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Author: Donna Huber

Chagas Disease Drug Discovery: Multiparametric Lead Optimization against Trypanosoma cruzi in Acylaminobenzothiazole Series

Acylaminobenzothiazole hits were identified as potential inhibitors of Trypanosoma cruzi replication, a parasite responsible for Chagas disease. We selected compound 1 for lead optimization, aiming to improve in parallel its anti-T. cruzi activity (IC50 = 0.63 μM) and its human metabolic stability (human clearance = 9.57 mL/min/g). A total of 39 analogues of 1 were synthesized and tested in vitro. We established a multiparametric structure-activity relationship, allowing optimization of antiparasite activity, physicochemical parameters, and ADME properties. We identified compound 50 as an advanced lead with an improved anti-T. cruzi activity in vitro (IC50 = 0.079 μM) and an enhanced metabolic stability (human clearance = 0.41 mL/min/g) and opportunity for the oral route of administration. After tolerability assessment, 50 demonstrated a promising in vivo efficacy.

Charlotte Fleau, Angel Padilla, Juan Miguel-Siles, Maria T. Quesada-Campos, Isabel Saiz-Nicolas, Ignacio Cotillo, Juan Cantizani Perez, Rick L. Tarleton, Maria Marco, Gilles Courtemanche. J Med Chem. 2019. doi: 10.1021/acs.jmedchem.9b01429.

Field Relevant Variation in Ambient Temperature Modifies Density-Dependent Establishment of Plasmodium falciparum Gametocytes in Mosquitoes

The relationship between Plasmodium falciparum gametocyte density and infections in mosquitoes is central to understanding the rates of transmission with important implications for control. Here, we determined whether field relevant variation in environmental temperature could also modulate this relationship. Anopheles stephensi were challenged with three densities of P. falciparum gametocytes spanning a ~10-fold gradient, and housed under diurnal/daily temperature range (“DTR”) of 9°C (+5°C and -4°C) around means of 20, 24, and 28°C. Vector competence was quantified as the proportion of mosquitoes infected with oocysts in the midguts (oocyst rates) or infectious with sporozoites in the salivary glands (sporozoite rates) at peak periods of infection for each temperature to account for the differences in development rates. In addition, oocyst intensities were also recorded from infected midguts and the overall study replicated across three separate parasite cultures and mosquito cohorts. While vector competence was similar at 20 DTR 9°C and 24 DTR 9°C, oocyst and sporozoite rates were also comparable, with evidence, surprisingly, for higher vector competence in mosquitoes challenged with intermediate gametocyte densities. For the same gametocyte densities however, severe reductions in the sporozoite rates was accompanied by a significant decline in overall vector competence at 28 DTR 9°C, with gametocyte density per se showing a positive and linear effect at this temperature. Unlike vector competence, oocyst intensities decreased with increasing temperatures with a predominantly positive and linear association with gametocyte density, especially at 28 DTR 9°C. Oocyst intensities across individual infected midguts suggested temperature-specific differences in mosquito susceptibility/resistance: at 20 DTR 9°C and 24 DTR 9°C, dispersion (aggregation) increased in a density-dependent manner but not at 28 DTR 9°C where the distributions were consistently random. Limitations notwithstanding, our results suggest that variation in temperature could modify seasonal dynamics of infectious reservoirs with implications for the design and deployment of transmission-blocking vaccines/drugs.

Ashutosh K. Pathak, Justine C. Shiau, Matthew B. Thomas and Courtney C. Murdock, Front Microbiol. 2019 Nov 15;10:2651. doi: 10.3389/fmicb.2019.02651. eCollection 2019.

Cultivation-assisted genome of Candidatus Fukatsuia symbiotica; the enigmatic ‘X-type’ symbiont of aphids

Heritable symbionts are common in terrestrial arthropods and often provide beneficial services to hosts. Unlike obligate, nutritional symbionts that largely persist under strict host control within specialized host cells, heritable facultative symbionts exhibit large variation in within-host lifestyles and services rendered with many retaining the capacity to transition among roles. One enigmatic symbiont, Candidatus Fukatsuia symbiotica, frequently infects aphids with reported roles ranging from pathogen, defensive symbiont, mutualism exploiter and nutritional co-obligate symbiont. Here we used an in vitro culture-assisted protocol to sequence the genome of a facultative strain of Fukatsuia from pea aphids (Acyrthosiphon pisum). Phylogenetic and genomic comparisons indicate that Fukatsuia is an aerobic heterotroph, which together with Regiella insecticola and Hamiltonella defensa form a clade of heritable facultative symbionts within the Yersiniaceae (Enterobacteriales). These three heritable facultative symbionts largely share overlapping inventories of genes associated with housekeeping functions, metabolism, and nutrient acquisition, while varying in complements of mobile DNA. One unusual feature of Fukatsuia is its strong tendency to occur as a co-infection with H. defensa. However, the overall similarity of gene inventories among aphid heritable facultative symbionts suggest that metabolic complementarity is not the basis for co-infection, unless playing out on a H. defensa strain-specific basis. We also compared the pea aphid Fukatsuia with a strain from the aphid Cinara confinis (Lachninae) where it is reported to have transitioned to co-obligate status to support decaying Buchnera function. Overall the two genomes are very similar with no clear genomic signatures consistent with such a transition, which suggests co-obligate status in C. confinis was a recent event.

V Patel, G Chevignon, A Manzano-Marín, J W Brandt, M R Strand, J A Russell, K M Oliver. 2019 Cultivation-assisted genome of Candidatus Fukatsuia symbiotica; the enigmatic ‘X-type’ symbiont of aphids. Genome Biology and Evolution, evz252, https://doi.org/10.1093/gbe/evz252

Sharing the Knowledge: NIH Award Supports Expanded Genomics Data Resource

By: Alan Flurry

A team led by scientists at the University of Pennsylvania and University of Georgia provides thousands of researchers around the world with access to the Eukaryotic Pathogen Genomics Database (EuPathDB.org), a collection of resources for analyzing large-scale datasets associated with microbial pathogens. These include the parasites responsible for malaria, sleeping sickness, and toxoplasmosis; the fungi responsible for thrush, aspergillosis and Valley Fever; and many other important diseases. In parallel, a team led by investigators at the University of Notre Dame has been responsible for similar resources covering invertebrate vectors of disease (VectorBase.org), including the mosquitoes transmitting malaria, Zika, and yellow fever, the ticks responsible for Lyme disease and Rocky Mountain Spotted Fever, and others.

To ensure that this important work continues, the National Institute of Allergy and Infectious Diseases, a part of the National Institutes of Health, has awarded a new contract to integrate these resources, worth up to $7.2 million in 2019-2020. The five‐year award for this project, rebranded as VEuPathDB.org (The Eukaryotic Pathogen, Host & Vector Genomics Resource) could total as much as $38.4 million if all associated options are exercised.

The patterns revealed by such “Big Data” provide insight into important diseases, permit the development of diagnostic methods, and define drug and vaccine targets. But to be useful, these immense datasets must be sensibly organized and made conveniently accessible to the researchers worldwide. The integrated VEuPathDB database hosts data on thousands of genomes, representing hundreds of species, along with extensive information on isolate provenance, gene function and the like.

The award is based at Penn, and directed by David S Roos, E Otis Kendall Professor of Biology in the School of Arts & Sciences. Key subcontracts include the University of Georgia (Joint PI Jessica C. Kissinger, Distinguished Professor of Genetics and Bioinformatics in the Franklin College of Arts and Sciences and the Center for Tropical and Emerging Global Diseases), University of Notre Dame (Joint PI Mary Ann McDowell, Associate Professor of Biological Sciences at the Eck Institute for Global Health).  Additional co-investigators include Professors Christian Stoeckert of Penn’s Perelman School of Medicine, Mark Caddick of the University of Liverpool, George K Christophides of Imperial College London, and Paul Flicek, Associate Director of the EMBL-EBI (European Bioinformatics Institute).

“It is wonderful to see the continued investment by NIH, the Wellcome Trust and others in resources that make performing much needed global research on infectious diseases both easier and better,” Kissinger said. “Datasets are larger and more complex than ever due to significant advances in technology. These breakthroughs create challenges for making the resulting data truly accessible and usable by the average researcher.  We strive to remove barriers, integrate diverse data and accelerate the speed with which new hypotheses can be generated and ideas tested both in silico and in the lab.”

“A critical aspect of this now joint program will be its accessibility throughout the world, empowering any infectious disease investigator to interrogate these highly complex databases in comprehensible and productive ways,” said Dan Colley, UGA professor of microbiology and member of the CTEGD who has conducted extensive research on n schistosomiasis in western Kenya. “These databases have led, and the merged data base will lead, to the design of new drugs and studies on how to better control and eliminate these major public health challenges, such as malaria, toxoplasmosis, yellow fever, eastern equine encephalitis and Lyme disease.”

“Since its conception, corresponding with the release of the first parasite genomes, EuPathDB has been a transformative tool in our search for a better understanding of human disease and parasite biology,” said Stephen Hadjuk, Professor Emeritus of biochemistry & molecular biology at UGA whose lab investigates trypanosomes, the causative agent of an human African sleeping sickness. “Today, it’s difficult to imagine any serious research on parasites and host pathology that doesn’t rely, at least to some extent, on EuPathDB. The decision to incorporate the vectors database into the eukaryotic pathogens database was brilliant, and makes this is an exciting new chapter in the EuPathDB story.”

“Innumerable investigators, including my own laboratory, rely on daily access to the high quality genomic and functional datasets made available by the VEuPathDB Project,” says Keith Gull, Professor of Molecular Microbiology at Oxford University.  “Sustainable support for such resources is imperative if we are to capitalize on the promise of modern technologies for scientific discovery and translational application.”  Joe Heitman, James B Duke Professor / Chair of Molecular Genetics & Microbiology at Duke University agrees: “Inclusion of fungal pathogens under the BRC umbrella has greatly enhanced our ability to study important human mycoses.  Cross-species comparisons provide insights into the biology and pathogenesis of these fascinating organisms, which can be deadly – but can also serve as workhorses for valuable biotechnology development.”

Originally published at https://www.franklin.uga.edu/news/stories/2019/sharing-knowledge-nih-award-supports-expanded-genomics-data-resource

Modeling approaches to predicting persistent hotspots in SCORE studies for gaining control of schistosomiasis mansoni in Kenya and Tanzania

BACKGROUND:

Some villages, labeled “persistent hotspots (PHS),” fail to respond adequately in regard to prevalence and intensity of infection to mass drug administration (MDA) for schistosomiasis. Early identification of PHS, for example, before initiating or after a year or two of MDA could help guide programmatic decision-making.

METHODS:

In a study with multiple rounds of MDA, data collected prior to the third MDA were used to predict PHS. We assessed six predictive approaches using data from before MDA and after 2 rounds of annual MDA from Kenya and Tanzania.

RESULTS:

Generalized linear models with variable selection possessed relatively stable performance compared to tree-based methods. Models applied to Kenya data alone or combined data from Kenya and Tanzania could reach over 80% predictive accuracy, while predicting PHS for Tanzania was challenging. Models developed from one country and validated in another failed to achieve satisfactory performance. Several Year 3 variables were identified as key predictors.

CONCLUSIONS:

Statistical models applied to Year 3 data could help predict PHS and guide program decisions, with infection intensity, prevalence of heavy infections (≥400 eggs/gram of feces), and total prevalence being particularly important factors. Additional studies including more variables and locations could help in developing generalizable models.

Ye Shen, Meng-Hsuan Sung, Charles H King, Sue Binder, Nupur Kittur, Christopher C Whalen, Daniel G Colley. J Infect Dis. 2019 Oct 17. pii: jiz529. doi: 10.1093/infdis/jiz529

Comparison of transcriptomes of an orthotospovirus vector and non-vector thrips species

Thrips transmit one of the most devastating plant viruses worldwide–tomato spotted wilt tospovirus (TSWV). Tomato spotted wilt tospovirus is a type species in the genus Orthotospovirus and family Tospoviridae. Although there are more than 7,000 thrips species, only nine thrips species are known to transmit TSWV. In this study, we investigated the molecular factors that could affect thrips ability to transmit TSWV. We assembled transcriptomes of a vector, Frankliniella fusca [Hinds], and a non-vector, Frankliniella tritici [Fitch], and performed qualitative comparisons of contigs associated with virus reception, virus infection, and innate immunity. Annotations of Ffusca and Ftritici contigs revealed slight differences across biological process and molecular functional groups. Comparison of virus cell surface receptors revealed that homologs of integrin were present in both species. However, homologs of another receptor, heperan sulfate, were present in Ffusca alone. Contigs associated with virus replication were identified in both species, but a contig involved in inhibition of virus replication (radical s-adenosylmethionine) was only present in the non-vector, Ftritici. Additionally, some differences in immune signaling pathways were identified between vector and non-vector thrips. Detailed investigations are necessary to functionally characterize these differences between vector and non-vector thrips and assess their relevance in orthotospovirus transmission.

Shrestha A, Champagne DE, Culbreath AK, Abney MR, Srinivasan R (2019) Comparison of transcriptomes of an orthotospovirus vector and non-vector thrips species. PLoS ONE 14(10): e0223438. https://doi.org/10.1371/journal.pone.0223438

Anibamine and Its Analogues: Potent Antiplasmodial Agents from Aniba citrifolia

In our continuing search for novel natural products with antiplasmodial activity, an extract of Aniba citrifolia was found to have good activity, with an IC50 value less than 1.25 μg/mL. After bioassay-directed fractionation, the known indolizinium alkaloid anibamine (1) and the new indolizinium alkaloid anibamine B (2) were isolated as the major bioactive constituents, with antiplasmodial IC50 values of 0.170 and 0.244 μM against the drug-resistant Dd2 strain of Plasmodium falciparum. The new coumarin anibomarin A (3), the new norneolignan anibignan A (5), and six known neolignans (712) were also obtained. The structures of all the isolated compounds were determined based on analyses of 1D and 2D NMR spectroscopic and mass spectrometric data, and the absolute configuration of anibignan A (5) was assigned from its ECD spectrum. Evaluation of a library of 28 anibamine analogues (1340) indicated that quaternary charged analogues had IC50 values as low as 58 nM, while uncharged analogues were inactive or significantly less active. Assessment of the potential effects of anibamine and its analogues on the intraerythrocytic stages and morphological development of P. falciparum revealed substantial activity against ring stages for compounds with two C-10 side chains, while those with only one C-10 side chain exhibited substantial activity against trophozoite stages, suggesting different mechanisms of action.

Yongle Du, Ana Lisa Valenciano, Yumin Dai, Yi Zheng, Feng Zhang, Yan Zhang, Jason Clement, Michael Goetz, David G. I. Kingston, Maria B. Cassera. 2019. J Nat Prod. doi: 10.1021/acs.jnatprod.9b00724.

Sylvatic cycles of arboviruses in non-human primates

Arboviruses infecting people primarily exist in urban transmission cycles involving urban mosquitoes in densely populated tropical regions. For dengue, chikungunya, Zika and yellow fever viruses, sylvatic (forest) transmission cycles also exist in some regions and involve non-human primates and forest-dwelling mosquitoes. Here we review the investigation methods and available data on sylvatic cycles involving non-human primates and dengue, chikungunya, Zika and yellow fever viruses in Africa, dengue viruses in Asia and yellow fever virus in the Americas. We also present current putative data that Mayaro, o’nyong’nyong, Oropouche, Spondweni and Lumbo viruses exist in sylvatic cycles.

Matthew John Valentine, Courtney Cuin Murdock & Patrick John Kelly. 2019. Parasit Vectors.;12(1):463. doi: 10.1186/s13071-019-3732-0

Mapping Schistosoma mansoni endemicity in Rwanda: a critical assessment of geographical disparities arising from circulating cathodic antigen versus Kato-Katz diagnostics

Background

Schistosomiasis is a neglected tropical disease caused by Schistosoma parasites. Intervention relies on identifying high-risk regions, yet rapid Schistosoma diagnostics (Kato-Katz stool assays (KK) and circulating cathodic antigen urine assays (CCA)) yield different prevalence estimates. We mapped Smansoni prevalence and delineated at-risk regions using a survey of schoolchildren in Rwanda, where Schistosoma mansoni is an endemic parasite. We asked if different diagnostics resulted in disparities in projected infection risk.

 

Methods

Infection data was obtained from a 2014 Rwandan school-based survey that used KK and CCA diagnostics. Across 386 schools screened by CCA (N = 19,217). To allow for uncertainty when interpreting ambiguous CCA trace readings, which accounted for 28.8% of total test results, we generated two presence-absence datasets: CCA trace as positive and CCA trace as negative. Samples (N = 9,175) from 185 schools were also screened by KK. We included land surface temperature (LST) and the Normalized Difference Vegetation and Normalized Difference Water Indices (NDVI, NDWI) as predictors in geostatistical regressions.

 

Findings

Across 8,647 children tested by both methods, prevalence was 35.93% for CCA trace as positive, 7.21% for CCA trace as negative and 1.95% for KK. LST was identified as a risk factor using KK, whereas NDVI was a risk factor for CCA models. Models predicted high endemicity in Northern and Western regions of Rwanda, though the CCA trace as positive model identified additional high-risk areas that were overlooked by the other methods. Estimates of current burden for children at highest risk (boys aged 5–9 years) varied by an order of magnitude, with 671,856 boys projected to be infected by CCA trace as positive and only 60,453 projected by CCA trace as negative results.

 

Conclusions

Our findings show that people in Rwanda’s Northern, Western and capital regions are at high risk of Smansoni infection. However, variation in identification of environmental risk factors and delineation of at-risk regions using different diagnostics likely provides confusing messages to disease intervention managers. Further research and statistical analyses, such as latent class analysis, can be used to improve CCA result classification and assess its use in guiding treatment regimes.

 

Clark NJ, Umulisa I, Ruberanziza E, Owada K, Colley DG, Ortu G, et al. (2019) Mapping Schistosoma mansoni endemicity in Rwanda: a critical assessment of geographical disparities arising from circulating cathodic antigen versus Kato-Katz diagnostics. PLoS Negl Trop Dis 13(9): e0007723. https://doi.org/10.1371/journal.pntd.0007723