parasites infecting lemurs (Bp 100; Fig.1 ). This parasite appeared as most closely related to Plasmodium sp. C-MAP-2012 (which has been found in Hapalemur griseus ( Pacheco et al., 2011)) , although the statistical support for this association was moderate (Bp 59; Fig.1 ). The patristic distance between these two sequences was 0.21 substitution per site (s.s —1), which is both higher than the 0.17 s s —1 observed between the two most closely related lemur parasite sequences known to date ( Plasmodium sp. B-MAP.2012 and E-MAP-2011) and higher than the maximum 0.14 s s —1 observed between P. reichenowii (clade C1, infecting chimpanzees) and P. praefalciparum sequences (clade G1, infecting gorillas; Rayner et al., 2011). Coinfections with both lemur malaria parasites were not observed.

A. Springer et al. / International Journal for Parasitology: Parasites and Wildlife 4 (2015) 385 ‾ 395 389

Age seemed to affect Plasmodium spp. test outcome, with older animals being more likely to test positive ( Fig. 2 ). In fact, none of the animals up to the age of 5 years were infected (n = 18), 50% of animals between age 6 and 10 were infected (n = 10) and all animals over the age of 10 tested positive (n = 8). However, in the GLMM including age as well as sex, season and group size as fixed factors, age only tended to have a statistically significant effect ( Table 2). Sex, season and group size had no significant effect on Plasmodium spp. infection status.

3.2. Babesia infections and potential Plasmodium ‾ Babesia interaction

Prevalence of Babesia infections was 55.6%. Again, test results for only 1 individual (age 7) changed from negative to positive during the study period. The Babesia sequence amplified (EMBL accession no. LN869519) was the same in all samples and showed 94% identity to different Babesia canis isolates. We did not microscopically observe any Babesia parasite stages in Diff-quick ® -stained blood smears.

Only age significantly affected Babesia sp. test outcome, but contrary to the Plasmodium sp. infections, young animals were more likely to test positive ( Table 3, Fig. 2 ). Only 3 animals (6, 7 and 11 years old) were co-infected with Plasmodium sp. and Babesia sp. Based on this complementary age pattern, the low number of coinfections, and because neither age, sex, season nor group size influenced Plasmodium spp. infections, we tested Babesia infection status as a predictor of Plasmodium infection, and found a highly significant negative effect ( Table 4).

Figures

Fig.1. Maximum likelihood tree of malaria parasite cytochrome b sequences (P.= Plasmodium).The clade formed by lemur malaria parasites is blue. The two sequences detected in this study are highlighted with grey rectangles. Bootstrap values are reported above branches when>50. The scale is in substitution per site.

Fig. 2. Age-dependence of Babesia sp. infections (grey) and Plasmodium sp. infections (black).The lines represent the predicted values according to the two different GLMMs.