RESUMO
The prolyl-tRNA synthetase (PRS) is a validated drug target for febrifugine and its synthetic analog halofuginone (HFG) against multiple apicomplexan parasites including Plasmodium falciparum and Toxoplasma gondii. Here, a novel ATP-mimetic centered on 1-(pyridin-4-yl) pyrrolidin-2-one (PPL) scaffold has been validated to bind to Toxoplasma gondii PRS and kill toxoplasma parasites. PPL series exhibited potent inhibition at the cellular (T. gondii parasites) and enzymatic (TgPRS) levels compared to the human counterparts. Cell-based chemical mutagenesis was employed to determine the mechanism of action via a forward genetic screen. Tg-resistant parasites were analyzed with wild-type strain by RNA-seq to identify mutations in the coding sequence conferring drug resistance by computational analysis of variants. DNA sequencing established two mutations, T477A and T592S, proximal to terminals of the PPL scaffold and not directly in the ATP, tRNA, or L-pro sites, as supported by the structural data from high-resolution crystal structures of drug-bound enzyme complexes. These data provide an avenue for structure-based activity enhancement of this chemical series as anti-infectives.
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Aminoacil-tRNA Sintetases , Toxoplasma , Toxoplasmose , Humanos , Toxoplasma/genética , Descoberta de Drogas , Aminoacil-tRNA Sintetases/genética , Trifosfato de AdenosinaRESUMO
The genomes of the malaria-causing Plasmodium parasites encode a protein fused of 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) and dihydropteroate synthase (DHPS) domains that catalyze sequential reactions in the folate biosynthetic pathway. Whereas higher organisms derive folate from their diet and lack the enzymes for its synthesis, most eubacteria and a number of lower eukaryotes including malaria parasites synthesize tetrahydrofolate via DHPS. Plasmodium falciparum (Pf) and Plasmodium vivax (Pv) HPPK-DHPSs are currently targets of drugs like sulfadoxine (SDX). The SDX effectiveness as an antimalarial drug is increasingly diminished by the rise and spread of drug-resistant mutations. Here, we present the crystal structure of PvHPPK-DHPS in complex with four substrates/analogs, revealing the bifunctional PvHPPK-DHPS architecture in an unprecedented state of enzymatic activation. SDX's effect on HPPK-DHPS is due to 4-amino benzoic acid (pABA) mimicry, and the PvHPPK-DHPS structure sheds light on the SDX-binding cavity, as well as on mutations that effect SDX potency. We mapped five dominant drug resistance mutations in PvHPPK-DHPS: S382A, A383G, K512E/D, A553G, and V585A, most of which occur individually or in clusters proximal to the pABA-binding site. We found that these resistance mutations subtly alter the intricate enzyme/pABA/SDX interactions such that DHPS affinity for pABA is diminished only moderately, but its affinity for SDX is changed substantially. In conclusion, the PvHPPK-DHPS structure rationalizes and unravels the structural bases for SDX resistance mutations and highlights architectural features in HPPK-DHPSs from malaria parasites that can form the basis for developing next-generation anti-folate agents to combat malaria parasites.
Assuntos
Di-Hidropteroato Sintase/química , Difosfotransferases/química , Malária Vivax/tratamento farmacológico , Plasmodium vivax/química , Sulfadoxina/química , Aminoácidos/química , Aminoácidos/genética , Cristalografia por Raios X , Di-Hidropteroato Sintase/genética , Difosfotransferases/genética , Resistência a Medicamentos/genética , Humanos , Malária Vivax/parasitologia , Mutação , Plasmodium falciparum , Plasmodium vivax/genética , Plasmodium vivax/patogenicidade , Sulfadoxina/uso terapêutico , Tetra-Hidrofolatos/químicaRESUMO
Cladosporin (CLD) is a fungal metabolite that kills the malaria parasite via inhibiting its cytoplasmic lysyl-tRNA synthetase (KRS) and abrogating protein translation. Here we provide structural and drug selectivity analyses on CLD interacting residues in apo and holo KRSs from Plasmodium falciparum, Homo sapiens, Cryptosporidium parvum, and Mycobacterium ulcerans. We show that both gross and subtle alterations in protein backbone and sidechains drive the active site structural plasticity that allows integration of CLD in KRSs. The ligand-induced fit of CLD in PfKRS is marked by closure and stabilization of three disordered loops and one alpha helix. However, these structural rearragements are not evident in KRS-CLD complexes from H. sapiens, C. parvum, or M. ulcerans. Strikingly, CLD fits into the MuKRS active site due to a remarkable rotameric alteration in its clash-prone methionine residue that provides accommodation for the methyl moiety in CLD. Although the high concentrations of drugs used for Hs, Cp, and MuKRS-CLD complexes in co-crystallization studies enable elucidation of a structural framework for understanding drug binding in KRSs, we propose that these data should be concurrently assessed via biochemical studies of potency and drug selectivity given the poor cell-based activity of CLD against human and bacterial cells. Our comprehensive analyses of KRS-CLD interactions, therefore, highlight vital issues in structure-based drug discovery studies.
Assuntos
Isocumarinas/metabolismo , Lisina-tRNA Ligase/metabolismo , Plasmodium falciparum/enzimologia , Cryptosporidium parvum/enzimologia , Isocumarinas/química , Lisina-tRNA Ligase/química , Mycobacterium ulcerans/enzimologia , Ligação ProteicaRESUMO
The γ-secretase protease and associated regulated intramembrane proteolysis play an important role in controlling receptor-mediated intracellular signaling events, which have a central role in Alzheimer disease, cancer progression, and immune surveillance. An increasing number of γ-secretase substrates have a role in cytokine signaling, including the IL-6 receptor, IL-1 receptor type I, and IL-1 receptor type II. In this study, we show that following TNF-converting enzyme-mediated ectodomain shedding of TNF type I receptor (TNFR1), the membrane-bound TNFR1 C-terminal fragment is subsequently cleaved by γ-secretase to generate a cytosolic TNFR1 intracellular domain. We also show that clathrin-mediated internalization of TNFR1 C-terminal fragment is a prerequisite for efficient γ-secretase cleavage of TNFR1. Furthermore, using in vitro and in vivo model systems, we show that in the absence of presenilin expression and γ-secretase activity, TNF-mediated JNK activation was prevented, assembly of the TNFR1 pro-apoptotic complex II was reduced, and TNF-induced apoptosis was inhibited. These observations demonstrate that TNFR1 is a γ-secretase substrate and suggest that γ-secretase cleavage of TNFR1 represents a new layer of regulation that links the presenilins and the γ-secretase protease to pro-inflammatory cytokine signaling.
Assuntos
Secretases da Proteína Precursora do Amiloide/metabolismo , Apoptose , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Animais , Linhagem Celular , Quimiocina CXCL1/metabolismo , Ativação Enzimática , Deleção de Genes , Humanos , MAP Quinase Quinase 4/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Presenilinas/genética , Presenilinas/metabolismo , Proteólise , Transdução de SinaisRESUMO
Trimethylamine (TMA) is a tertiary amine with a characteristic fishy odour. It is synthesised from dietary constituents, including choline, L-carnitine, betaine and lecithin by the action of microbial enzymes during both healthy and diseased conditions in humans. Trimethylaminuria (TMAU) is a disease typified by its association with the characteristic fishy odour because of decreased TMA metabolism and excessive TMA excretion. Besides TMAU, a number of other diseases are associated with abnormal levels of TMA, including renal disorders, cancer, obesity, diabetes, cardiovascular diseases and neuropsychiatric disorders. Aside from its role in pathobiology, TMA is a precursor of trimethylamine-N-oxide that has been associated with an increased risk of athero-thrombogenesis. Additionally, TMA is a major air pollutant originating from vehicular exhaust, food waste and animal husbandry industry. The adverse effects of TMA need to be monitored given its ubiquitous presence in air and easy absorption through human skin. In this review, we highlight multifaceted attributes of TMA with an emphasis on its physiological, pathological and environmental impacts. We propose a clinical surveillance of human TMA levels that can fully assess its role as a potential marker of microbial dysbiosis-based diseases.
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Aterosclerose/metabolismo , Erros Inatos do Metabolismo/metabolismo , Metilaminas/urina , Neoplasias/metabolismo , Insuficiência Renal/metabolismo , Trombose/metabolismo , Poluentes Atmosféricos/análise , Poluentes Atmosféricos/metabolismo , Animais , Aterosclerose/patologia , Dieta , Humanos , Erros Inatos do Metabolismo/patologia , Metilaminas/análise , Metilaminas/metabolismo , Neoplasias/patologia , Odorantes/análise , Insuficiência Renal/patologia , Fatores de Risco , Trombose/patologiaRESUMO
Aminoacyl-tRNA synthetases (aaRSs) are essential enzymes in protein translation machinery that provide the charged tRNAs needed for protein synthesis. Over the past decades, aaRSs have been studied as anti-parasitic, anti-bacterial, and anti-fungal drug targets. This study focused on the cytoplasmic glutamyl-tRNA synthetase (GluRS) from Plasmodium falciparum, which belongs to class Ib in aaRSs. GluRS unlike most other aaRSs requires tRNA to activate its cognate amino acid substrate L-Glutamate (L-Glu), and fails to form an intermediate adenylate complex in the absence of tRNA. The crystal structures of the Apo, ATP, and ADP-bound forms of Plasmodium falciparum glutamyl-tRNA synthetase (PfGluRS) were solved at 2.1 Å, 2.2 Å, and 2.8 Å respectively. The structural comparison of the Apo- and ATP-bound holo-forms of PfGluRS showed considerable conformational changes in the loop regions around the ATP-binding pocket of the enzyme. Biophysical characterization of the PfGluRS showed binding of the enzyme substrates L-Gluand ATP.. The sequence and structural conservation were evident across GluRS compared to other species. The structural dissection of the PfGluRS gives insight into the critical residues involved in the binding of ATP substrate, which can be harvested to develop new antimalarial drugs.
Assuntos
Aminoacil-tRNA Sintetases , Glutamato-tRNA Ligase , Glutamato-tRNA Ligase/genética , Glutamato-tRNA Ligase/química , Glutamato-tRNA Ligase/metabolismo , Plasmodium falciparum/genética , Plasmodium falciparum/metabolismo , Sequência de Aminoácidos , Aminoacil-tRNA Sintetases/genética , Aminoacil-tRNA Sintetases/química , Aminoacil-tRNA Sintetases/metabolismo , RNA de Transferência/metabolismo , Trifosfato de Adenosina/metabolismoRESUMO
Purpose: Keratoconjunctivitis sicca (KCS) or dry eye disease (DED) is a multifactorial disease that results in discomfort, visual disturbance, and tear film instability with potential damage to the ocular surface. A pilot study was undertaken to determine if there were any major substantial differences in the ocular microbiome in DED patients versus healthy controls. Methods: The bacterial communities residing in the conjunctiva of patients with DED (n = 4) and healthy controls (n = 4) were assessed by 16S ribosomal RNA (rRNA) gene sequencing of the V4-V5 region. Results: The phyla Proteobacteria, Actinobacteria, Bacteroidetes, and Firmicutes were most dominant and accounted for 97% and 94.5% of all bacterial sequences in patients and controls, respectively. At the genus level, 27 bacterial genera were found with more than two-fold difference between patients and controls. Four of these - Acinetobacter, Corynebacterium, Lactobacillus, and Pseudomonas spp. - dominated the ocular microbiome of all subjects, but were proportionately lower in DED (16.5%) compared to controls (37.7%). Several bacterial genera were found to be unique in DED (34) and controls (24). Conclusion: This pilot study is an attempt to profile the ocular microbiome in patients with DED that demonstrated a higher concentration of microbial DNA compared to controls, with Firmicutes phyla dominating the bacterial population in patients with DED.
Assuntos
Síndromes do Olho Seco , Microbiota , Humanos , Projetos Piloto , Túnica Conjuntiva/microbiologia , Síndromes do Olho Seco/diagnóstico , Bactérias/genética , Lágrimas , Estudos de Casos e ControlesRESUMO
BACKGROUND: Malaria infection can result in distinct clinical outcomes from asymptomatic to severe. The association between patho-physiological changes and molecular changes in the host, and their correlation with severity of malaria progression is not fully understood. METHODS: In this study, we addressed mass spectrometry-based temporal profiling of serum metabolite levels from mice infected with Plasmodium berhgei (strain ANKA). RESULTS: We show global perturbations and identify changes in specific metabolites in correlation with disease progression. While metabolome-wide changes were apparent in late-stage malaria, a subset of metabolites exhibited highly correlated changes with disease progression. These metabolites changed early on following infection and either continued or maintained the change as mice developed severe disease. Some of these have the potential to be sentinel metabolites for severe malaria. Moreover, glycolytic metabolites, purine nucleotide precursors, tryptophan and its bioactive derivatives were many fold decreased in late-stage disease. Interestingly, uric acid, a metabolic waste reported to be elevated in severe human malaria, increased with disease progression, and subsequently appears to be detoxified into allantoin. This detoxification mechanism is absent in humans as they lack the enzyme uricase. CONCLUSIONS: We have identified candidate marker metabolites that may be of relevance in the context of human malaria.
Assuntos
Malária , Parasitos , Camundongos , Animais , Humanos , Metabolômica , Malária/parasitologia , Metaboloma , Progressão da Doença , Plasmodium bergheiRESUMO
The specificity of each aminoacyl-tRNA synthetase (aaRS) for its cognate amino acid ensures correct tRNA esterification and allows fidelity in protein synthesis. The aaRSs discriminate based on the chemical properties of their amino acid substrates and structural features of the binding pockets. In this study, we characterized aspartyl-(DRS) and asparaginyl-tRNA synthetase (NRS) from Plasmodium falciparum to determine the basis of their specificity towards L-asp and L-asn respectively. The negatively charged L-asp and its analogue L-asn differ only in their side-chain groups i.e., -OH and -NH2. Further, the amino acid binding sites are highly conserved within these two enzymes. Analysis of the substrate (L-asp/L-asn) binding sites across species revealed two highly conserved residues in PfDRS (D408 and K372) and PfNRS (E395 and L360) that are involved in recognition of the Oδ2/Nδ2 of L-asp/L-asn respectively. These residues were mutated and swapped between the D408âE in PfDRS and the corresponding E395âD in PfNRS. A similar approach was employed for residue number K372âL in PfDRS and L360âK in PfNRS. The mutated PfDRSD408E retained its enzymatic activity during step 1 of aminoacylation reaction towards L-asp and L-asn and esterified tRNAAsp with L-asp like wild type enzyme, while the PfDRSK372L was rendered enzymatically inactive. The correspondingly mutated PfNRSE395D was enzymatically inactive. The mutated PfNRSL360K had an altered specificity and esterified tRNAAsn with non-cognate amino acid L-asp and not L-asn. These data suggest that the residue K372 is crucial for the enzymatic activity of PfDRS while the residue L360 in PfNRS imparts specificity towards L-asn.
Assuntos
Aspartato-tRNA Ligase , Plasmodium falciparum , Substituição de Aminoácidos , Aminoácidos/metabolismo , Aspartato-tRNA Ligase/química , Aspartato-tRNA Ligase/genética , Aspartato-tRNA Ligase/metabolismo , Plasmodium falciparum/genética , Plasmodium falciparum/metabolismo , RNA de Transferência/metabolismo , Aminoacil-RNA de Transferência , Especificidade por SubstratoRESUMO
Background: In 2012, the World Health Organization (WHO) released the Global Plan for Insecticide Resistance Management in malaria vectors to stress the need to address insecticide resistance. In a prospective multi-centric study commissioned by the Indian Council of Medical Research (ICMR), we assessed the insecticide susceptibility status of the primary malaria vectors in India from 2017 through 2019. Methods: The insecticide susceptibility status of the prevalent primary malaria vectors - An. culicifacies, An. fluviatilis, An. stephensi, An. minimus and An. baimaii and secondary malaria vectors - An. aconitus, An. annularis and An. philippinensis/nivepes from 328 villages in 79 districts of 15 states of India were assessed following the WHO method mainly to insecticides used in vector control, organochlorine (DDT), organophosphate (malathion), and other pyrethroids (alpha-cypermethrin, cyfluthrin, lambda-cyhalothrin and permethrin). The study sites were selected as suggested by the National Vector Borne Disease Control Programme. Results: The primary malaria vector An. culicifacies showed resistance to DDT (50/50 districts including two districts of Northeastern India), malathion (27/44 districts), and deltamethrin (17/44 districts). This species was resistant to DDT alone in 19 districts, double resistant to DDT-malathion in 16 districts, double resistant to DDT-deltamethrin in 6 districts, and triple resistant to DDT-malathion-deltamethrin in 9 districts. An. minimus and An. baimaii were susceptible in Northeastern India while An. fluviatilis and the secondary malaria vector An. annularis was resistant to DDT in Jharkhand. Conclusion: In this study we report that among the primary vectors An. culicifacies is predominantly resistant to multiple insecticides. Our data suggest that periodic monitoring of insecticide susceptibility is vital. The national malaria program can take proactive steps for insecticide resistance management to continue its push toward malaria elimination in India.
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Malaria is a parasitic illness caused by the genus Plasmodium from the apicomplexan phylum. Five plasmodial species of P. falciparum (Pf), P. knowlesi, P. malariae, P. ovale, and P. vivax (Pv) are responsible for causing malaria in humans. According to the World Malaria Report 2020, there were 229 million cases and ~ 0.04 million deaths of which 67% were in children below 5 years of age. While more than 3 billion people are at risk of malaria infection globally, antimalarial drugs are their only option for treatment. Antimalarial drug resistance keeps arising periodically and thus threatens the main line of malaria treatment, emphasizing the need to find new alternatives. The availability of whole genomes of P. falciparum and P. vivax has allowed targeting their unexplored plasmodial enzymes for inhibitor development with a focus on multistage targets that are crucial for parasite viability in both the blood and liver stages. Over the past decades, aminoacyl-tRNA synthetases (aaRSs) have been explored as anti-bacterial and anti-fungal drug targets, and more recently (since 2009) aaRSs are also the focus of antimalarial drug targeting. Here, we dissect the structure-based knowledge of the most advanced three aaRSs-lysyl- (KRS), prolyl- (PRS), and phenylalanyl- (FRS) synthetases in terms of development of antimalarial drugs. These examples showcase the promising potential of this family of enzymes to provide druggable targets that stall protein synthesis upon inhibition and thereby kill malaria parasites selectively.
Assuntos
Aminoacil-tRNA Sintetases/química , Antimaláricos/química , Inibidores Enzimáticos/química , Lisina-tRNA Ligase/química , Fenilalanina-tRNA Ligase/química , Plasmodium falciparum/efeitos dos fármacos , Proteínas de Protozoários/química , Aminoacil-tRNA Sintetases/antagonistas & inibidores , Aminoacil-tRNA Sintetases/genética , Aminoacil-tRNA Sintetases/metabolismo , Antimaláricos/farmacologia , Domínio Catalítico , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Expressão Gênica , Humanos , Lisina-tRNA Ligase/antagonistas & inibidores , Lisina-tRNA Ligase/genética , Lisina-tRNA Ligase/metabolismo , Malária Falciparum/tratamento farmacológico , Malária Falciparum/parasitologia , Modelos Moleculares , Fenilalanina-tRNA Ligase/antagonistas & inibidores , Fenilalanina-tRNA Ligase/genética , Fenilalanina-tRNA Ligase/metabolismo , Plasmodium falciparum/química , Plasmodium falciparum/enzimologia , Plasmodium falciparum/genética , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Proteínas de Protozoários/antagonistas & inibidores , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologiaRESUMO
BACKGROUND: Mosquito-borne diseases have a devastating impact on human civilization. A few species of Anopheles mosquitoes are responsible for malaria transmission, and while there has been a reduction in malaria-related deaths worldwide, growing insecticide resistance is a cause for concern. Aedes mosquitoes are known vectors of viral infections, including dengue, yellow fever, chikungunya, and Zika. Aminoacyl-tRNA synthetases (aaRSs) are key players in protein synthesis and are potent anti-infective drug targets. The structure-function activity relationship of aaRSs in mosquitoes (in particular, Anopheles and Aedes spp.) remains unexplored. METHODS: We employed computational techniques to identify aaRSs from five different mosquito species (Anopheles culicifacies, Anopheles stephensi, Anopheles gambiae, Anopheles minimus, and Aedes aegypti). The VectorBase database ( https://vectorbase.org/vectorbase/app ) and web-based tools were utilized to predict the subcellular localizations (TargetP-2.0, UniProt, DeepLoc-1.0), physicochemical characteristics (ProtParam), and domain arrangements (PfAM, InterPro) of the aaRSs. Structural models for prolyl (PRS)-, and phenylalanyl (FRS)-tRNA synthetases-were generated using the I-TASSER and Phyre protein modeling servers. RESULTS: Among the vector species, a total of 37 (An. gambiae), 37 (An. culicifacies), 37 (An. stephensi), 37 (An. minimus), and 35 (Ae. aegypti) different aaRSs were characterized within their respective mosquito genomes. Sequence identity amongst the aaRSs from the four Anopheles spp. was > 80% and in Ae. aegypti was > 50%. CONCLUSIONS: Structural analysis of two important aminoacyl-tRNA synthetases [prolyl (PRS) and phenylanalyl (FRS)] of Anopheles spp. suggests structural and sequence similarity with potential antimalarial inhibitor [halofuginone (HF) and bicyclic azetidine (BRD1369)] binding sites. This suggests the potential for repurposing of these inhibitors against the studied Anopheles spp. and Ae. aegypti.
Assuntos
Aedes/efeitos dos fármacos , Aminoacil-tRNA Sintetases/antagonistas & inibidores , Anopheles/efeitos dos fármacos , Dengue/transmissão , Inseticidas/farmacologia , Malária/transmissão , Mosquitos Vetores/efeitos dos fármacos , Aedes/enzimologia , Aedes/genética , Sequência de Aminoácidos , Aminoacil-tRNA Sintetases/química , Aminoacil-tRNA Sintetases/genética , Animais , Anopheles/enzimologia , Anopheles/genética , Sistemas de Liberação de Medicamentos , Descoberta de Drogas , Genômica , Humanos , Resistência a Inseticidas , Modelos Estruturais , Mosquitos Vetores/enzimologia , Mosquitos Vetores/genética , Alinhamento de Sequência , Relação Estrutura-AtividadeRESUMO
COVID-19 is an acute respiratory distress syndrome and is often accompanied by gastrointestinal symptoms. The SARS-CoV-2 has been traced not only in nasopharyngeal and mid-nasal swabs but also in stool and rectal swabs of COVID-19 patients. The gut microbiota is important for an effective immune response as it ensures that unfavorable immune reactions in lungs and other vital organs are regulated. The human gut-lung microbiota interplay provides a framework for therapies in the treatment and management of several pulmonary diseases and infections. Here, we have collated data from COVID-19 studies, which suggest that bacterial co-infections as well as the gut-lung cross talk may be important players in COVID-19 disease prognosis. Our analyses suggests a role of gut microbiome in pathogen infections as well as in an array of excessive immune reactions during and post COVID-19 infection recovery period.
RESUMO
To counter the coronavirus disease 2019 (COVID-19) pandemic, each country must design sustainable control plans given the inherent disparities in wealth and healthcare systems. Most malaria-endemic countries run well-entrenched malaria control programs via their established frameworks for diagnosis, case management, treatment and overall surveillance. We propose that the malaria control infrastructures can be partially co-opted for launching sustainable COVID-19 mitigation plans.
Assuntos
COVID-19/prevenção & controle , Atenção à Saúde , Planejamento em Saúde , Malária/prevenção & controle , Pandemias , COVID-19/epidemiologia , Doenças Endêmicas , Programas Governamentais , Humanos , SARS-CoV-2RESUMO
The global spread of sulfadoxine (Sdx, S) and pyrimethamine (Pyr, P) resistance is attributed to increasing number of mutations in DHPS and DHFR enzymes encoded by malaria parasites. The association between drug resistance mutations and SP efficacy is complex. Here we provide an overview of the geographical spread of SP resistance mutations in Plasmodium falciparum (Pf) and Plasmodium vivax (Pv) encoded dhps and dhfr genes. In addition, we have collated the mutation data and mapped it on to the three-dimensional structures of DHPS and DHFR which have become available. Data from genomic databases and 286 studies were collated to provide a comprehensive landscape of mutational data from 2005 to 2019. Our analyses show that the Pyr-resistant double mutations are widespread in Pf/PvDHFR (P. falciparum â¼61% in Asia and the Middle East, and in the Indian sub-continent; in P. vivax â¼33% globally) with triple mutations prevailing in Africa (â¼66%) and South America (â¼33%). For PfDHPS, triple mutations dominate South America (â¼44%), Asia and the Middle East (â¼34%) and the Indian sub-continent (â¼27%), while single mutations are widespread in Africa (â¼45%). Contrary to the status for P. falciparum, Sdx-resistant single point mutations in PvDHPS dominate globally. Alarmingly, highly resistant quintuple and sextuple mutations are rising in Africa (PfDHFR-DHPS) and Asia (Pf/PvDHFR-DHPS). Structural analyses of DHFR and DHPS proteins in complexes with substrates/drugs have revealed that resistance mutations map proximal to Sdx and Pyr binding sites. Thus new studies can focus on discovery of novel inhibitors that target the non-substrate binding grooves in these two validated malaria parasite drug targets.
Assuntos
Antimaláricos , Plasmodium falciparum/efeitos dos fármacos , Antimaláricos/farmacologia , Di-Hidropteroato Sintase/genética , Combinação de Medicamentos , Resistência a Medicamentos , Genótipo , Mutação , Plasmodium falciparum/genética , Pirimetamina/farmacologia , Sulfadoxina/farmacologia , Tetra-Hidrofolato Desidrogenase/genéticaRESUMO
We describe the epidemiological characteristics and associated risk factors of those presenting at a large testing centre for SARS-CoV-2 infection. This is a retrospective record review of individuals who underwent SARS-CoV-2 testing by reverse transcription-polymerase chain reaction (RT-PCR) at a high-throughput national-level government facility located in the north of India. Samples collected from 6 April to 31 December 2020 are included in this work and represent four highly populous regions. Additionally, there was a prospective follow-up of 1729 cases through telephone interviews from 25 May 2020 to 20 June 2020. Descriptive analysis has been performed for profiling clinic-epidemiological aspects of suspect cases. Multivariable logistic regression analysis was undertaken to determine risk factors that are associated with SARS-CoV-2 test positivity and symptom status. A total of 125 600 participants' details have been included in this report. The mean (s.d.) age of the participants was 33.1 (±15.3) years and 66% were male. Among these tested, 9515 (7.6%) were positive for COVID-19. A large proportion of positive cases were asymptomatic. In symptomatic positive cases, the commonest symptoms were cough and fever. Increasing age (groups 20-59 and ≥60 years compared to age group less than 5 years), male sex, history of international travel, symptoms for SARS-CoV-2, and participants from Delhi and Madhya Pradesh were positively associated with SARS-CoV-2 test positivity. Having co-morbidity, risk behaviours and intra-familial positivity were associated with a positive odds ratio for exhibiting SARS-CoV-2 symptoms. Intensified testing and isolation of cases, identification of both asymptomatic and symptomatic individuals and additional care of those with co-morbidities and risk behaviours will all be collectively important for disease containment in India. Reasons for differentials in testing between men and women remain an important area for in-depth study. The increased deployment of vaccines is likely to impact the trajectory of COVID-19 in the coming time, and therefore our data will serve as a comparative resource as India experiences the second wave of infection in light of newer variants that are likely to accelerate disease spread.
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Teste de Ácido Nucleico para COVID-19/estatística & dados numéricos , COVID-19/epidemiologia , Adolescente , Adulto , Fatores Etários , Idoso , Comorbidade , Feminino , Humanos , Índia , Masculino , Pessoa de Meia-Idade , Fatores de Risco , Fatores Sexuais , Viagem/estatística & dados numéricosRESUMO
BACKGROUND: A surveillance system is the foundation for disease prevention and control. Malaria surveillance is crucial for tracking regional and temporal patterns in disease incidence, assisting in recorded details, timely reporting, and frequency of analysis. OBJECTIVE: In this study, we aim to develop an integrated surveillance graphical app called FeverTracker, which has been designed to assist the community and health care workers in digital surveillance and thereby contribute toward malaria control and elimination. METHODS: FeverTracker uses a geographic information system and is linked to a web app with automated data digitization, SMS text messaging, and advisory instructions, thereby allowing immediate notification of individual cases to district and state health authorities in real time. RESULTS: The use of FeverTracker for malaria surveillance is evident, given the archaic paper-based surveillance tools used currently. The use of the app in 19 tribal villages of the Dhalai district in Tripura, India, assisted in the surveillance of 1880 suspected malaria patients and confirmed malaria infection in 93.4% (114/122; Plasmodium falciparum), 4.9% (6/122; P vivax), and 1.6% (2/122; P falciparum/P vivax mixed infection) of cases. Digital tools such as FeverTracker will be critical in integrating disease surveillance, and they offer instant data digitization for downstream processing. CONCLUSIONS: The use of this technology in health care and research will strengthen the ongoing efforts to eliminate malaria. Moreover, FeverTracker provides a modifiable template for deployment in other disease systems.
RESUMO
Scaffold modules known as aminoacyl-tRNA synthetase (aaRS)-interacting multifunctional proteins (AIMPs), such as AIMP1/p43, AIMP2/p38 and AIMP3/p18, are important in driving the assembly of multi-aaRS (MARS) complexes in eukaryotes. Often, AIMPs contain an N-terminal glutathione S-transferase (GST)-like domain and a C-terminal OB-fold tRNA-binding domain. Recently, the apicomplexan-specific Plasmodium falciparum p43 protein (Pfp43) has been annotated as an AIMP and its tRNA binding, tRNA import and membrane association have been characterized. The crystal structures of both the N- and C-terminal domains of the Plasmodium vivax p43 protein (Pvp43), which is an ortholog of Pfp43, have been resolved. Analyses reveal the overall oligomeric structure of Pvp43 and highlight several notable features that show Pvp43 to be a soluble, cytosolic protein. The dimeric assembly of the N-terminal GST-like domain of Pvp43 differs significantly from canonical GST dimers, and it is tied to the C-terminal tRNA-binding domain via a linker region. This work therefore establishes a framework for dissecting the additional roles of p43 orthologs in eukaryotic multi-protein MARS complexes.
Assuntos
Plasmodium vivax/química , Proteínas de Protozoários/química , Cristalografia por Raios X , Ligação Proteica , Domínios Proteicos , Multimerização Proteica , RNA de Transferência/metabolismoRESUMO
Artemisinin-based combination therapy (ACT) offers highly successful treatment of malaria. Emergence and spread of Plasmodium falciparum (Pf) parasites with decreased susceptibility to ACT in South-East Asia has caused concern worldwide. The current accepted criteria to assess artemisinin (ART) resistance relies upon data on treatment failure, delayed parasite clearance at day 3 (DPC3), parasite clearance half-life (PCHL) and in-vitro/ex-vivo ring stage survival assays (RSAs). Interestingly, some studies suggest that DPC3 does not provide a distinct separation between ART sensitive/resistant strains, and RSA differences may also be inconclusive. More recently, recrudescence of ART treated Pf, independent of the presence of Kelch 13 (K13) mutation (C580Y), has been reported in the monkey malaria model suggesting that genes other than K13 like coronin, dhps, dhfr, crt, mdr1 and plasmepsin1 may contribute towards ACT failure. Here we have collated the distribution of K13 mutants from Pf strains in South Asia. A total of fifty Pf-K13 mutations have been studied for ART resistance in South Asia of which nine have been validated while eleven are potentials for ART resistance. The remaining thirty K13 mutations have been reported from various locations in South Asia but lack corroborative clinical data on ART resistance/ACT failure. Of the fifty, fourteen K13 mutations have been identified in India including four novel mutations (S549Y, G625R, N657H, D702N). Structural mapping of these K13 mutations does not offer any coherent explanation for their contribution towards ART resistance as they are scattered in the K13 structure. Thus, K13 mutations likely provide only a partial synopsis, and we propose that all suspect cases of ACT failure be assessed by: 1) DPC3, 2) PCHL, 3) in-vitro/ex-vivo RSAs and 4) GWAS data in an effort to annotate the resistance status of the parasites. These efforts may help in surveillance and containment of ART resistance/ACT failure in South Asia.
Assuntos
Antimaláricos/farmacologia , Artemisininas/farmacologia , Resistência a Medicamentos/genética , Repetição Kelch/genética , Mutação , Plasmodium falciparum/genética , Antimaláricos/uso terapêutico , Artemisininas/uso terapêutico , Ásia/epidemiologia , Quimioterapia Combinada , Doenças Endêmicas , Índia/epidemiologia , Malária Falciparum/tratamento farmacológico , Malária Falciparum/epidemiologia , Malária Falciparum/parasitologia , Plasmodium falciparum/efeitos dos fármacosRESUMO
BACKGROUND: A century ago, pantheras were abundant across Asia. Illegal hunting and trading along with loss of habitat have resulted in the designation of Panthera as a genus of endangered species. In addition to the onslaught from humans, pantheras are also susceptible to outbreaks of several infectious diseases, including babesiosis. The latter is a hemoprotozoan disease whose causative agents are the eukaryotic parasites of the apicomplexan genus Babesia. Babesiosis affects a varied range of animals including humans (Homo sapiens), bovines (e.g. Bos taurus), pantheras (e.g. Panthera tigris, P. leo, P. pardus) and equines. Babesia spp. are transmitted by the tick vector Ixodes scapularis or ticks of domestic animals, namely Rhipicephalus (Boophilus) microplus and R. (B.) decoloratus. At the level of protein translation within these organisms, the conserved aminoacyl tRNA synthetase (aaRS) family offers an opportunity to identify the sequence and structural differences in the host (Panthera) and parasites (Babesia spp.) in order to exploit these for drug targeting Babesia spp. METHODS: Using computational tools we investigated the genomes of Babesia spp. and Panthera tigris so as to annotate their aaRSs. The sequences were analysed and their subcellular localizations were predicted using Target P1.1, SignalP 3.0, TMHMM v.2.0 and Deeploc 1.0 web servers. Structure-based analysis of the aaRSs from P. tigris and its protozoan pathogens Babesia spp. was performed using Phyre2 and chimera. RESULTS: We identified 33 (B. bovis), 34 (B. microti), 33 (B. bigemina) and 33 (P. tigris) aaRSs in these respective organisms. Poor sequence identity (~ 20-50%) between aaRSs from Babesia spp. and P. tigris was observed and this merits future experiments to validate new drug targets against Babesia spp. CONCLUSIONS: Overall this work provides a foundation for experimental investigation of druggable aaRSs from Babesia sp. in an effort to control Babesiosis in Panthera.