Publications by authors named "Vida Ahyong"

19 Publications

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The COVID-19 epidemic in Madagascar: clinical description and laboratory results of the first wave, march-september 2020.

Influenza Other Respir Viruses 2021 Feb 15. Epub 2021 Feb 15.

Virology Unit, Institut Pasteur de Madagascar, Antananarivo, Madagascar.

Background: Following the first detection of SARS-CoV-2 in passengers arriving from Europe on 19 March 2020, Madagascar took several mitigation measures to limit the spread of the virus in the country.

Methods: Nasopharyngeal and/or oropharyngeal swabs were collected from travellers to Madagascar, suspected SARS-CoV-2 cases and contact of confirmed cases. Swabs were tested at the national reference laboratory using real-time RT-PCR. Data collected from patients were entered in an electronic database for subsequent statistical analysis. All distribution of laboratory-confirmed cases were mapped, and six genomes of viruses were fully sequenced.

Results: Overall, 26,415 individuals were tested for SARS-CoV-2 between 18 March and 18 September 2020, of whom 21.0% (5,553/26,145) returned positive. Among laboratory-confirmed SARS-CoV-2-positive patients, the median age was 39 years (IQR: 28-52), and 56.6% (3,311/5,553) were asymptomatic at the time of sampling. The probability of testing positive increased with age with the highest adjusted odds ratio of 2.2 [95% CI: 1.9-2.5] for individuals aged 49 years and more. Viral strains sequenced belong to clades 19A, 20A and 20B indicative of several independent introduction of viruses.

Conclusions: Our study describes the first wave of the COVID-19 in Madagascar. Despite early strategies in place Madagascar could not avoid the introduction and spread of the virus. More studies are needed to estimate the true burden of disease and make public health recommendations for a better preparation to another wave.
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http://dx.doi.org/10.1111/irv.12845DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8013501PMC
February 2021

Rapid deployment of SARS-CoV-2 testing: The CLIAHUB.

Authors:
Emily D Crawford Irene Acosta Vida Ahyong Erika C Anderson Shaun Arevalo Daniel Asarnow Shannon Axelrod Patrick Ayscue Camillia S Azimi Caleigh M Azumaya Stefanie Bachl Iris Bachmutsky Aparna Bhaduri Jeremy Bancroft Brown Joshua Batson Astrid Behnert Ryan M Boileau Saumya R Bollam Alain R Bonny David Booth Michael Jerico B Borja David Brown Bryan Buie Cassandra E Burnett Lauren E Byrnes Katelyn A Cabral Joana P Cabrera Saharai Caldera Gabriela Canales Gloria R Castañeda Agnes Protacio Chan Christopher R Chang Arthur Charles-Orszag Carly Cheung Unseng Chio Eric D Chow Y Rose Citron Allison Cohen Lillian B Cohn Charles Chiu Mitchel A Cole Daniel N Conrad Angela Constantino Andrew Cote Tre'Jon Crayton-Hall Spyros Darmanis Angela M Detweiler Rebekah L Dial Shen Dong Elias M Duarte David Dynerman Rebecca Egger Alison Fanton Stacey M Frumm Becky Xu Hua Fu Valentina E Garcia Julie Garcia Christina Gladkova Miriam Goldman Rafael Gomez-Sjoberg M Grace Gordon James C R Grove Shweta Gupta Alexis Haddjeri-Hopkins Pierce Hadley John Haliburton Samantha L Hao George Hartoularos Nadia Herrera Melissa Hilberg Kit Ying E Ho Nicholas Hoppe Shayan Hosseinzadeh Conor J Howard Jeffrey A Hussmann Elizabeth Hwang Danielle Ingebrigtsen Julia R Jackson Ziad M Jowhar Danielle Kain James Y S Kim Amy Kistler Oriana Kreutzfeld Jessie Kulsuptrakul Andrew F Kung Charles Langelier Matthew T Laurie Lena Lee Kun Leng Kristoffer E Leon Manuel D Leonetti Sophia R Levan Sam Li Aileen W Li Jamin Liu Heidi S Lubin Amy Lyden Jennifer Mann Sabrina Mann Gorica Margulis Diana M Marquez Bryan P Marsh Calla Martyn Elizabeth E McCarthy Aaron McGeever Alexander F Merriman Lauren K Meyer Steve Miller Megan K Moore Cody T Mowery Tanzila Mukhtar Lusajo L Mwakibete Noelle Narez Norma F Neff Lindsay A Osso Diter Oviedo Suping Peng Maira Phelps Kiet Phong Peter Picard Lindsey M Pieper Neha Pincha Angela Oliveira Pisco Angela Pogson Sergei Pourmal Robert R Puccinelli Andreas S Puschnik Elze Rackaityte Preethi Raghavan Madhura Raghavan James Reese Joseph M Replogle Hanna Retallack Helen Reyes Donald Rose Marci F Rosenberg Estella Sanchez-Guerrero Sydney M Sattler Laura Savy Stephanie K See Kristin K Sellers Paula Hayakawa Serpa Maureen Sheehy Jonathan Sheu Sukrit Silas Jessica A Streithorst Jack Strickland Doug Stryke Sara Sunshine Peter Suslow Renaldo Sutanto Serena Tamura Michelle Tan Jiongyi Tan Alice Tang Cristina M Tato Jack C Taylor Iliana Tenvooren Erin M Thompson Edward C Thornborrow Eric Tse Tony Tung Marc L Turner Victoria S Turner Rigney E Turnham Mary J Turocy Trisha V Vaidyanathan Ilia D Vainchtein Manu Vanaerschot Sara E Vazquez Anica M Wandler Anne Wapniarski James T Webber Zara Y Weinberg Alexandra Westbrook Allison W Wong Emily Wong Gajus Worthington Fang Xie Albert Xu Terrina Yamamoto Ying Yang Fauna Yarza Yefim Zaltsman Tina Zheng Joseph L DeRisi

PLoS Pathog 2020 10 28;16(10):e1008966. Epub 2020 Oct 28.

Chan Zuckerberg Biohub, San Francisco, California, United States of America.

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http://dx.doi.org/10.1371/journal.ppat.1008966DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7592773PMC
October 2020

IDseq-An open source cloud-based pipeline and analysis service for metagenomic pathogen detection and monitoring.

Gigascience 2020 10;9(10)

Chan Zuckerberg Biohub, 499 Illinois St, San Francisco, CA 94158, USA.

Background: Metagenomic next-generation sequencing (mNGS) has enabled the rapid, unbiased detection and identification of microbes without pathogen-specific reagents, culturing, or a priori knowledge of the microbial landscape. mNGS data analysis requires a series of computationally intensive processing steps to accurately determine the microbial composition of a sample. Existing mNGS data analysis tools typically require bioinformatics expertise and access to local server-class hardware resources. For many research laboratories, this presents an obstacle, especially in resource-limited environments.

Findings: We present IDseq, an open source cloud-based metagenomics pipeline and service for global pathogen detection and monitoring (https://idseq.net). The IDseq Portal accepts raw mNGS data, performs host and quality filtration steps, then executes an assembly-based alignment pipeline, which results in the assignment of reads and contigs to taxonomic categories. The taxonomic relative abundances are reported and visualized in an easy-to-use web application to facilitate data interpretation and hypothesis generation. Furthermore, IDseq supports environmental background model generation and automatic internal spike-in control recognition, providing statistics that are critical for data interpretation. IDseq was designed with the specific intent of detecting novel pathogens. Here, we benchmark novel virus detection capability using both synthetically evolved viral sequences and real-world samples, including IDseq analysis of a nasopharyngeal swab sample acquired and processed locally in Cambodia from a tourist from Wuhan, China, infected with the recently emergent SARS-CoV-2.

Conclusion: The IDseq Portal reduces the barrier to entry for mNGS data analysis and enables bench scientists, clinicians, and bioinformaticians to gain insight from mNGS datasets for both known and novel pathogens.
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http://dx.doi.org/10.1093/gigascience/giaa111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7566497PMC
October 2020

Complete Genome Sequence of a Novel Coronavirus (SARS-CoV-2) Isolate from Bangladesh.

Microbiol Resour Announc 2020 Jun 11;9(24). Epub 2020 Jun 11.

Child Health Research Foundation, Dhaka, Bangladesh.

The complete genome sequence of a novel coronavirus (severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2]) isolate obtained from a nasopharyngeal swab from a patient with COVID-19 in Bangladesh is reported.
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http://dx.doi.org/10.1128/MRA.00568-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7291105PMC
June 2020

Rapid metagenomic characterization of a case of imported COVID-19 in Cambodia.

bioRxiv 2020 Mar 5. Epub 2020 Mar 5.

Virology Unit, Institut Pasteur du Cambodge, Phnom Penh, Cambodia.

Rapid production and publication of pathogen genome sequences during emerging disease outbreaks provide crucial public health information. In resource-limited settings, especially near an outbreak epicenter, conventional deep sequencing or bioinformatics are often challenging. Here we successfully used metagenomic next generation sequencing on an iSeq100 Illumina platform paired with an open-source bioinformatics pipeline to quickly characterize Cambodia's first case of COVID-2019.
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http://dx.doi.org/10.1101/2020.03.02.968818DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7217139PMC
March 2020

Co-occurrence of Species in Children From Eastern Ethiopia, and Their Association With Environmental Enteric Dysfunction, Diarrhea, and Host Microbiome.

Front Public Health 2020 15;8:99. Epub 2020 Apr 15.

The Ohio State University, Columbus, OH, United States.

High prevalence during early childhood has been associated with stunting and environmental enteric dysfunction (EED), especially in low resource settings. This study assessed the prevalence, diversity, abundance, and co-occurrence of spp. in stools from children in a rural area of eastern Ethiopia and their association with microbiome, diarrhea, and EED in children. Stool samples ( = 100) were collected from randomly selected children (age range: 360-498 days) in five kebeles in Haramaya District, Ethiopia. Diarrhea, compromised gut permeability, and gut inflammation were observed in 48, 45, and 57% of children, respectively. prevalence and species diversity were assessed using PCR and meta-total RNA sequencing (MeTRS). The prevalence of spp. in the children's stools was 50% (41-60%) by PCR and 88% (80-93.6%) by MeTRS ( < 0.01). Further, seven species ( sp. RM6137, uncultured sp., and sp. RM12175) were detected by MeTRS in at least 40% of children stools in high abundance (>1.76-log read per million per positive stool sample). Four clusters of species (5-12 species per cluster) co-occurred in the stool samples, suggesting that colonization of children may have occurred through multiple reservoirs or from a reservoir in which several species may co-inhabit. No associations between spp., EED, and diarrhea were detected in this cross-sectional study; however, characteristic microbiome profiles were identified based on the prevalence of spp., EED severity, and diarrhea. Forty-seven bacterial species were correlated with , and 13 of them also correlated with gut permeability, gut inflammation and/or EED severity. Forty-nine species not correlated with were correlated with gut permeability, gut inflammation, EED severity and/or diarrhea. This study demonstrated that (1) in addition to and , multiple non-thermophilic spp. (i.e., , and ) were frequently detected in the children's stools and (2) the , gut permeability, gut inflammation, EED severity, and diarrhea were associated with characteristic microbiome composition. Additional spatial and longitudinal studies are needed to identify environmental reservoirs and sources of infection of children with disparate species and to better define their associations with EED in low-income countries.
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http://dx.doi.org/10.3389/fpubh.2020.00099DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7174729PMC
April 2020

A Metabolic Dependency for Host Isoprenoids in the Obligate Intracellular Pathogen Rickettsia parkeri Underlies a Sensitivity to the Statin Class of Host-Targeted Therapeutics.

mSphere 2019 11 13;4(6). Epub 2019 Nov 13.

Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA

Gram-negative bacteria in the order have an obligate intracellular growth requirement, and some species cause human diseases such as typhus and spotted fever. The bacteria have evolved a dependence on essential nutrients and metabolites from the host cell as a consequence of extensive genome reduction. However, it remains largely unknown which nutrients they acquire and whether their metabolic dependency can be exploited therapeutically. Here, we describe a genetic rewiring of bacterial isoprenoid biosynthetic pathways in the that has resulted from reductive genome evolution. Furthermore, we investigated whether the spotted fever group species scavenges isoprenoid precursors directly from the host. Using targeted mass spectrometry, we found that infection caused decreases in host isoprenoid products and concomitant increases in bacterial isoprenoid metabolites. Additionally, we report that treatment of infected cells with statins, which inhibit host isoprenoid synthesis, prohibited bacterial growth. We show that growth inhibition correlates with changes in bacterial size and shape that mimic those caused by antibiotics that inhibit peptidoglycan biosynthesis, suggesting that statins lead to an inhibition of cell wall synthesis. Altogether, our results describe a potential Achilles' heel of obligate intracellular pathogens that can potentially be exploited with host-targeted therapeutics that interfere with metabolic pathways required for bacterial growth. Obligate intracellular pathogens, which include viruses as well as certain bacteria and eukaryotes, are a subset of infectious microbes that are metabolically dependent on and unable to grow outside an infected host cell because they have lost or lack essential biosynthetic pathways. In this study, we describe a metabolic dependency of the bacterial pathogen on host isoprenoid molecules that are used in the biosynthesis of downstream products, including cholesterol, steroid hormones, and heme. Bacteria make products from isoprenoids, such as an essential lipid carrier for making the bacterial cell wall. We show that bacterial metabolic dependency can represent a potential Achilles' heel and that inhibiting host isoprenoid biosynthesis with the FDA-approved statin class of drugs inhibits bacterial growth by interfering with the integrity of the cell wall. This work supports the potential to treat infections by obligate intracellular pathogens through inhibition of host biosynthetic pathways that are susceptible to parasitism.
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http://dx.doi.org/10.1128/mSphere.00536-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6854040PMC
November 2019

RNAi screen reveals a role for PACSIN2 and caveolins during bacterial cell-to-cell spread.

Mol Biol Cell 2019 08 26;30(17):2124-2133. Epub 2019 Jun 26.

Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139.

is a human bacterial pathogen that disseminates through host tissues using a process called cell-to-cell spread. This critical yet understudied virulence strategy resembles a vesicular form of intercellular trafficking that allows to move between host cells without escaping the cell. Interestingly, eukaryotic cells can also directly exchange cellular components via intercellular communication pathways (e.g., -endocytosis) using cell-cell adhesion, membrane trafficking, and membrane remodeling proteins. Therefore, we hypothesized that would hijack these types of host proteins during spread. Using a focused RNA interference screen, we identified 22 host genes that are important for spread. We then found that caveolins (CAV1 and CAV2) and the membrane sculpting F-BAR protein PACSIN2 promote protrusion engulfment during spread, and that PACSIN2 specifically localizes to protrusions. Overall, our study demonstrates that host intercellular communication pathways may be coopted during bacterial spread and that specific trafficking and membrane remodeling proteins promote bacterial protrusion resolution.
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http://dx.doi.org/10.1091/mbc.E19-04-0197DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6743452PMC
August 2019

The Plasmodium falciparum cytoplasmic translation apparatus: a promising therapeutic target not yet exploited by clinically approved anti-malarials.

Malar J 2018 Dec 12;17(1):465. Epub 2018 Dec 12.

Department of Biochemistry and Biophysics, University of California San Francisco, San Francisco, CA, USA.

Background: The continued spectre of resistance to existing anti-malarials necessitates the pursuit of novel targets and mechanisms of action for drug development. One class of promising targets consists of the 80S ribosome and its associated components comprising the parasite translational apparatus. Development of translation-targeting therapeutics requires a greater understanding of protein synthesis and its regulation in the malaria parasite. Research in this area has been limited by the lack of appropriate experimental methods, particularly a direct measure of parasite translation.

Methods: An in vitro method directly measuring translation in whole-cell extracts from the malaria parasite Plasmodium falciparum, the PfIVT assay, and a historically-utilized indirect measure of translation, S35-radiolabel incorporation, were compared utilizing a large panel of known translation inhibitors as well as anti-malarial drugs.

Results: Here, an extensive pharmacologic assessment of the PfIVT assay is presented, using a wide range of known inhibitors demonstrating its utility for studying activity of both ribosomal and non-ribosomal elements directly involved in translation. Further, the superiority of this assay over a historically utilized indirect measure of translation, S35-radiolabel incorporation, is demonstrated. Additionally, the PfIVT assay is utilized to investigate a panel of clinically approved anti-malarial drugs, many with unknown or unclear mechanisms of action, and show that none inhibit translation, reaffirming Plasmodium translation to be a viable alternative drug target. Within this set, mefloquine is unambiguously found to lack translation inhibition activity, despite having been recently mischaracterized as a ribosomal inhibitor.

Conclusions: This work exploits a direct and reproducible assay for measuring P. falciparum translation, demonstrating its value in the continued study of protein synthesis in malaria and its inhibition as a drug target.
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http://dx.doi.org/10.1186/s12936-018-2616-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6292128PMC
December 2018

Natural Variation in and Underlie Condition-Specific Growth Defects in .

G3 (Bethesda) 2018 01 4;8(1):239-251. Epub 2018 Jan 4.

Pacific Northwest Research Institute, Seattle, Washington 98122

Despite their ubiquitous use in laboratory strains, naturally occurring loss-of-function mutations in genes encoding core metabolic enzymes are relatively rare in wild isolates of Here, we identify a naturally occurring serine auxotrophy in a sake brewing strain from Japan. Through a cross with a honey wine (white tecc) brewing strain from Ethiopia, we map the minimal medium growth defect to , which encodes 3-phosphoserine aminotransferase and is orthologous to the human disease gene, To investigate the impact of this polymorphism under conditions of abundant external nutrients, we examine growth in rich medium alone or with additional stresses, including the drugs caffeine and rapamycin and relatively high concentrations of copper, salt, and ethanol. Consistent with studies that found widespread effects of different auxotrophies on RNA expression patterns in rich media, we find that the loss-of-function allele dominates the quantitative trait locus (QTL) landscape under many of these conditions, with a notable exacerbation of the effect in the presence of rapamycin and caffeine. We also identify a major-effect QTL associated with growth on salt that maps to the gene encoding the sodium exporter, We demonstrate that the salt phenotype is largely driven by variation in the promoter, which harbors a deletion that removes binding sites for the Mig1 and Nrg1 transcriptional repressors. Thus, our results identify natural variation associated with both coding and regulatory regions of the genome that underlie strong growth phenotypes.
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http://dx.doi.org/10.1534/g3.117.300392DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5765352PMC
January 2018

A potent antimalarial benzoxaborole targets a Plasmodium falciparum cleavage and polyadenylation specificity factor homologue.

Nat Commun 2017 03 6;8:14574. Epub 2017 Mar 6.

Department of Medicine, University of California, Box 0811, San Francisco, California 94143, USA.

Benzoxaboroles are effective against bacterial, fungal and protozoan pathogens. We report potent activity of the benzoxaborole AN3661 against Plasmodium falciparum laboratory-adapted strains (mean IC 32 nM), Ugandan field isolates (mean ex vivo IC 64 nM), and murine P. berghei and P. falciparum infections (day 4 ED 0.34 and 0.57 mg kg, respectively). Multiple P. falciparum lines selected in vitro for resistance to AN3661 harboured point mutations in pfcpsf3, which encodes a homologue of mammalian cleavage and polyadenylation specificity factor subunit 3 (CPSF-73 or CPSF3). CRISPR-Cas9-mediated introduction of pfcpsf3 mutations into parental lines recapitulated AN3661 resistance. PfCPSF3 homology models placed these mutations in the active site, where AN3661 is predicted to bind. Transcripts for three trophozoite-expressed genes were lost in AN3661-treated trophozoites, which was not observed in parasites selected or engineered for AN3661 resistance. Our results identify the pre-mRNA processing factor PfCPSF3 as a promising antimalarial drug target.
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http://dx.doi.org/10.1038/ncomms14574DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5343452PMC
March 2017

Open Source Drug Discovery with the Malaria Box Compound Collection for Neglected Diseases and Beyond.

Authors:
Wesley C Van Voorhis John H Adams Roberto Adelfio Vida Ahyong Myles H Akabas Pietro Alano Aintzane Alday Yesmalie Alemán Resto Aishah Alsibaee Ainhoa Alzualde Katherine T Andrews Simon V Avery Vicky M Avery Lawrence Ayong Mark Baker Stephen Baker Choukri Ben Mamoun Sangeeta Bhatia Quentin Bickle Lotfi Bounaadja Tana Bowling Jürgen Bosch Lauren E Boucher Fabrice F Boyom Jose Brea Marian Brennan Audrey Burton Conor R Caffrey Grazia Camarda Manuela Carrasquilla Dee Carter Maria Belen Cassera Ken Chih-Chien Cheng Worathad Chindaudomsate Anthony Chubb Beatrice L Colon Daisy D Colón-López Yolanda Corbett Gregory J Crowther Noemi Cowan Sarah D'Alessandro Na Le Dang Michael Delves Joseph L DeRisi Alan Y Du Sandra Duffy Shimaa Abd El-Salam El-Sayed Michael T Ferdig José A Fernández Robledo David A Fidock Isabelle Florent Patrick V T Fokou Ani Galstian Francisco Javier Gamo Suzanne Gokool Ben Gold Todd Golub Gregory M Goldgof Rajarshi Guha W Armand Guiguemde Nil Gural R Kiplin Guy Michael A E Hansen Kirsten K Hanson Andrew Hemphill Rob Hooft van Huijsduijnen Takaaki Horii Paul Horrocks Tyler B Hughes Christopher Huston Ikuo Igarashi Katrin Ingram-Sieber Maurice A Itoe Ajit Jadhav Amornrat Naranuntarat Jensen Laran T Jensen Rays H Y Jiang Annette Kaiser Jennifer Keiser Thomas Ketas Sebastien Kicka Sunyoung Kim Kiaran Kirk Vidya P Kumar Dennis E Kyle Maria Jose Lafuente Scott Landfear Nathan Lee Sukjun Lee Adele M Lehane Fengwu Li David Little Liqiong Liu Manuel Llinás Maria I Loza Aristea Lubar Leonardo Lucantoni Isabelle Lucet Louis Maes Dalu Mancama Nuha R Mansour Sandra March Sheena McGowan Iset Medina Vera Stephan Meister Luke Mercer Jordi Mestres Alvine N Mfopa Raj N Misra Seunghyun Moon John P Moore Francielly Morais Rodrigues da Costa Joachim Müller Arantza Muriana Stephen Nakazawa Hewitt Bakela Nare Carl Nathan Nathalie Narraidoo Sujeevi Nawaratna Kayode K Ojo Diana Ortiz Gordana Panic George Papadatos Silvia Parapini Kailash Patra Ngoc Pham Sarah Prats David M Plouffe Sally-Ann Poulsen Anupam Pradhan Celia Quevedo Ronald J Quinn Christopher A Rice Mohamed Abdo Rizk Andrea Ruecker Robert St Onge Rafaela Salgado Ferreira Jasmeet Samra Natalie G Robinett Ulrich Schlecht Marjorie Schmitt Filipe Silva Villela Francesco Silvestrini Robert Sinden Dennis A Smith Thierry Soldati Andreas Spitzmüller Serge Maximilian Stamm David J Sullivan William Sullivan Sundari Suresh Brian M Suzuki Yo Suzuki S Joshua Swamidass Donatella Taramelli Lauve R Y Tchokouaha Anjo Theron David Thomas Kathryn F Tonissen Simon Townson Abhai K Tripathi Valentin Trofimov Kenneth O Udenze Imran Ullah Cindy Vallieres Edgar Vigil Joseph M Vinetz Phat Voong Vinh Hoan Vu Nao-Aki Watanabe Kate Weatherby Pamela M White Andrew F Wilks Elizabeth A Winzeler Edward Wojcik Melanie Wree Wesley Wu Naoaki Yokoyama Paul H A Zollo Nada Abla Benjamin Blasco Jeremy Burrows Benoît Laleu Didier Leroy Thomas Spangenberg Timothy Wells Paul A Willis

PLoS Pathog 2016 07 28;12(7):e1005763. Epub 2016 Jul 28.

Medicines for Malaria Venture, Geneva, Switzerland.

A major cause of the paucity of new starting points for drug discovery is the lack of interaction between academia and industry. Much of the global resource in biology is present in universities, whereas the focus of medicinal chemistry is still largely within industry. Open source drug discovery, with sharing of information, is clearly a first step towards overcoming this gap. But the interface could especially be bridged through a scale-up of open sharing of physical compounds, which would accelerate the finding of new starting points for drug discovery. The Medicines for Malaria Venture Malaria Box is a collection of over 400 compounds representing families of structures identified in phenotypic screens of pharmaceutical and academic libraries against the Plasmodium falciparum malaria parasite. The set has now been distributed to almost 200 research groups globally in the last two years, with the only stipulation that information from the screens is deposited in the public domain. This paper reports for the first time on 236 screens that have been carried out against the Malaria Box and compares these results with 55 assays that were previously published, in a format that allows a meta-analysis of the combined dataset. The combined biochemical and cellular assays presented here suggest mechanisms of action for 135 (34%) of the compounds active in killing multiple life-cycle stages of the malaria parasite, including asexual blood, liver, gametocyte, gametes and insect ookinete stages. In addition, many compounds demonstrated activity against other pathogens, showing hits in assays with 16 protozoa, 7 helminths, 9 bacterial and mycobacterial species, the dengue fever mosquito vector, and the NCI60 human cancer cell line panel of 60 human tumor cell lines. Toxicological, pharmacokinetic and metabolic properties were collected on all the compounds, assisting in the selection of the most promising candidates for murine proof-of-concept experiments and medicinal chemistry programs. The data for all of these assays are presented and analyzed to show how outstanding leads for many indications can be selected. These results reveal the immense potential for translating the dispersed expertise in biological assays involving human pathogens into drug discovery starting points, by providing open access to new families of molecules, and emphasize how a small additional investment made to help acquire and distribute compounds, and sharing the data, can catalyze drug discovery for dozens of different indications. Another lesson is that when multiple screens from different groups are run on the same library, results can be integrated quickly to select the most valuable starting points for subsequent medicinal chemistry efforts.
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http://dx.doi.org/10.1371/journal.ppat.1005763DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4965013PMC
July 2016

Antimalarial Benzoxaboroles Target Plasmodium falciparum Leucyl-tRNA Synthetase.

Antimicrob Agents Chemother 2016 08 22;60(8):4886-95. Epub 2016 Jul 22.

Department of Medicine, University of California, San Francisco, California, USA

There is a need for new antimalarials, ideally with novel mechanisms of action. Benzoxaboroles have been shown to be active against bacteria, fungi, and trypanosomes. Therefore, we investigated the antimalarial activity and mechanism of action of 3-aminomethyl benzoxaboroles against Plasmodium falciparum Two 3-aminomethyl compounds, AN6426 and AN8432, demonstrated good potency against cultured multidrug-resistant (W2 strain) P. falciparum (50% inhibitory concentration [IC50] of 310 nM and 490 nM, respectively) and efficacy against murine Plasmodium berghei infection when administered orally once daily for 4 days (90% effective dose [ED90], 7.4 and 16.2 mg/kg of body weight, respectively). To characterize mechanisms of action, we selected parasites with decreased drug sensitivity by culturing with stepwise increases in concentration of AN6426. Resistant clones were characterized by whole-genome sequencing. Three generations of resistant parasites had polymorphisms in the predicted editing domain of the gene encoding a P. falciparum leucyl-tRNA synthetase (LeuRS; PF3D7_0622800) and in another gene (PF3D7_1218100), which encodes a protein of unknown function. Solution of the structure of the P. falciparum LeuRS editing domain suggested key roles for mutated residues in LeuRS editing. Short incubations with AN6426 and AN8432, unlike artemisinin, caused dose-dependent inhibition of [(14)C]leucine incorporation by cultured wild-type, but not resistant, parasites. The growth of resistant, but not wild-type, parasites was impaired in the presence of the unnatural amino acid norvaline, consistent with a loss of LeuRS editing activity in resistant parasites. In summary, the benzoxaboroles AN6426 and AN8432 offer effective antimalarial activity and act, at least in part, against a novel target, the editing domain of P. falciparum LeuRS.
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http://dx.doi.org/10.1128/AAC.00820-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4958189PMC
August 2016

Identification of Plasmodium falciparum specific translation inhibitors from the MMV Malaria Box using a high throughput in vitro translation screen.

Malar J 2016 Mar 17;15:173. Epub 2016 Mar 17.

Department of Biochemistry and Biophysics, University of California San Francisco, San Francisco, CA, USA.

Background: A major goal in the search for new anti-malarial compounds is to identify new mechanisms of action or new molecular targets. While cell-based, growth inhibition-based screening have enjoyed tremendous success, an alternative approach is to specifically assay a given pathway or essential cellular process.

Methods: Here, this work describes the development of a plate-based, in vitro luciferase assay to probe for inhibitors specific to protein synthesis in Plasmodium falciparum through the use of an in vitro translation system derived from the parasite.

Results: Using the Medicines for Malaria Venture's Malaria Box as a pilot, 400 bioactive compounds with minimal human cytotoxicity profiles were screened, identifying eight compounds that displayed greater potency against the P. falciparum translation machinery relative to a mammalian translation system. Dose-response curves were determined in both translation systems to further characterize the top hit compound (MMV008270).

Conclusions: This assay will be useful not only in future anti-malarial screening efforts but also in the investigation of P. falciparum protein synthesis and essential processes in P. falciparum biology.
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http://dx.doi.org/10.1186/s12936-016-1231-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4794828PMC
March 2016

Genome-wide regulatory dynamics of translation in the asexual blood stages.

Elife 2014 Dec 10;3. Epub 2014 Dec 10.

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States.

The characterization of the transcriptome and proteome of has been a tremendous resource for the understanding of the molecular physiology of this parasite. However, the translational dynamics that link steady-state mRNA with protein levels are not well understood. In this study, we bridge this disconnect by measuring genome-wide translation using ribosome profiling, through five stages of the blood phase developmental cycle. Our findings show that transcription and translation are tightly coupled, with overt translational control occurring for less than 10% of the transcriptome. Translationally regulated genes are predominantly associated with merozoite egress functions. We systematically define mRNA 5' leader sequences, and 3' UTRs, as well as antisense transcripts, along with ribosome occupancy for each, and establish that accumulation of ribosomes on 5' leaders is a common transcript feature. This work represents the highest resolution and broadest portrait of gene expression and translation to date for this medically important parasite.
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http://dx.doi.org/10.7554/eLife.04106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4371882PMC
December 2014

Aneuploidy underlies a multicellular phenotypic switch.

Proc Natl Acad Sci U S A 2013 Jul 28;110(30):12367-72. Epub 2013 Jun 28.

Institute for Systems Biology, Seattle, WA 98109, USA.

Although microorganisms are traditionally used to investigate unicellular processes, the yeast Saccharomyces cerevisiae has the ability to form colonies with highly complex, multicellular structures. Colonies with the "fluffy" morphology have properties reminiscent of bacterial biofilms and are easily distinguished from the "smooth" colonies typically formed by laboratory strains. We have identified strains that are able to reversibly toggle between the fluffy and smooth colony-forming states. Using a combination of flow cytometry and high-throughput restriction-site associated DNA tag sequencing, we show that this switch is correlated with a change in chromosomal copy number. Furthermore, the gain of a single chromosome is sufficient to switch a strain from the fluffy to the smooth state, and its subsequent loss to revert the strain back to the fluffy state. Because copy number imbalance of six of the 16 S. cerevisiae chromosomes and even a single gene can modulate the switch, our results support the hypothesis that the state switch is produced by dosage-sensitive genes, rather than a general response to altered DNA content. These findings add a complex, multicellular phenotype to the list of molecular and cellular traits known to be altered by aneuploidy and suggest that chromosome missegregation can provide a quick, heritable, and reversible mechanism by which organisms can toggle between phenotypes.
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http://dx.doi.org/10.1073/pnas.1301047110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3725063PMC
July 2013

Asexual populations of the human malaria parasite, Plasmodium falciparum, use a two-step genomic strategy to acquire accurate, beneficial DNA amplifications.

PLoS Pathog 2013 23;9(5):e1003375. Epub 2013 May 23.

Department of Chemistry, University of Washington, Seattle, Washington, United States of America.

Malaria drug resistance contributes to up to a million annual deaths. Judicious deployment of new antimalarials and vaccines could benefit from an understanding of early molecular events that promote the evolution of parasites. Continuous in vitro challenge of Plasmodium falciparum parasites with a novel dihydroorotate dehydrogenase (DHODH) inhibitor reproducibly selected for resistant parasites. Genome-wide analysis of independently-derived resistant clones revealed a two-step strategy to evolutionary success. Some haploid blood-stage parasites first survive antimalarial pressure through fortuitous DNA duplications that always included the DHODH gene. Independently-selected parasites had different sized amplification units but they were always flanked by distant A/T tracks. Higher level amplification and resistance was attained using a second, more efficient and more accurate, mechanism for head-to-tail expansion of the founder unit. This second homology-based process could faithfully tune DNA copy numbers in either direction, always retaining the unique DNA amplification sequence from the original A/T-mediated duplication for that parasite line. Pseudo-polyploidy at relevant genomic loci sets the stage for gaining additional mutations at the locus of interest. Overall, we reveal a population-based genomic strategy for mutagenesis that operates in human stages of P. falciparum to efficiently yield resistance-causing genetic changes at the correct locus in a successful parasite. Importantly, these founding events arise with precision; no other new amplifications are seen in the resistant haploid blood stage parasite. This minimizes the need for meiotic genetic cleansing that can only occur in sexual stage development of the parasite in mosquitoes.
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http://dx.doi.org/10.1371/journal.ppat.1003375DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3662640PMC
December 2013