Mosquito immune responses and compatibility between Plasmodium parasites and anopheline mosquitoes

<p>Abstract</p> <p>Background</p> <p>Functional screens based on dsRNA-mediated gene silencing identified several <it>Anopheles gambiae </it>genes that limit <it>Plasmodium berghei </it>infection. However, some of the genes identified in these screens have no effect on the human malaria parasite <it>Plasmodium falciparum</it>; raising the question of whether different mosquito effector genes mediate anti-parasitic responses to different <it>Plasmodium </it>species.</p> <p>Results</p> <p>Four new <it>An. gambiae </it>(G3) genes were identified that, when silenced, have a different effect on <it>P. berghei </it>(Anka 2.34) and <it>P. falciparum </it>(3D7) infections. Orthologs of these genes, as well as <it>LRIM1 </it>and <it>CTL4</it>, were also silenced in <it>An. stephensi </it>(Nijmegen Sda500) females infected with <it>P. yoelii </it>(17XNL). For five of the six genes tested, silencing had the same effect on infection in the <it>P. falciparum-An. gambiae </it>and <it>P. yoelii-An. stephensi </it>parasite-vector combinations. Although silencing <it>LRIM1 </it>or <it>CTL4 </it>has no effect in <it>An. stephensi </it>females infected with <it>P. yoelii</it>, when <it>An. gambiae </it>is infected with the same parasite, silencing these genes has a dramatic effect. In <it>An. gambiae </it>(G3), TEP1, LRIM1 or LRIM2 silencing reverts lysis and melanization of <it>P. yoelii</it>, while <it>CTL4 </it>silencing enhances melanization.</p> <p>Conclusion</p> <p>There is a broad spectrum of compatibility, the extent to which the mosquito immune system limits infection, between different <it>Plasmodium </it>strains and particular mosquito strains that is mediated by TEP1/LRIM1 activation. The interactions between highly compatible animal models of malaria, such as <it>P. yoelii </it>(17XNL)-<it>An. stephensi </it>(Nijmegen Sda500), is more similar to that of <it>P. falciparum </it>(3D7)-<it>An. gambiae </it>(G3).</p

Transcriptional analysis of an immune-responsive serine protease from Indian malarial vector, Anopheles culicifacies

<p>Abstract</p> <p>Background</p> <p>The main vector for transmission of malaria in India is the <it>Anopheles culicifacies </it>mosquito species, a naturally selected subgroup of which is completely refractory (R) to transmission of the malaria parasite, <it>Plasmodium vivax</it>;</p> <p>Results</p> <p>Here, we report the molecular characterization of a serine protease (<it>acsp30</it>)-encoding gene from <it>A. culicifacies</it>, which was expressed in high abundance in the refractory strain compared to the susceptible (S) strain. The transcriptional upregulation of <it>acsp30 </it>upon <it>Plasmodium </it>challenge in the refractory strain coincided with ookinete invasion of mosquito midgut. Gene organization and primary sequence of <it>acsp30 </it>were identical in the R and S strains suggesting a divergent regulatory status of <it>acsp30 </it>in these strains. To examine this further, the upstream regulatory sequences of <it>acsp30 </it>were isolated, cloned and evaluated for the presence of promoter activity. The 702 bp upstream region of <it>acsp30 </it>from the two strains revealed sequence divergence. The promoter activity measured by luciferase-based reporter assay was shown to be 1.5-fold higher in the R strain than in the S. Gel shift experiments demonstrated a differential recruitment of nuclear proteins to upstream sequences of <it>acsp30 </it>as well as a difference in the composition of nuclear proteins in the two strains, both of which might contribute to the relative abundance of <it>acsp30 </it>in the R strain;</p> <p>Conclusion</p> <p>The specific upregulation of <it>acsp30 </it>in the R strain only in response to <it>Plasmodium </it>infection is suggestive of its role in contributing the refractory phenotype to the <it>A. culicifacies </it>mosquito population.</p

Hundreds of dual-stage antimalarial molecules discovered by a functional gametocyte screen.

Plasmodium falciparum stage V gametocytes are responsible for parasite transmission, and drugs targeting this stage are needed to support malaria elimination. We here screen the Tres Cantos Antimalarial Set (TCAMS) using the previously developed P. falciparum female gametocyte activation assay (Pf FGAA), which assesses stage V female gametocyte viability and functionality using Pfs25 expression. We identify over 400 compounds with activities <2 μM, chemically classified into 57 clusters and 33 singletons. Up to 68% of the hits are chemotypes described for the first time as late-stage gametocyte-targeting molecules. In addition, the biological profile of 90 compounds representing the chemical diversity is assessed. We confirm in vitro transmission-blocking activity of four of the six selected molecules belonging to three distinct scaffold clusters. Overall, this TCAMS gametocyte screen provides 276 promising antimalarial molecules with dual asexual/sexual activity, representing starting points for target identification and candidate selection

An Epithelial Serine Protease, AgESP, Is Required for Plasmodium Invasion in the Mosquito Anopheles gambiae

Background: Plasmodium parasites need to cross the midgut and salivary gland epithelia to complete their life cycle in the mosquito. However, our understanding of the molecular mechanism and the mosquito genes that participate in this process is still very limited. Methodology/Principal Findings: We identified an Anopheles gambiae epithelial serine protease (AgESP) that is constitutively expressed in the submicrovillar region of mosquito midgut epithelial cells and in the basal side of the salivary glands that is critical for Plasmodium parasites to cross these two epithelial barriers. AgESP silencing greatly reduces Plasmodium berghei and Plasmodium falciparum midgut invasion and prevents the transcriptional activation of gelsolin, a key regulator of actin remodeling and a reported Plasmodium agonist. AgESP expression is highly induced in midgut cells invaded by Plasmodium, suggesting that this protease also participates in the apoptotic response to invasion. In salivary gland epithelial cells, AgESP is localized on the basal side–the surface with which sporozoites interact. AgESP expression in the salivary gland is also induced in response to P. berghei and P. falciparum sporozoite invasion, and AgESP silencing significantly reduces the number of sporozoites that invade this organ. Conclusion: Our findings indicate that AgESP is required for Plasmodium parasites to effectively traverse the midgut and salivary gland epithelial barriers. Plasmodium parasites need to modify the actin cytoskeleton of mosquito epithelial cells t

A tetraoxane-based antimalarial drug candidate that overcomes PfK13-C580Y dependent artemisinin resistance.

K13 gene mutations are a primary marker of artemisinin resistance in Plasmodium falciparum malaria that threatens the long-term clinical utility of artemisinin-based combination therapies, the cornerstone of modern day malaria treatment. Here we describe a multinational drug discovery programme that has delivered a synthetic tetraoxane-based molecule, E209, which meets key requirements of the Medicines for Malaria Venture drug candidate profiles. E209 has potent nanomolar inhibitory activity against multiple strains of P. falciparum and P. vivax in vitro, is efficacious against P. falciparum in in vivo rodent models, produces parasite reduction ratios equivalent to dihydroartemisinin and has pharmacokinetic and pharmacodynamic characteristics compatible with a single-dose cure. In vitro studies with transgenic parasites expressing variant forms of K13 show no cross-resistance with the C580Y mutation, the primary variant observed in Southeast Asia. E209 is a superior next generation endoperoxide with combined pharmacokinetic and pharmacodynamic features that overcome the liabilities of artemisinin derivatives

Potent Plasmodium falciparum gametocytocidal compounds identified by exploring the kinase inhibitor chemical space for dual active antimalarials

OBJECTIVES : Novel chemical tools to eliminate malaria should ideally target both the asexual parasites and transmissible gametocytes. Several imidazopyridazines (IMPs) and 2-aminopyridines (2-APs) have been described as potent antimalarial candidates targeting lipid kinases. However, these have not been extensively explored for stage-specific inhibition of gametocytes in Plasmodium falciparum parasites. Here we provide an in-depth evaluation of the gametocytocidal activity of compounds from these chemotypes and identify novel starting points for dual-acting antimalarials. METHODS : We evaluated compounds against P. falciparum gametocytes using several assay platforms for cross-validation and stringently identified hits that were further profiled for stage specificity, speed of action and ex vivo efficacy. Physicochemical feature extraction and chemogenomic fingerprinting were applied to explore the kinase inhibition susceptibility profile. RESULTS : We identified 34 compounds with submicromolar activity against late stage gametocytes, validated across several assay platforms. Of these, 12 were potent at 1000-fold selectivity towards the parasite over mammalian cells. Front-runner compounds targeted mature gametocytes within 48 h and blocked transmission to mosquitoes. The resultant chemogenomic fingerprint of parasites treated with the lead compounds revealed the importance of targeting kinases in asexual parasites and gametocytes. CONCLUSIONS : This study encompasses an in-depth evaluation of the kinase inhibitor space for gametocytocidal activity. Potent lead compounds have enticing dual activities and highlight the importance of targeting the kinase superfamily in malaria elimination strategies.The South African Medical Research Council (SAMRC) Self-initiated Research (to JN) and Strategic Health Initiatives Partnerships (MRC-SHIP) programmes to L.B., T.C., D.M. K.C. further acknowledges the SAMRC for funding of the extramural Drug Discovery and Development Research Unit at UCT. The SAMRC is acknowledged for funding of the UP ISMC (LMB) and WRIM (TLC) as Collaborating Centres for Malaria Research. The Council for Scientific and Industrial Research and the 3R Foundation (project 118–10) to D.M. We thank the Medicines for Malaria Venture and South African Technology Innovation Agency (TIA) for funding to K.C. (Project MMV09/0002). The University of Cape Town, University of Pretoria, and South African Research Chairs Initiative of the Department of Science and Technology, administered through the South African National Research Foundation are gratefully acknowledged for support to K.C. and L.B. (UID84627). JN was supported through an International Society for Infectious Diseases grant.https://academic.oup.com/jac2019-05-01hj2018Biochemistr

Leave no one behind: response to new evidence and guidelines for the management of cryptococcal meningitis in low-income and middle-income countries

In 2018, WHO issued guidelines for the diagnosis, prevention, and management of HIV-related cryptococcal disease. Two strategies are recommended to reduce the high mortality associated with HIV-related cryptococcal meningitis in low-income and middle-income countries (LMICs): optimised combination therapies for confirmed meningitis cases and cryptococcal antigen screening programmes for ambulatory people living with HIV who access care. WHO's preferred therapy for the treatment of HIV-related cryptococcal meningitis in LMICs is 1 week of amphotericin B plus flucytosine, and the alternative therapy is 2 weeks of fluconazole plus flucytosine. In the ACTA trial, 1-week (short course) amphotericin B plus flucytosine resulted in a 10-week mortality of 24% (95% CI −16 to 32) and 2 weeks of fluconazole and flucytosine resulted in a 10-week mortality of 35% (95% CI −29 to 41). However, with widely used fluconazole monotherapy, mortality because of HIV-related cryptococcal meningitis is approximately 70% in many African LMIC settings. Therefore, the potential to transform the management of HIV-related cryptococcal meningitis in resource-limited settings is substantial. Sustainable access to essential medicines, including flucytosine and amphotericin B, in LMICs is paramount and the focus of this Personal View

Isolation and Characterization of a marine bacterium belonging to the genus <i style="">Alkaligenes </i>capable of the complete mineralization of the dibenzothiophene

391-397Following enrichment on benzoate five bacterial isolates capable of aerobic growth on dibenzothiophene were obtained from sea water collected off the west coast of India near the city of Goa. Sampling location had been previously contaminated with furnace oil spilled from a tanker. Classified as marine bacteria by their requirement for and optimum growth on 3% NaCl, all the isolates were identified by morphological observations and biochemical tests as belonging to the genes Alkaligenes. A single isolates producing intense red pigmented colony when growing on dibenzothiophene was chosen for further studies and designated as Strain JR110. During catabolism of dibenzothiophene intermediates were observed after initial dissolution and complete mineralization was achieved. JR110, independently confirmed as belonging to the genus Alkaligenes by Im Tech, India was deposited in their culture collection as MTCC3317

Fighting Shigella by Blocking Its Disease-Causing Toxin

Shiga toxin is an AB5 toxin produced by Shigella species, while related toxins are produced by Shiga toxin-producing Escherichia coli (STEC). Infection by Shigella can lead to bloody diarrhea followed by the often fatal hemolytic uremic syndrome (HUS). In the present paper, we aimed for a simple and effective toxin inhibitor by comparing three classes of carbohydrate-based inhibitors: glycodendrimers, glycopolymers, and oligosaccharides. We observed a clear enhancement in potency for multivalent inhibitors, with the divalent and tetravalent compounds inhibiting in the millimolar and micromolar range, respectively. However, the polymeric inhibitor based on galabiose was the most potent in the series exhibiting nanomolar inhibition. Alginate and chitosan oligosaccharides also inhibit Shiga toxin and may be used as a prophylactic drug during shigella outbreaks