Kinetoplastid parasites—trypanosomes and leishmanias—infect millions of humans and cause economically devastating diseases of livestock, and the few existing drugs have serious deficiencies. Benzoxaborole-based compounds are very promising potential novel anti-trypanosomal therapies, with candidates already in human and animal clinical trials. We investigated the mechanism of action of several benzoxaboroles, including AN7973, an early candidate for veterinary trypanosomosis. In all kinetoplastids, transcription is polycistronic. Individual mRNA 5'-ends are created by trans splicing of a short leader sequence, with coupled polyadenylation of the preceding mRNA. Treatment of Trypanosoma brucei with AN7973 inhibited trans splicing within 1h, as judged by loss of the Y-structure splicing intermediate, reduced levels of mRNA, and accumulation of peri-nuclear granules. Methylation of the spliced leader precursor RNA was not affected, but more prolonged AN7973 treatment caused an increase in S-adenosyl methionine and methylated lysine. Together, the results indicate that mRNA processing is a primary target of AN7973. Polyadenylation is required for kinetoplastid trans splicing, and the EC50 for AN7973 in T. brucei was increased three-fold by over-expression of the T. brucei cleavage and polyadenylation factor CPSF3, identifying CPSF3 as a potential molecular target. Molecular modeling results suggested that inhibition of CPSF3 by AN7973 is feasible. Our results thus chemically validate mRNA processing as a viable drug target in trypanosomes. Several other benzoxaboroles showed metabolomic and splicing effects that were similar to those of AN7973, identifying splicing inhibition as a common mode of action and suggesting that it might be linked to subsequent changes in methylated metabolites. Granule formation, splicing inhibition and resistance after CPSF3 expression did not, however, always correlate and prolonged selection of trypanosomes in AN7973 resulted in only 1.5-fold resistance. It is therefore possible that the modes of action of oxaboroles that target trypanosome mRNA processing might extend beyond CPSF3 inhibition.

The entomological efficacy of using 25% deltamethrin EC insecticide-treated bednets (ITNs) was evaluated against the sandfly Lutzomyia longipalpis Lutz and Neiva (Diptera: Psychodidae), the principal vector of zoonotic visceral leishmaniasis (ZVL) in Latin America. A crossover field study in Amazon Brazil (Marajó Island, Pará State) demonstrated that, compared with untreated nets, the insecticide increased the barrier effect of nets by 39% (95% confidence interval [CI] 34-44%), reduced human landing rates by 80% (95% CI 62-90%) and increased the 24-h mortality rate from 0% to 98% (95% CI 93-99%) inside ITNs. The presence of an ITN also reduced the human landing rate on unprotected persons outside the net in the same room by 56% (95% CI 52-59%), and increased 24-h mortality to 68% (95% CI 62-73%) compared to 0.4% (0.1-2.0%) in untreated houses. The reduction in human landing rates in ITN rooms was associated with a doubling in the proportion of sandflies alighting on walls compared with that in untreated rooms, which was attributed to insecticide-induced excito-repellency. There was no evidence that sandflies were diverted onto unprotected hosts. Human landing catches inside houses peaked between 19.00 hours and 23.00 hours and declined steadily to zero at 02.00 hours and thereafter. House-to-house questionnaires established that only 34% of households owned at least one net (median two, range 1-8), only 20% of the population slept under a net (33% of 0-5-year-old children), and the majority (73%) of the population slept in hammocks. Combined data pertaining to sleeping times for children and sandfly activity period indicate that > 50% of sandfly bites inside houses, and substantially more outside houses, were taken before a third of children were potentially protected by a net. This study demonstrates the clear entomological efficacy of ITNs against Lu. longipalpis in this endemic region. The effectiveness of ITNs at preventing ZVL infection and disease has still to be evaluated.

Graphical abstract

Animal African trypanosomiasis (AAT) is a significant socioeconomic burden for sub-Saharan Africa due to its huge impact on livestock health. Existing therapies including those based upon Minor Groove Binders (MGBs), such as the diamidines, which have been used for decades, have now lost efficacy in some places due to the emergence of resistant parasites. Consequently, the need for new

Trypanosoma congolense causes the most economically important animal trypanosomosis in Africa. In South Africa, a rinderpest pandemic of the 1890s removed many host animals, resulting in the near-eradication of most tsetse species. Further suppression was achieved through spraying with dichlorodiphenyltrichloroethane (DDT); however, residual populations of Glossina austeni and G. brevipalpis remained in isolated pockets. A total of 506 of these tsetse flies were captured in the Hluhluwe-iMfolozi Park, the St Lucia Wetland Park and Boomerang commercial farm. The polymerase chain reaction (PCR) was used to determine the infection rate and frequency of mixed infections of these flies. Additionally, 473 blood samples were collected from cattle at communal diptanks and a commercial farm in the area and each one examined by the haematocrit centrifugation technique (HCT). Furthermore, buffy coats from these blood samples were spotted onto FTA Elute cards and the DNA extracted from each one tested using 3 separate PCRs. The HCT revealed the presence of trypanosomes in only 6.6 % of the blood samples; by contrast, species-specific PCR detected trypanosome DNA in 50 % of the samples. The species-specific PCR detected trypanosome DNA in 17 % of the tsetse flies, compared with the nested PCR targeting rDNA which detected trypanosome DNA in only 14 % of the samples. Over time, the transmission of Savannah-type T. congolense and Kilifi-type T. congolense as mixed infections could have an impact on disease manifestation in different hosts in the area

This study details the synthesis and biological evaluation of a collection of 19 structurally related Minor Groove Binders (MGBs), derived from the natural product distamycin, which were designed to probe antifungal and antimycobacterial activity. From this initial set, we report several MGBs that are worth more detailed investigation and optimisation. MGB-4, MGB-317 and MGB-325 have promising MICs of 2, 4 and 0.25 μg/mL, respectively, against the fungus C. neoformans.MGB-353 and MGB-354 have MICs of 3.1 μM against the mycobacterium M. tuberculosis. The selectivity and activity of these compounds is related to their physicochemical properties and the cell wall/membrane characteristics of the infective agents.

African animal trypanosomosis (AAT) is caused by the tsetse fly-transmitted protozoans Trypanosoma congolense and T. vivax and leads to huge agricultural losses throughout sub-Saharan Africa. Three drugs are available to treat nagana in cattle (diminazene diaceturate, homidium chloride, and isometamidium chloride). With increasing reports of drug-resistant populations, new molecules should be investigated as potential candidates to combat nagana. Dicationic compounds have been demonstrated to have excellent efficacy against different kinetoplastid parasites. This study therefore evaluated the activities of 37 diamidines, using in vitro and ex vivo drug sensitivity assays. The 50% inhibitory concentrations obtained ranged from 0.007 to 0.562 g/ml for T. congolense and from 0.019 to 0.607 g/ml for T. vivax. On the basis of these promising results, 33 of these diamidines were further examined using in vivo mouse models of infection. Minimal curative doses of 1.25 mg/kg of body weight for both T. congolense-and T. vivax-infected mice were seen when the diamidines were administered intraperitoneally (i.p.) over 4 consecutive days. From these observations, 15 of these 33 diamidines were then further tested in vivo, using a single bolus dose for administration. The total cure of mice infected with T. congolense and T. vivax was seen with single i.p. doses of 5 and 2.5 mg/kg, respectively. This study identified a selection of diamidines which could be considered lead compounds for the treatment of nagana.

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