Phenotypic and Genetic Divergence among Poison Frog Populations in a Mimetic Radiation

The evolution of Müllerian mimicry is, paradoxically, associated with high levels of diversity in color and pattern. In a mimetic radiation, different populations of a species evolve to resemble different models, which can lead to speciation. Yet there are circumstances under which initial selection for divergence under mimicry may be reversed. Here we provide evidence for the evolution of extensive phenotypic divergence in a mimetic radiation in Ranitomeya imitator, the mimic poison frog, in Peru. Analyses of color hue (spectral reflectance) and pattern reveal substantial divergence between morphs. However, we also report that there is a “transition-zone� with mixed phenotypes. Analyses of genetic structure using microsatellite variation reveals some differentiation between populations, but this does not strictly correspond to color pattern divergence. Analyses of gene flow between populations suggest that, while historical levels of gene flow were low, recent levels are high in some cases, including substantial gene flow between some color pattern morphs. We discuss possible explanations for these observations

The TgsGP gene is essential for resistance to human serum in Trypanosoma brucei gambiense

Trypanosoma brucei gambiense causes 97% of all cases of African sleeping sickness, a fatal disease of sub-Saharan Africa. Most species of trypanosome, such as T. b. brucei, are unable to infect humans due to the trypanolytic serum protein apolipoprotein-L1 (APOL1) delivered via two trypanosome lytic factors (TLF-1 and TLF-2). Understanding how T. b. gambiense overcomes these factors and infects humans is of major importance in the fight against this disease. Previous work indicated that a failure to take up TLF-1 in T. b. gambiense contributes to resistance to TLF-1, although another mechanism is required to overcome TLF-2. Here, we have examined a T. b. gambiense specific gene, TgsGP, which had previously been suggested, but not shown, to be involved in serum resistance. We show that TgsGP is essential for resistance to lysis as deletion of TgsGP in T. b. gambiense renders the parasites sensitive to human serum and recombinant APOL1. Deletion of TgsGP in T. b. gambiense modified to uptake TLF-1 showed sensitivity to TLF-1, APOL1 and human serum. Reintroducing TgsGP into knockout parasite lines restored resistance. We conclude that TgsGP is essential for human serum resistance in T. b. gambiense

Cathepsin-L can resist lysis by human serum in Trypanosoma brucei brucei.

Closely related African trypanosomes cause lethal diseases but display distinct host ranges. Specifically, Trypanosoma brucei brucei causes nagana in livestock but fails to infect humans, while Trypanosoma brucei gambiense and Trypanosoma brucei rhodesiense cause sleeping sickness in humans. T. b. brucei fails to infect humans because it is sensitive to innate immune complexes found in normal human serum known as trypanolytic factor (TLF) 1 and 2; the lytic component is apolipoprotein-L1 in both TLFs. TLF resistance mechanisms of T. b. gambiense and T. b. rhodesiense are now known to arise through either gain or loss-of-function, but our understanding of factors that render T. b. brucei susceptible to lysis by human serum remains incomplete. We conducted a genome-scale RNA interference (RNAi) library screen for reduced sensitivity to human serum. Among only four high-confidence 'hits' were all three genes previously shown to sensitize T. b. brucei to human serum, the haptoglobin-haemoglobin receptor (HpHbR), inhibitor of cysteine peptidase (ICP) and the lysosomal protein, p67, thereby demonstrating the pivotal roles these factors play. The fourth gene identified encodes a predicted protein with eleven trans-membrane domains. Using chemical and genetic approaches, we show that ICP sensitizes T. b. brucei to human serum by modulating the essential cathepsin, CATL, a lysosomal cysteine peptidase. A second cathepsin, CATB, likely to be dispensable for growth in in vitro culture, has little or no impact on human-serum sensitivity. Our findings reveal major and novel determinants of human-serum sensitivity in T. b. brucei. They also shed light on the lysosomal protein-protein interactions that render T. b. brucei exquisitely sensitive to lytic factors in human serum, and indicate that CATL, an important potential drug target, has the capacity to resist these factors

Mechanism of Trypanosoma brucei gambiense resistance to human serum.

The African parasite Trypanosoma brucei gambiense accounts for 97% of human sleeping sickness cases. T. b. gambiense resists the specific human innate immunity acting against several other tsetse-fly-transmitted trypanosome species such as T. b. brucei, the causative agent of nagana disease in cattle. Human immunity to some African trypanosomes is due to two serum complexes designated trypanolytic factors (TLF-1 and -2), which both contain haptoglobin-related protein (HPR) and apolipoprotein LI (APOL1). Whereas HPR association with haemoglobin (Hb) allows TLF-1 binding and uptake via the trypanosome receptor TbHpHbR (ref. 5), TLF-2 enters trypanosomes independently of TbHpHbR (refs 4, 5). APOL1 kills trypanosomes after insertion into endosomal/lysosomal membranes. Here we report that T. b. gambiense resists TLFs via a hydrophobic beta-sheet of the T. b. gambiense-specific glycoprotein (TgsGP), which prevents APOL1 toxicity and induces stiffening of membranes upon interaction with lipids. Two additional features contribute to resistance to TLFs: reduction of sensitivity to APOL1 requiring cysteine protease activity, and TbHpHbR inactivation due to a L210S substitution. According to such a multifactorial defence mechanism, transgenic expression of T. b. brucei TbHpHbR in T. b. gambiense did not cause parasite lysis in normal human serum. However, these transgenic parasites were killed in hypohaptoglobinaemic serum, after high TLF-1 uptake in the absence of haptoglobin (Hp) that competes for Hb and receptor binding. TbHpHbR inactivation preventing high APOL1 loading in hypohaptoglobinaemic serum may have evolved because of the overlapping endemic area of T. b. gambiense infection and malaria, the main cause of haemolysis-induced hypohaptoglobinaemia in western and central Africa

Decoding the network of Trypanosoma brucei proteins that determines sensitivity to apolipoprotein-L1

In contrast to Trypanosoma brucei gambiense and T. b. rhodesiense (the causative agents of human African trypanosomiasis), T. b. brucei is lysed by apolipoprotein-L1 (apoL1)-containing human serum trypanolytic factors (TLF), rendering it non-infectious to humans. While the mechanisms of TLF1 uptake, apoL1 membrane integration, and T. b. gambiense and T. b. rhodesiense apoL1-resistance have been extensively characterised, our understanding of the range of factors that drive apoL1 action in T. b. brucei is limited. Selecting our bloodstream-form T. b. brucei RNAi library with recombinant apoL1 identified an array of factors that supports the trypanocidal action of apoL1, including six putative ubiquitin modifiers and several proteins putatively involved in membrane trafficking; we also identified the known apoL1 sensitivity determinants, TbKIFC1 and the V-ATPase. Most prominent amongst the novel apoL1 sensitivity determinants was a putative ubiquitin ligase. Intriguingly, while loss of this ubiquitin ligase reduces parasite sensitivity to apoL1, its loss enhances parasite sensitivity to TLF1-dominated normal human serum, indicating that free and TLF1-bound apoL1 have contrasting modes-of-action. Indeed, loss of the known human serum sensitivity determinants, p67 (lysosomal associated membrane protein) and the cathepsin-L regulator, ‘inhibitor of cysteine peptidase’, had no effect on sensitivity to free apoL1. Our findings highlight a complex network of proteins that influences apoL1 action, with implications for our understanding of the anti-trypanosomal action of human serum