An X-Domain Phosphoinositide Phospholipase C (PI-PLC-like) of Trypanosoma brucei Has a Surface Localization and Is Essential for Proliferation.

Trypanosoma brucei is the causative agent of African trypanosomiasis, a deadly disease that affects humans and cattle. There are very few drugs to treat it, and there is evidence of mounting resistance, raising the need for new drug development. Here, we report the presence of a phosphoinositide phospholipase C (TbPI-PLC-like), containing an X and a PDZ domain, that is similar to the previously characterized TbPI-PLC1. TbPI-PLC-like only possesses the X catalytic domain and does not have the EF-hand, Y, and C2 domains, having instead a PDZ domain. Recombinant TbPI-PLC-like does not hydrolyze phosphatidylinositol 4,5-bisphosphate (PIP2) and does not modulate TbPI-PLC1 activity in vitro. TbPI-PLC-like shows a plasma membrane and intracellular localization in permeabilized cells and a surface localization in non-permeabilized cells. Surprisingly, knockdown of TbPI-PLC-like expression by RNAi significantly affected proliferation of both procyclic and bloodstream trypomastigotes. This is in contrast with the lack of effect of downregulation of expression of TbPI-PLC1.

Malaria prevention knowledge, attitudes, and practices in Zambezia Province, Mozambique.

BACKGROUND: In Mozambique, socio-economic and cultural factors influence the wide adoption of disease preventive measures that are relevant for malaria control strategies to promote early recognition of disease, prompt seeking of medical care, sleeping under insecticide-treated nets (ITNs), and taking intermittent preventive treatment for pregnant women. However, there is a critical information gap regarding previous and ongoing malaria social and behavioural change (SBC) interventions. The aim of this study is to assess the knowledge, attitudes, practices of beneficiaries of SBC interventions. METHODS: A descriptive cross-sectional survey was undertaken in 2018 in two rural districts of Zambezia Province, Mozambique. A structured questionnaire was administered to 773 randomly selected households. Respondents were the adult heads of the households. Descriptive statistics were done. RESULTS: The main results show that 96.4% of respondents recalled hearing about malaria in the previous 6 months, 90.0% had knowledge of malaria prevention, and 70.0% of preventive measures. Of the 97.7% respondents that had received ITNs through a mass ITN distribution campaign, 81.7% had slept under an ITN the night before the survey. In terms of source of health information, 70.5% mentioned the role of community volunteers in dissemination of malaria prevention messages, 76.1% of respondents considered worship places (churches and mosques) to be the main places where they heard key malaria prevention messages, and 79.1% asserted that community dialogue sessions helped them better understand how to prevent malaria. CONCLUSIONS: Results show that volunteers/activists/teachers played an important role in dissemination of key malaria prevention messages, which brought the following successes: community actors are recognized and people have knowledge of malaria transmission, signs and symptoms, preventive measures, and where to get treatment. There is, however, room for improvement on SBC messaging regarding some malaria symptoms (anaemia and convulsions) and operational research is needed to ascertain the drivers of malaria prevalence and inform the SBC approach.

Malaria prevention knowledge, attitudes, and practices in Zambezia Province, Mozambique

Background: In Mozambique, socio-economic and cultural factors influence the wide adoption of disease preventive measures that are relevant for malaria control strategies to promote early recognition of disease, prompt seeking of medical care, sleeping under insecticide-treated nets (ITNs), and taking intermittent preventive treatment for pregnant women. However, there is a critical information gap regarding previous and ongoing malaria social and behavioural change (SBC) interventions. The aim of this study is to assess the knowledge, attitudes, practices of beneficiaries of SBC interventions. Methods: A descriptive cross-sectional survey was undertaken in 2018 in two rural districts of Zambezia Province, Mozambique. A structured questionnaire was administered to 773 randomly selected households. Respondents were the adult heads of the households. Descriptive statistics were done. Results: The main results show that 96.4% of respondents recalled hearing about malaria in the previous 6 months, 90.0% had knowledge of malaria prevention, and 70.0% of preventive measures. Of the 97.7% respondents that had received ITNs through a mass ITN distribution campaign, 81.7% had slept under an ITN the night before the survey. In terms of source of health information, 70.5% mentioned the role of community volunteers in dissemination of malaria prevention messages, 76.1% of respondents considered worship places (churches and mosques) to be the main places where they heard key malaria prevention messages, and 79.1% asserted that community dialogue sessions helped them better understand how to prevent malaria. Conclusions: Results show that volunteers/activists/teachers played an important role in dissemination of key malaria prevention messages, which brought the following successes: community actors are recognized and people have knowledge of malaria transmission, signs and symptoms, preventive measures, and where to get treatment. There is, however, room for improvement on SBC messaging regarding some malaria symptoms (anaemia and convulsions) and operational research is needed to ascertain the drivers of malaria prevalence and inform the SBC approach.

Synthesis and in vitro evaluation of new 5-substituted 6-nitroimidazooxazoles as antikinetoplastid agents.

In continuation of our pharmacomodulation work on the nitroimidazooxazole series, we report the synthesis of new 5-substituted 6-nitroimidazooxazole derivatives. Our aim was to evaluate how functionalization of the 5-position of the 6-nitroimidazooxazole scaffold affects antileishmanial and antitrypanosomal in vitro activities. Twenty-one original compounds were synthesized and evaluated for their in vitro antileishmanial (L. donovani) and antitrypanosomal (T. cruzi) properties. Pallado-catalyzed cross-coupling reactions were used to introduce an aryl or ethynyl aryl substituent in 5-position from a 5-brominated-6-nitroimidazooxazole starting product. Unfortunately, the first series of compounds bearing an aryl group in 5-position presented limited in vitro activities against L. donovani and T. cruzi, with IC50 > 10 µM (vs 0.18 µM and 2.31 µM for the reference drugs amphotericin B and benznidazole respectively). Interestingly, the second series of compounds bearing an ethynyl aryl substituent in 5-position showed more promising, particularly against T. cruzi. Compounds 6a, 6b, 6c, 6g and 6h had better activity than the reference drug benznidazole (0.92 µM ≤ IC50 ≤ 2.18 µM vs IC50 = 2.31 µM), whereas the non-functionalized 2-methyl-6-nitro-2,3-dihydroimidazo [2,1-b]oxazole 2 was not active against T. cruzi (IC50 > 10 µM).

The acidocalcisome inositol-1,4,5-trisphosphate receptor of Trypanosoma brucei is stimulated by luminal polyphosphate hydrolysis products.

Acidocalcisomes are acidic calcium stores rich in polyphosphate (polyP) and are present in trypanosomes and also in a diverse range of other organisms. Ca2+ is released from these organelles through a channel, inositol 1,4,5-trisphosphate receptor (TbIP3R), which is essential for growth and infectivity of the parasite Trypanosoma brucei However, the mechanism by which TbIP3R controls Ca2+ release is unclear. In this work, we expressed TbIP3R in a chicken B lymphocyte cell line in which the genes for all three vertebrate IP3Rs were stably ablated (DT40-3KO). We show that IP3-mediated Ca2+ release depends on Ca2+ but not on ATP concentration and is inhibited by heparin, caffeine, and 2-aminomethoxydiphenyl borate (2-APB). Excised patch clamp recordings from nuclear membranes of DT40 cells expressing only TbIP3R disclosed that luminal inorganic orthophosphate (Pi) or pyrophosphate (PPi), and neutral or alkaline pH can stimulate IP3-generated currents. In contrast, polyP or acidic pH did not induce these currents, and nuclear membranes obtained from cells expressing rat IP3R were unresponsive to polyP or its hydrolysis products. Our results are consistent with the notion that polyP hydrolysis products within acidocalcisomes or alkalinization of their luminal pH activate TbIP3R and Ca2+ release. We conclude that TbIP3R is well-adapted to its role as the major Ca2+ release channel of acidocalcisomes in T. brucei.

Prevention of palatable diet-induced hyperphagia in rats by central injection of a VEGFR kinase inhibitor.

Our previous studies have demonstrated a critical role of a VEGFR-like signaling pathway in hunger-driven overeating of sugar-rich food in Drosophila larvae. In the current study, we investigate whether the VEGFR signaling mechanism plays a similar role in the feeding behavior of vertebrates using male Sprague Dawley rats. Young rats were treated intracerebroventrically (i.c.v.) with a single dose (2 µg) of VEGFR2 Tyrosine Kinase Inhibitor V (VTKI-V), an N-cyclopropylnaphthamide compound that selectively inhibits the kinase activity of VEGFR2 at subnanomolar concentrations. We find that animals treated with VTKI-V showed markedly attenuated overconsumption of palatable food that is sweet and fatty. The subsequent meal pattern analysis reveals that is achieved by consumption of smaller, shorter meals. Furthermore, i.c.v. injection of VTKI-V decreased body weight gain in animals fed with CHOW or palatable food. These inhibitory effects of the drug were detectable within 24h and persisted for at least five days. Given that body weight was affected by the drug regardless of diet while food intake was selectively altered in palatable diet fed animals, these results raise the possibility that i.c.v. injection of VTKI-V may interfere the functions of two separate VEGFR-mediated mechanisms: one promotes overconsumption of palatable food, and the other mediates body weight gain.

Duplication of the Asymmetric Leaves1/Rough Sheath 2/Phantastica (ARP) gene precedes the explosive radiation of the Ruschioideae.

The Mesembryanthemoideae and Ruschioideae subfamilies are a major component of the Greater Cape Floristic Region in southern Africa. The Ruschioideae show an astonishing diversity of leaf shape and growth forms. Although 1,585 species are recognised within the morphologically diverse Ruschioideae, these species show minimal variation in plastid DNA sequence. We have investigated whether changes in selected leaf development transcription factors underpin the recent, rapid diversification of this large group of succulent plants. Degenerate primers designed to conserved regions of Asymmetric Leaves1/Rough Sheath 2/Phantastica (ARP) and the Class III HD-ZIP family of genes, were used to amplify sequences corresponding to these genes from several species within the Mesembryanthemoideae and Ruschioideae subfamilies. Two members of the Class III HD-ZIP family were identified in both the Mesembryanthemoideae and Ruschioideae, and were derived from an ancient gene duplication event that preceded the divergence of gymnosperms and angiosperms. While a single ARP orthologue was identified in the Mesembryanthemoideae, two paralogues, ARPa and ARPb, were identified in the Ruschioideae subfamily. ARPa was present in all species of Ruschioideae analysed in this study. ARPb has been lost from the Apatesieae and Dorotheantheae tribes, which form an early evolutionary branch from the Ruschieae tribe, as well as from selected species within the Ruschieae. The recent duplication and subsequent selected gene loss of the ARP transcription factor correlates with the rapid diversification of plant forms in the Ruschioideae.