Background: Sporozoites isolated from the salivary glands of Plasmodium-infected mosquitoes are a prerequisite for several basic and pre-clinical applications. Although salivary glands are pooled to maximize sporozoite recovery, insufficient yields pose logistical and analytical hurdles; thus, predicting yields prior to isolation would be valuable. Preceding oocyst densities in the midgut is an obvious candidate. However, it is unclear whether current understanding of its relationship with sporozoite densities can be used to maximize yields, or whether it can capture the potential density-dependence in rates of sporozoite invasion of the salivary glands.
Methods: This study presents a retrospective analysis of Anopheles stephensi mosquitoes infected with two strains of the rodent-specific Plasmodium berghei. Mean oocyst densities were estimated in the midguts earlier in the infection (11-15 days post-blood meal), with sporozoites pooled from the salivary glands later in the infection (17-29 days). Generalized linear mixed effects models were used to determine if (1) mean oocyst densities can predict sporozoite yields from pooled salivary glands, (2) whether these densities can capture differences in rates of sporozoite invasion of salivary glands, and (3), if the interaction between oocyst densities and time could be leveraged to boost overall yields.
Results: The non-linear effect of mean oocyst densities confirmed the role of density-dependent constraints in limiting yields beyond certain oocyst densities. Irrespective of oocyst densities however, the continued invasion of salivary glands by the sporozoites boosted recoveries over time (17-29 days post-blood meal) for either parasite strain.
Conclusions: Sporozoite invasion of the salivary glands over time can be leveraged to maximize yields for P. berghei. In general, however, invasion of the salivary glands over time is a critical fitness determinant for all Plasmodium species (extrinsic incubation period, EIP). Thus, delaying sporozoite collection could, in principle, substantially reduce dissection effort for any parasite within the genus, with the results also alluding to the potential for changes in sporozoites densities over time to modify infectivity for the next host.
The skeletal muscle of a mouse infected with Trypanosoma cruzi is shown under a microscope. (Submitted by Fernando Sanchez)
The condition affects tens of millions across the Americas but lacks effective treatments
Researchers from the University of Georgia have discovered a potential treatment for Chagas disease, marking the first medication with promise to successfully and safely target the parasitic infection in more than 50 years.
Human clinical trials of the drug, an antiparasitic compound known as AN15368, will hopefully begin in the next few years.
Distinguished Research Professor Dr. Rick Tarleton of the Center for Tropical and Emerging Global Diseases at the Paul D. Coverdell Center for Biomedical and Health Sciences on Thursday, May 8, 2008. Dr. Tarleton is researching the effects of drug treatments on Chagas’ disease.
“I’m very optimistic,” said Rick Tarleton, corresponding author of the study and a UGA Athletic Association Distinguished Professor in the Franklin College of Arts and Sciences and member of the Center for Tropical and Emerging Global Diseases. “I think it has a really strong chance of being a real solution, not just a stand-in for something that works better than the drugs we currently have.”
The new drug works by targeting the parasite that causes the disease, Trypanosoma cruzi, also known as T. cruzi.
Nearly all people infected with the parasite experience flu-like symptoms such as fever, headaches and vomiting. However, after their immune response kicks in, their symptoms may subside.
But for 30% to 40% of patients, the infection can result in severe heart damage that can be both debilitating and life-threatening.
New drug is 100% effective in eliminating T. cruzi
Published in Nature Microbiology, the study found the new medication was 100% effective in curing mice, as well as non-human primates that were naturally infected by the parasite at a research facility in Texas. The animals also experienced no significant side effects from exposure to the drug.
Over the past several decades, previous treatment candidates went straight from experimental infections in mice to human clinical trials, where they failed to cure the infection. The new drug’s efficacy in non-human primates bodes well for how it will perform in humans.
“We’ve got something that is as close to effective as it can be in what is as close to a human as it could be, and there aren’t any side effects. That really de-risks it by a lot going into humans,” Tarleton said. “It doesn’t make it fail-safe, but it moves it much further along.”
Current medications to treat T. cruzi infection not ideal
T. cruzi is carried by blood-sucking insects known as kissing bugs. The insects can be found throughout North, Central and South America.
In addition to a nasty bite, the creatures carry the T. cruzi parasite, which is transmitted through their fecal matter. Victims can become infected when they unknowingly rub the insect’s feces into their eyes, nose or an open wound.
The infection may also be transmitted through organ transplants, from a pregnant person to their fetus or through contaminated food. However, infections from these pathways are less common.
The go-to medications used to treat Chagas aren’t terrible, Tarleton said, but they’re not ideal. They can pack some serious side effects and they’re not reliably effective, but they’re currently the only treatment option.
Patients also have to take the drugs for two months. And even the common but mild side effects like headache or nausea get old after a few weeks. As a result, about one in five people being treated for the disease stop taking their medications before they have a chance to cure the infection.
“Plus they have variable efficacy, and it’s not predictable,” Tarleton said. “I think most physicians in Latin America have to say, ‘We have a drug. It’s going to make you feel bad, and two months later after we finish it, we’re not really going to be able to tell you if it worked or not.’
“It’s really not a good inducement to take the medication.”
Chagas disease common in Latin American countries
Tens of millions of people across the Americas are infected with the parasite that causes Chagas disease. But it doesn’t get much media attention.
It’s most common in Latin American countries, particularly in low-income areas where housing isn’t ideal. Some of the countries with the highest rates of the disease include Bolivia, Venezuela, Argentina, Chile, Mexico and Brazil.
In homes with thatched roofs, mud walls or inadequate protection from the elements, kissing bugs thrive, making infection more likely.
Chagas disease poses significant risk to pets
The Centers for Disease Control and Prevention estimates around 300,000 people infected with the parasite currently live in the U.S. But because the condition isn’t a huge threat in places with good housing options, Chagas disease treatment and prevention doesn’t get much research funding.
There is growing concern about the T. cruzi infection rate among outdoor pets in the U.S., however. Working dogs and other pets that spend extended periods of time outside are contracting the parasite at an alarming rate.
“There are areas where the infection rates are 20% to 30% new infections per year,” Tarleton said. “Those tend to be severe infections where the dogs either die or develop a disease that makes them unable to work.”
Tarleton hopes to partner with veterinary pharmaceutical companies in the future to create a drug to treat the infection in pets as a means of funding diagnostics and medication purchases in Latin America.
For the present study, Tarleton partnered with colleagues at Anacor Pharmaceuticals, Texas A&M University, the University of Texas, the University of Kansas and Pfizer. Angel Padilla, Wei Wang, Dylan Orr, Brooke White, Arlene George and Huifeng Shen from UGA’s Center for Tropical and Emerging Global Diseases and the Department of Cellular Biology are co-authors on the paper.
Rick Tarleton and colleagues recently published their new study, “Discovery of an orally active benzoxaborole prodrug effective in the treatment of Chagas disease in non-human primates” in Nature Microbiology. Check out these news stories about the study.
Behind the paper: New Hope for Treatment of a Very Neglected, Neglected Tropical Disease (Nature Microbiology)
Possible new treatment identified for neglected tropical disease (Science.org)
Researchers discover potential treatment for Chagas disease (Mirage News)
Researchers discover potential treatment for Chagas disease (Science Daily)
Researchers discover potential treatment for Chagas disease (Medical Xpress)
Chagas disease potential treatment: ‘I think it has a really strong chance of being a real solution’ (Outbreak News Today)
Promising New Drug May Effectively Treat Chagas Disease (Technology Networks)
Researchers discover potential treatment for Chagas disease (Newswise)
Trypanosoma cruzi, the agent of Chagas disease, probably infects tens of millions of people, primarily in Latin America, causing morbidity and mortality. The options for treatment and prevention of Chagas disease are limited and underutilized. Here we describe the discovery of a series of benzoxaborole compounds with nanomolar activity against extra- and intracellular stages of T. cruzi. Leveraging both ongoing drug discovery efforts in related kinetoplastids, and the exceptional models for rapid drug screening and optimization in T. cruzi, we have identified the prodrug AN15368 that is activated by parasite carboxypeptidases to yield a compound that targets the messenger RNA processing pathway in T. cruzi. AN15368 was found to be active in vitro and in vivo against a range of genetically distinct T. cruzi lineages and was uniformly curative in non-human primates (NHPs) with long-term naturally acquired infections. Treatment in NHPs also revealed no detectable acute toxicity or long-term health or reproductive impact. Thus, AN15368 is an extensively validated and apparently safe, clinically ready candidate with promising potential for prevention and treatment of Chagas disease.
Angel M. Padilla, Wei Wang, Tsutomu Akama, David S. Carter, Eric Easom, Yvonne Freund, Jason S. Halladay, Yang Liu, Sarah A. Hamer, Carolyn L. Hodo, Gregory K. Wilkerson, Dylan Orr, Brooke White, Arlene George, Huifeng Shen, Yiru Jin, Michael Zhuo Wang, Susanna Tse, Robert T. Jacobs & Rick L. Tarleton. Nat Microbiol (2022). https://doi.org/10.1038/s41564-022-01211-y
Anautogenous mosquitoes must blood feed on a vertebrate host to produce eggs. Each gonadotrophic cycle is subdivided into a sugar-feeding previtellogenic phase that produces primary follicles and a blood meal-activated vitellogenic phase in which large numbers of eggs synchronously mature and are laid. Multiple endocrine factors including juvenile hormone (JH), insulin-like peptides (ILPs), ovary ecdysteroidogenic hormone (OEH) and 20-hydroxyecdysone (20E) coordinate each gonadotrophic cycle. Egg formation also requires nutrients from feeding that are stored in the fat body. Regulation of egg formation is best understood in Aedes aegypti but the role different endocrine factors play in regulating nutrient mobilization and storage remains unclear. In this study, we report that adult female Ae. aegypti maintained triacylglycerol (TAG) stores during the previtellogenic phase of the first gonadotrophic cycle while glycogen stores declined. In contrast, TAG and glycogen stores were rapidly mobilized during the vitellogenic phase and then replenishment. Several genes encoding enzymes with functions in TAG and glycogen metabolism were differentially expressed in the fat body, which suggested regulation was mediated in part at the transcriptional level. Gain of function assays indicated that stored nutrients were primarily mobilized by adipokinetic hormone (AKH) while juvenoids and OEH regulated replenishment. ILP3 further showed evidence of negatively regulating certain lipolytic enzymes. Loss of function assays further indicated AKH depends on the AKH receptor (AKHR) for function. Altogether, our results indicate that the opposing activities of different hormones regulate nutrient stores during a gonadotrophic cycle in Ae. aegypti. This article is protected by copyright. All rights reserved.
The resilience of Plasmodium vivax, the most widely-distributed malaria-causing parasite in humans, is attributed to its ability to produce dormant liver forms known as hypnozoites, which can activate weeks, months, or even years after an initial mosquito bite. The factors underlying hypnozoite formation and activation are poorly understood, as is the parasite’s influence on the host hepatocyte. Here, we shed light on transcriptome-wide signatures of both the parasite and the infected host cell by sequencing over 1,000 P. vivax-infected hepatocytes at single-cell resolution. We distinguish between replicating schizonts and hypnozoites at the transcriptional level, identifying key differences in transcripts encoding for RNA-binding proteins associated with cell fate. In infected hepatocytes, we show that genes associated with energy metabolism and antioxidant stress response are upregulated, and those involved in the host immune response downregulated, suggesting both schizonts and hypnozoites alter the host intracellular environment. The transcriptional markers in schizonts, hypnozoites, and infected hepatocytes revealed here pinpoint potential factors underlying dormancy and can inform therapeutic targets against P. vivax liver-stage infection.
Anthony A Ruberto, Steven P Maher, Amélie Vantaux, Chester J Joyner, Caitlin Bourke, Balu Balan, Aaron Jex, Ivo Mueller, Benoit Witkowski, Dennis E Kyle. Front Cell Infect Microbiol. 2022 Aug 25;12:986314. doi: 10.3389/fcimb.2022.986314. eCollection 2022.
Apicomplexan parasites cause persistent mortality and morbidity worldwide through diseases including malaria, toxoplasmosis, and cryptosporidiosis. Ca2+ signaling pathways have been repurposed in these eukaryotic pathogens to regulate parasite-specific cellular processes governing the replicative and lytic phases of the infectious cycle, as well as the transition between them. Despite the presence of conserved Ca2+-responsive proteins, little is known about how specific signaling elements interact to impact pathogenesis. We mapped the Ca2+-responsive proteome of the model apicomplexan T. gondii via time-resolved phosphoproteomics and thermal proteome profiling. The waves of phosphoregulation following PKG activation and stimulated Ca2+ release corroborate known physiological changes but identify specific proteins operating in these pathways. Thermal profiling of parasite extracts identified many expected Ca2+-responsive proteins, such as parasite Ca2+-dependent protein kinases. Our approach also identified numerous Ca2+-responsive proteins that are not predicted to bind Ca2+, yet are critical components of the parasite signaling network. We characterized protein phosphatase 1 (PP1) as a Ca2+-responsive enzyme that relocalized to the parasite apex upon Ca2+ store release. Conditional depletion of PP1 revealed that the phosphatase regulates Ca2+ uptake to promote parasite motility. PP1 may thus be partly responsible for Ca2+-regulated serine/threonine phosphatase activity in apicomplexan parasites.
The proposed model of how host gut microbiota promotes parasite survival. (Figure 6)
Studying the microbial symbionts of eukaryotic hosts has revealed a range of interactions that benefit host biology. Most eukaryotes are also infected by parasites that adversely affect host biology for their own benefit. However, it is largely unclear whether the ability of parasites to develop in hosts also depends on host-associated symbionts, e.g., the gut microbiota. Here, we studied the parasitic wasp Leptopilina boulardi (Lb) and its host Drosophila melanogaster. Results showed that Lb successfully develops in conventional hosts (CN) with a gut microbiota but fails to develop in axenic hosts (AX) without a gut microbiota. We determined that developing Lb larvae consume fat body cells that store lipids. We also determined that much larger amounts of lipid accumulate in fat body cells of parasitized CN hosts than parasitized AX hosts. CN hosts parasitized by Lb exhibited large increases in the abundance of the bacterium Acetobacter pomorum in the gut, but did not affect the abundance of Lactobacillus fructivorans which is another common member of the host gut microbiota. However, AX hosts inoculated with A. pomorum and/or L. fructivorans did not rescue development of Lb. In contrast, AX larvae inoculated with A. pomorum plus other identified gut community members including a Bacillus sp. substantially rescued Lb development. Rescue was further associated with increased lipid accumulation in host fat body cells. Insulin-like peptides increased in brain neurosecretory cells of parasitized CN larvae. Lipid accumulation in the fat body of CN hosts was further associated with reduced Bmm lipase activity mediated by insulin/insulin-like growth factor signaling (IIS). Altogether, our results identify a previously unknown role for the gut microbiota in defining host permissiveness for a parasite. Our findings also identify a new paradigm for parasite manipulation of host metabolism that depends on insulin signaling and the gut microbiota.
Sicong Zhou, Yueqi Lu, Jiani Chen, Zhongqiu Pan, Lan Pang, Ying Wang, Qichao Zhang, Michael R Strand, Xue-Xin Chen, Jianhua Huang. ISME J. 2022 Aug 8. doi: 10.1038/s41396-022-01301-z.
In June 2021, the World Health Organization (WHO), recognizing the need for new diagnostics to support the control and elimination of onchocerciasis, published the target product profiles (TPPs) of new tests that would support the two most immediate needs: (a) mapping onchocerciasis in areas of low prevalence and (b) deciding when to stop mass drug administration programs. In both instances, the test should ideally detect an antigen specific for live, adult O. volvulus female worms. The preferred format is a field-deployable rapid test. For mapping, the test needs to be ≥ 60% sensitive and ≥ 99.8% specific, while to support stopping decisions, the test must be ≥ 89% sensitive and ≥ 99.8% specific. The requirement for extremely high specificity is dictated by the need to detect with sufficient statistical confidence the low seroprevalence threshold set by WHO. Surveys designed to detect a 1-2% prevalence of a given biomarker, as is the case here, cannot tolerate more than 0.2% of false-positives. Otherwise, the background noise would drown out the signal. It is recognized that reaching and demonstrating such a stringent specificity criterion will be challenging, but test developers can expect to be assisted by national governments and implementing partners for adequately powered field validation.
Marco A. Biamonte ,Paul T. Cantey,Yaya I. Coulibaly,Katherine M. Gass,Louise C. Hamill,Christopher Hanna,Patrick J. Lammie,Joseph Kamgno,Thomas B. Nutman,David W. Oguttu,Dieudonné P. Sankara,Wilma A. Stolk,Thomas R. Unnasch. PLoS Negl Trop Dis. 2022 Aug 3;16(8):e0010682. doi: 10.1371/journal.pntd.0010682.
E3-SCF (Skp1/cullin-1/F-box protein) polyubiquitin ligases activate the proteasomal degradation of over a thousand proteins, but the evolutionary diversification of the F-box protein (FBP) family of substrate receptor subunits has challenged their elucidation in protists. Here we expand the FBP candidate list in the social amoeba Dictyostelium and show that the Skp1 interactome is highly remodeled as cells transition from solitary growth to multicellular development. Importantly, a subset of candidate FBPs was less represented when the posttranslational hydroxylation and glycosylation of Skp1 was abrogated by deletion of the O2 -sensing Skp1 prolyl hydroxylase PhyA. A role for this Skp1 modification for SCF activity was indicated by partial rescue of development, which normally depends on high O2 and PhyA, of phyA -knockout cells by proteasomal inhibitors. Further examination of two FBPs, FbxwD and the Jumonji C protein JcdI, suggested that Skp1 was substituted by other factors in phyA-knockout cells. Although a double-knockout of jcdI and its paralog jcdH did not affect development, overexpression of JcdI increased its sensitivity to O2. JcdI, a non-heme dioxygenase shown to have physiological O2-dependence, is conserved across protists with its F-box and other domains, and related to the human oncogene JmjD6. Sensitization of JcdI-overexpression cells to O2 depended on its dioxygenase activity and other domains, but not its F-box, which may however be the mediator of its reduced levels in wild-type relative to Skp1 modification mutant cells. The findings suggest that activation of JcdI by O2 is tempered by homeostatic down-regulation via PhyA and association with Skp1.
Andrew W Boland, Elisabet Gas-Pascual, Braxton L Nottingham, Hanke van der Wel, M Osman Sheikh, Christopher M Schafer, Christopher M West. J Biol Chem. 2022 Aug 3;102305. doi: 10.1016/j.jbc.2022.102305.
