Protozoan Parasitism: From Omics to Prevention and Control | Book
Caister Academic Press
Luis Miguel de Pablos Torró1,2
and Jacob-Lorenzo Morales3
1Department of Parasitology, Universidad de Granada, Severo Ochoa s/n, 18001 Granada, Spain; 2Institute of Biotechnology, University of Granada, 18071 Granada, Spain; 3Inst. Univ. De Enfermedades Tropicales Y Salud Pública de Canarias, Universidad de La Laguna, San Cristóbal de La Laguna, Santa Cruz de Tenerife 38200, Spain
vi + 182
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Over the last fifteen years a large collection of protozoan parasite genomes have been sequenced, annotated and made available on public databases. This wealth of information has stimulated and fuelled new research into vaccine and drug discovery, new approaches for high-throughput experimentation and the genetic engeneering of key protozoan parasites.
In this timely book expert international authors review the current hot-topics in this area to provide an up-to-date overview. The book is divided into two parts. The first part is dedicated to Trypanosoma cruzi, an organism not only responsible for a serious tropical disease but also a good model system for fundamental and applied biology research. Part two of the book is dedicated to four other important protozoa: Leishmania, Naegleria, Entamoeba and Acanthamoeba. Chapters are written from a molecular biology and genomics perspective and focus on a common goal: the development of new strategies for the control and prevention of infection.
This volume is essential reading for anyone working on protozoan parasites in academia, government, and biotechnology companies.
Table of contents
1. From proteins to molecular targets: Trypanosoma cruzi proteomic insights in drug development
Giselle V-F Brunoro, Marcelle A Caminha and Rubem FS Menna-Barreto
Chagas disease is a neglected disease caused by the hemoflagellate protozoan Trypanosoma cruzi, which is endemic to Latin America, that emerges in non-endemic areas due to immigration. The parasite presents a complex life cycle, including two hosts (invertebrate and vertebrate) and three developmental stages (epimastigotes, trypomastigotes and amastigotes). The current chemotherapy is restricted to two nitroheterocycles, nifurtimox and benznidazole, which present limited efficacy depending on the disease phase. Multidisciplinary efforts have been made to identify promising novel parasite-specific key molecules to develop nitroderivative substitutes, increase trypanocidal activity and minimize host toxicity. In this context, deep analysis of drug pharmacokinetics and mechanisms of action is crucial to improve specificity and safety. We reviewed proteomics as a valuable tool for the identification of molecular targets of T. cruzi. Advances in parasite proteomic maps (different developmental stages and strains) were reviewed, emphasizing the promising targets, their biological applications, and possible follow-up analysis for the development of novel anti-T. cruzi drugs.
2. High throughput screenings: new breeze in anti-Trypanosoma cruzi drug discovery
Julio Alonso-Padilla and Ana Rodríguez
Development of therapeutic solutions for Chagas disease, which is caused by the protozoan Trypanosoma cruzi, has been historically neglected. Despite being the parasitic disease of highest impact in America, there is no vaccine in sight and the only two available drugs for treatment, benznidazole and nifurtimox, display severe toxicity. These drugs have good efficacy against acute T. cruzi infections and are relatively well tolerated by young patients. Nonetheless, the acute stage of the disease is mostly asymptomatic and largely remains undiagnosed and untreated. Cardiac and/or digestive life-threatening symptoms arise in the chronic stage, which is when diagnosis is generally achieved. At this stage, the efficacy of current drugs is diminished and their side effects prompt to ~20% of treatment discontinuation. Thus there is an urgent need for more efficacious and less toxic therapies for the estimated 7 million chronically infected patients. To meet this challenge, massive screenings have been recently performed over large chemical collections to identify new chemical structures fitted with anti-T. cruzi specific activity. The parasite´s biological complexity and the lack of detailed metabolic and biochemical information have favored a phenotypic over a target-based drug discovery strategy. The first fruits of these high throughput screening campaigns are now arriving at the pre-clinical steps of the drug developmental pathway.
3. Protein glycosylation in Trypanosoma cruzi and mass spectrometry-based strategies for glycan and glycoprotein characterization
Rebeca Kawahara, Joyce Saad, Walter Colli, Maria JM Alves and Giuseppe Palmisano
Trypanosoma cruzi is the protozoan parasite that causes Chagas disease. This parasite is genetically highly diverse and has a complex life cycle involving two different hosts and distinct developmental stages. The surface of the parasite is covered by a dense array of glycoproteins and glycolipids, which play important function in the differentiation processes and in the host-parasite interaction. The N- and O-glycan biosynthesis in T. cruzi has important features that involve unusual carbohydrates and glycosylation enzymes. The characterization of protein glycosylation in T. cruzi has unprecedented promise for the discovery of novel targets for drug design and therapeutic intervention in Chagas disease as well as to characterize the different T. cruzi strains. However, analytical methodologies for large-scale protein glycosylation analysis have faced challenges due to the extensive glycan micro- and macroheterogeneity. This chapter aims to provide an historical overview of key findings in T. cruzi protein glycosylation pathway, including the intermediates and enzymes involved especially in N-linked glycan biosynthesis and the analytical tools used to uncover the composition or structure of N and O-linked glycans. Moreover, we described how mass spectrometry is a powerful tool for large-scale characterizing the set of glycans and glycoproteins in T. cruzi.
4. High-throughput genetic engineering in Trypanosoma cruzi
Luis Miguel de Pablos
There are several reasons supporting the fact that kinetoplastid parasites are good models for solving biological questions: i) these parasites are among the most ancient and divergent eukaryotes, ii) the pathogenicity is dictated by dynamic cell/protein structures specific to this order of parasites and exposed to the host immune system, iii) the life cycle of this parasites can be replicated in vitro or in vivo to a nice extend iv) Their life cycles combine drastic developmental changes in response to environmental cues v) Kinetoplastid parasites can survive as extracellular or intracellular forms in a myriad of host including insects, plants or mammals. Thus, the versatility of scenarios faced by those cells makes them interesting to deep studies using genetic engineering. The emergency of new reverse genetic techniques such us gene editing using nucleases or new approaches for gene flanking homology DNA assembly (fusion PCR or Gibson assembly) for precise integration of different tags have been recently used and applied in the field of trypanosomatids. The aim of this chapter will be to review on advances and approaches designed for successful genetic engeneering on Trypanosoma cruzi.
5. Omics approaches for understanding gene expression in Leishmania: clues for tackling leishmaniasis
Jose M Requena, Pedro J Alcolea, Ana Alonso and Vicente Larraga
Leishmaniases, a group of parasitic diseases caused by species of the genus Leishmania, afflict millions of people across the globe and cause significant morbidity and mortality. Unfortunately, vaccine and chemotherapy options are limited. The advances in whole genome sequencing have led to renewed impetus in identifying druggable targets for future development of more effective treatments. Hence, the last decade has witnessed a revolution in our understanding of the Leishmania genomes through the completion of an increasing number of genome sequencing projects for several species and strains. However, the completion of a genome sequence is not the final product, rather it is just the beginning towards the objective of linking the wealth of data encoded in millions of bases to the biological processes of an organism. Moreover, the genome features (genomics) is only a part of the problem to be solved: the genome yields on transcription, the transcriptome, which in turn yields the proteome on translation, and ultimately the proteins either produce metabolites or are modulated by them. The size of genomics, transcriptomics, proteomics and metabolomics datasets has impelled a new way of analyzing data together with the development of potent bioinformatics tools. During their life cycles, Leishmania parasites undergo significant changes in their morphology and metabolism. These changes clearly demand a developmental regulation of differential gene expression. Moreover, it is now becoming clear that epigenetic control can also regulate other aspects of the parasitic life cycle, including the control of the switch from proliferative to developmental programs, and the adaptations required for host and cellular tropisms.
6. Natural Products in Human Leishmaniasis Therapy: Last Two Years of Research
J. Piñero, A. Lopez-Arencibia, M. Reyes-Batlle, I. Sifaoui, C. Wagner, A. Castillo, E. Córdoba-Lanús and J. Lorenzo-Morales
Leishmaniasis is a vector-borne neglected tropical disease caused by protozoan parasites of the Leishmania genus and transmitted by the female Phlebotomus and Lutzomyia sand flies. This disease affects the tropical as well as subtropical countries of the world and presents more than one clinical forms, with the visceral one considered fatal. Furthermore, this disease received limited funding for discovery, development and delivery of new tools. Chemotherapy is the major treatment option considered for the control of this disease in the infected people. The currently prescribed therapies still rely on pentavalent antimonials, pentamidine, paromomycin, liposomal amphotericin B, and miltefosine. However, their low efficacy, long-course treatment regimen, high toxicity, adverse side effects, induction of parasite resistance and high cost require the need for better drugs given that antileishmanial vaccines may not be available in the near future. An alternative to synthetic drugs is the search for anti-parasitic plant extracts or secondary metabolites derived from these sources. This chapter examines the later studies on natural bioactive compounds as sources of antileishmanicidal agents.
7. Naegleria fowleri: biology and pathogenesis
Ruqaiyyah Siddiqui and Naveed A Khan
Naegleria fowleri is a free-living protist pathogen, that can infect the central nervous system to produce fatal primary amoebic meningoencephaltis (PAM). The amoeba enters through the nasal cavity via contaminated water and reaches the the brain via olfactory neuroepithelial route to produce infection within days and almost always results in death (Symmers, 1969). A complete knowledge of this parasite, how it lives in the environment, and produces diseases is crucial for the rational development of preventative and therapeutic strategies.
8. Entamoeba histolytica: from high-throughput technology to new drugs
Anjan Debnath and Sharon L Reed
Amebiasis, caused by the protozoan parasite Entamoeba histolytica, is a major contributor to morbidity and mortality worldwide. Current therapy mainly depends on metronidazole, but it is not active against E. histolytica cysts that transmit amebiasis. Therefore, an additional luminal drug paromomycin is required for the treatment. Treatment with two drugs for 20 days leads to incomplete compliance. Moreover, in the absence of a back-up drug and a looming threat of drug resistance, the identification of new drugs for the treatment of amebiasis is a top priority. With the advent of high-throughput screening technology, we focused on developing an assay that could utilize robotics for rapid interrogation of compounds against E. histolytica. We developed a high-throughput screen for E. histolytica and used the validated assay to screen a smaller FDA-approved drug library and a larger kinase-targeted library. The screen of the kinase-targeted library identified six amebicidals and the FDA-approved drug library screen identified auranofin as amebicidal. This opened the avenue for repurposing the anti-arthritic drug, auranofin, for the treatment of amebiasis. Currently, auranofin has completed Phase I clinical trial for amebiasis and is undergoing Phase II clinical trial evaluation for the treatment of amebiasis and giardiasis.
9. Gene silencing and therapeutic targets against Acanthamoeba infections
Jacob Lorenzo-Morales, María Reyes-Batlle, Ines Sifaoui, Atteneri López-Arencibia, Olfa Chiboub, Aitor Rizo-Liendo, Carlos J Bethencourt-Estrella, Francisco Arnalich-Montiel, Pedro Rocha-Cabrera and José E Piñero
Pathogenic strains of Acanthamoeba genus are the causative agents of a sight threating infection of the cornea known as Acanthamoeba keratitis (AK). The incidence of AK has risen worldwide in the recent years which is highly related to an increase in the number of contact lens wearers, the main group of individuals at risk. Moreover, due to the ability of these amoebae to form a highly resistant cyst stage, most of the currently available treatments and contact lens maintenance solutions are not fully effective. The development of Acanthamoeba genome project, releasing the sequences of the whole genome of this pathogen as well as the availability of novel therapeutic approaches such as gene silencing using RNAi technology, has allowed the application of this approach for the elucidation of novel drug targets in Acanthamoeba. In this chapter, the advances in this field as well as current and future therapeutic approaches against AK are described.
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(EAN: 9781910190838 9781910190845 Subjects: [medical microbiology] [microbiology] [parasitology] )