The Toxoplasma gondii homolog of ATPase inhibitory factor 1 is critical for mitochondrial cristae maintenance and stress response
The production of energy in the form of ATP by the mitochondrial ATP synthase must be tightly controlled. One well-conserved form of regulation is mediated via ATPase inhibitory factor 1 (IF1), which governs ATP synthase activity and gene expression patterns through a cytoprotective process known as mitohormesis. In apicomplexans, the processes regulating ATP synthase activity are not fully elucidated. Using the model apicomplexan Toxoplasma gondii, we found that knockout and overexpression of TgIF1, the structural homolog of IF1, significantly affected gene expression. Additionally, TgIF1 overexpression resulted in the formation of a stable TgIF1 oligomer and increased the presence of higher order ATP synthase oligomers. We also show that parasites lacking TgIF1 exhibit reduced mitochondrial cristae density, and that while TgIF1 levels do not affect growth in conventional culture conditions, they are crucial for parasite survival under hypoxia. Interestingly, TgIF1 overexpression enhances recovery from oxidative stress, suggesting a mitohormetic function. In summary, while TgIF1 does not appear to play a role in ATP synthase regulation under conventional growth conditions, our work uncovers its potential role in adapting to the stressors faced by T. gondii and other apicomplexans throughout their intricate life cycles.
Madelaine M Usey, Anthony A Ruberto, Kaelynn V Parker, Diego Huet. Mol Biol Cell. 2024 Nov 27:mbcE24080344. doi: 10.1091/mbc.E24-08-0344.
A pathway for skin NTD diagnostic development
The present document outlines the next steps recommended by the Skin NTDs subgroup of the DTAG to operationalise individual disease TPPs and to advance the NTD road map approach of scaling up integrated approaches to skin NTDs.
Michael Marks, Sundeep Chaitanya Vedithi, Wendy W J van de Sande, Bruno Levecke, Anthony W Solomon, Kingsley Asiedu, Camilla L Ducker, Patrick Lammie, Daniel Argaw Dagne, Isra Cruz; WHO Diagnostic Technical Advisory Group on Neglected Tropical Diseases. PLoS Negl Trop Dis. 2024 Nov 25;18(11):e0012661. doi: 10.1371/journal.pntd.0012661. eCollection 2024 Nov.
Screening the Global Health Priority Box against Plasmodium berghei liver stage parasites using an inexpensive luciferase detection protocol
Background: Malaria, a disease caused by parasites of the genus Plasmodium, continues to impact many regions globally. The rise in resistance to artemisinin-based anti-malarial drugs highlights the need for new treatments. Ideally, new anti-malarials will kill the asymptomatic liver stages as well as the symptomatic blood stages. While blood stage screening assays are routine and efficient, liver stage screening assays are more complex and costly. To decrease the cost of liver stage screening, a previously reported luciferase detection protocol requiring only common laboratory reagents was adapted for testing against luciferase-expressing Plasmodium berghei liver stage parasites.
Methods: After optimizing cell lysis conditions, the concentration of reagents, and the density of host hepatocytes (HepG2), the protocol was validated with 28 legacy anti-malarials to show this simple protocol produces a stable signal useful for obtaining quality small molecule potency data similar to that obtained from a high content imaging endpoint. The protocol was then used to screen the Global Health Priority Box (GHPB) and confirm the potency of hits in dose-response assays. Selectivity was determined using a galactose-based, 72 h HepG2 assay to avoid missing mitochondrial-toxic compounds due to the Crabtree effect. Receiver-operator characteristic plots were used to retroactively characterize the screens’ predictive value.
Results: Optimal luciferase signal was achieved using a lower HepG2 seed density (5 × 103 cells/well of a 384-well microtitre plate) compared to many previously reported luciferase-based screens. While producing lower signal compared to a commercial alternative, this luciferase detection method was found much more stable, with a > 3 h half-life, and robust enough for producing dose-response plots with as few as 500 sporozoites/well. A screen of the GHPB resulted in 9 hits with selective activity against P. berghei liver schizonts, including MMV674132 which exhibited 30.2 nM potency. Retrospective analyses show excellent predictive value for both anti-malarial activity and cytotoxicity.
Conclusions: This method is suitable for high-throughput screening at a cost nearly 20-fold less than using commercial luciferase detection kits, thereby enabling larger liver stage anti-malarial screens and hit optimization make-test cycles. Further optimization of the hits detected using this protocol is ongoing.
Gia-Bao Nguyen, Caitlin A Cooper, Olivia McWhorter, Ritu Sharma, Anne Elliot, Anthony Ruberto, Rafael Freitas, Ashutosh K Pathak, Dennis E Kyle, Steven P Maher. Malar J. 2024 Nov 23;23(1):357. doi: 10.1186/s12936-024-05155-y.
That’s NOT a Nucleus in Your Red Blood Cell – People, Parasites, & Plagues podcast with Vasant Muralidharan
Vasant Muralidharan, associate professor in Franklin College‘s Department of Cellular Biology and member of CTEGD, is the featured guest in this episode of People, Parasites, and Plagues. He discusses his work with the Plasmodium parasite and his personal experience with malaria.
β-Carboline-3-carboxamide Antimalarials: Structure-Activity Relationship, ADME-Tox Studies, and Resistance Profiling
The development of parasite resistance to both artemisinin derivatives and their partner drugs jeopardizes the effectiveness of the artemisinin combination therapy. Thus, the discovery of new antimalarial drugs, with new mechanisms of action, is urgently needed. We recently disclosed that β-carboline 1a was orally efficacious in Plasmodium berghei-infected mice and that it showed low cross-resistance between susceptible Plasmodium falciparum and four different drug-resistant strains. In this report, we describe the synthesis and in vitro antimalarial evaluation of 91 new derivatives of 1a. The asexual blood stage growth inhibition data show a clear preference for a 3,4-dihalogenated, 3,5-dihalogenated, 3,4,5-trichloro-, or 4-trifluoromethyphenyl ring at the C1-position. The most potent compound, 3,4,5-trichlorophenyl-substituted 42a, is twice as potent as 1a. Six potent analogues were assessed for their drug-like properties, and four of these were subjected to in vitro barcoded cross-resistance profiling. Compounds 1a, 1m, 42a, and 42m showed no cross-resistance to 32 resistance mutations on the Dd2 genetic background and 10 resistance mutations on the 3D7 genetic background. These data suggest that compounds in this scaffold possess a novel mechanism of antimalarial action.
Jopaul Mathew, Bo Zhou, Reagan S Haney, Kevin A Kunz, Leticia S Do Amaral, Rudraneel Roy Chowdhury, Joshua H Butler, Haibo Li, Amarraj J Chakraborty, Anika Tabassum, Emily K Bremers, Emilio F Merino, Rachael Coyle, Marcus C S Lee, Delphine Baud, Stephen Brand, Maxim Totrov, Maria Belen Cassera, Paul R Carlier. ACS Infect Dis. 2024 Oct 28. doi: 10.1021/acsinfecdis.4c00653.
A myeloid differentiation-like protein in partnership with Toll5 from the pest insect Spodoptera litura senses baculovirus infection
Many types of viruses infect insects and other arthropods. In contrast, little is known about how arthropods sense viruses, although several innate immune pathways including Toll have antiviral functions. Large DNA viruses in the family Baculoviridae are used to control a number of pest insects. Here, we studied Spodoptera litura and Autographa californica multiple nucleopolyhedrovirus (AcMNPV) to test the hypothesis that one or more myeloid differentiation-like (ML) proteins and Toll family members sense baculoviruses. We identified 11 ML and 12 Toll genes in the S. litura genome. A series of experiments indicated that S. litura ML protein 11 (SlML-11) binds the budded form of AcMNPV and partners with S. litura Toll5 (SlToll5). SlML-11 also bound sphingomyelin (SPM), which is a component of the virion envelope. Disabling SlML-11 and SlToll5 increased susceptibility to infection, while priming larvae with SPM reduced susceptibility as measured by increased survival to the adult stage and clearance of AcMNPV from individuals that emerged as adults. We conclude that SPM is a pathogen-associated molecular pattern molecule while SlML-11 and SlToll5 interact to function as a pattern recognition receptor that senses AcMNPV.
Ruonan Zhang, Jielai Zhong, Yanjun Li, Mengge Li, Jie Zhang, Qihao Hu, Liang Wen, Xiaoxia Xu, Fengliang Jin, Wanying Yang, Yuzhen Lu, Michael R Strand, Xiao-Qiang Yu. Proc Natl Acad Sci U S A. 2024 Oct 29;121(44):e2415398121. doi: 10.1073/pnas.2415398121.
ANTI-MALARIAL ACTIVITY OF AMENTOFLAVONE ISOLATED FROM LEAF OF CALOPHYLLUM TOMENTOSUM WEIGHT
Calophyllum tomentosum belonging to Clusiaceae family is an Indian medicinal plant used as folklore medicine to cure various kinds of diseases reported in Ayurveda, and the leaves of the plant are also used as an active ingredient for the preparation of a botanical medicine known as ‘Punnaga’, ‘Surapunnaga’ and ‘Tamoil’ among other common names. Chemical profiling of the methanol extract of the defatted leaf revealed the presence of amentoflavone as one of the constituents along with coumarins, terpenoids, steroids, and apetalic acids. Structural determination of these amentoflavone has been conducted by chemical, spectral, and spectrometric methods in comparison with spectral values available in the literature and confirmed by a single crystal X-ray diffraction study. Amentoflavone (1) and its derivative (2-5) tested to check the efficacy of anti-malarial activity against Plasmodium falciparum. Amongst them, only tetra methoxy amentoflavone, (2) exhibited moderate anti-malarial activity with IC50 value 1.99 ± 0.42 µM against Plasmodium falciparum in comparison with artemisinin as control, whereas the other products possessed almost negligible activity although their structural skeletons are identical with little variation of number and nature of substituents. The structure activity relationship (SAR) of the active constituent and its derivatives is reported herein.
Ajoy Kumar Bauri, Joshua H Butler, Maria B Cassera, Sabine Foro. Chem Biodivers. 2024 Oct 14:e202401576. doi: 10.1002/cbdv.202401576.
Lipophilic bisphosphonates reduced cyst burden and ameliorated hyperactivity of mice chronically infected with Toxoplasma gondii
The current treatments for toxoplasmosis are only active against fast-growing tachyzoites, present in acute infections, with little effect on slow-growing bradyzoites within tissue cysts, present in latent chronic infections. The mitochondrion of Toxoplasma gondii is essential for its survival, and one of the major anti-parasitic drugs, atovaquone, inhibits the mitochondrial electron transport chain at the coenzyme Q:cytochrome c oxidoreductase site. Coenzyme Q (also known as ubiquinone [UQ]) consists of a quinone head and a lipophilic, isoprenoid tail that anchors UQ to membranes. The synthesis of the isoprenoid unit is essential for cell growth and is inhibited by lipophilic bisphosphonates, which inhibit the parasite growth. In this work, we investigated the effect of lipophilic bisphosphonates on the chronic stages of T. gondii. We discovered that three lipophilic bisphosphonates (BPH-1218, BPH-1236, and BPH-1238), effective for the acute infection, were also effective in controlling the development of chronic stages. We showed effectiveness by testing them against in vitro cysts and in vivo derived tissue cysts and, most importantly, these compounds reduced the cyst burden in the brains of chronically infected mice. We monitored the activity of infected mice non-invasively and continuously with a novel device termed the CageDot. A decrease in activity accompanied the acute phase, but mice recovered to normal activity and showed signs of hyperactivity when the chronic infection was established. Moreover, treatment with atovaquone or BPH-1218 ameliorated the hyperactivity observed during the chronic infection.IMPORTANCETreatment for toxoplasmosis is challenged by a lack of effective drugs to eradicate the chronic stages. Most of the drugs currently used are poorly distributed to the central nervous system, and they trigger allergic reactions in a large number of patients. There is a compelling need for safe and effective treatments for toxoplasmosis. Bisphosphonates (BPs) are analogs of inorganic pyrophosphate and are used for the treatment of bone disorders. BPs target the isoprenoid pathway and are effective against several experimental parasitic infections. Some lipophilic BPs can specifically inhibit the mitochondrial activity of Toxoplasma gondii by interfering with the mechanism by which ubiquinone is inserted into the inner mitochondrial membrane. In this work, we present the effect of three lipophilic BPs against T. gondii chronic stages. We also present a new strategy for the monitoring of animal activity during disease and treatment that is non-invasive and continuous.
Melissa A Sleda, Zaid F Pitafi, WenZhan Song, Eric Oldfield, Silvia N J Moreno. mBio. 2024 Oct 10:e0175624. doi: 10.1128/mbio.01756-24
Regional changes of maximum dose insecticide responses in diamondback moth (Lepidoptera: Plutellidae) populations from Georgia and Florida, USA
Maximum dose bioassays were conducted to assess the efficacy of multiple registered active ingredients for diamondback moth (DBM), Plutella xylostella (L.), control in Georgia and Florida from 2021 to 2023 as a follow-up to an earlier study. Low efficacy (<40% mortality) was recorded for the highest labeled rate of Bacillus thuringiensis var. kurstaki strain ATBS-351 in Georgia, as well as chlorantraniliprole, cyantraniliprole, and cyclaniliprole in Georgia and Florida. The active ingredients with the highest efficacy (>80% mortality) in both states were naled, emamectin benzoate, and spinetoram. Independent analysis of data by state indicated that the efficacy of bifenthrin, chlorantraniliprole, cyantraniliprole, tolfenpyrad, and methomyl was higher in Florida populations than in Georgia populations. In addition, a comparison of these data to a recent DBM maximum dose survey in the same region suggested that these DBM populations have rapidly developed high levels of resistance to cyantraniliprole and cyclaniliprole. This work provides growers in the region with a recent ranking of insecticide efficacy that documents the loss of control for certain active ingredients, which assists pest managers in the planning of ongoing insecticide rotations for DBM resistance management.
Thomas P Dunn, Paulo S G Cremonez, Amanda Furuya, Will S Brown, Mirela M Nagaoka, Chase B Powell, Alton N Sparks Jr, Hugh Smith, David G Riley, Donald E Champagne. J Econ Entomol. 2024 Oct 10:toae218. doi: 10.1093/jee/toae218.