In this review, we focus on the sequenced genomes of the pathogens Naegleria fowleri, Acanthamoeba spp. and Balamuthia mandrillaris, and the remarkable discoveries regarding the pathogenicity and genetic information of these organisms, using techniques related to the various omics branches like genomics, transcriptomics, and proteomics. Currently, novel data produced through comparative genomics analyses and both differential gene and protein expression in these free-living amoebas have allowed for breakthroughs to identify genes unique to N. fowleri, genes with active transcriptional activity, and their differential expression in conditions of modified virulence. Furthermore, orthologous genes of the various nuclear genomes within the Naegleria and Acanthamoeba genera have been clustered. The proteome of B. mandrillaris has been reconstructed through transcriptome data, and its mitochondrial genome structure has been thoroughly described with a unique characteristic that has come to light: a type I intron with the capacity of interrupting genes through its self-splicing ribozymes activity. With the integration of data derived from the diverse omic sciences, there is a potential approximation that reflects the molecular complexity required for the identification of virulence factors, as well as crucial information regarding the comprehension of the molecular mechanisms with which these interact. Altogether, these breakthroughs could contribute to radical advances in both the fields of therapy design and medical diagnosis in the foreseeable future.
BACKGROUND: Application des sciences de l’omique à l’étude de Naegleria fowleri, Acanthamoeba spp. et Balamuthia mandrillaris : état actuel et projections futures.
UNASSIGNED: Dans cette revue, l’accent est mis sur les génomes séquencés des agents pathogènes Naegleria fowleri, Acanthamoeba spp. et Balamuthia mandrillaris, et les découvertes remarquables concernant la pathogénicité et l’information génétique de ces organismes, en utilisant des techniques liées aux diverses branches de l’omique comme la génomique, la transcriptomique et la protéomique. Actuellement, de nouvelles données produites par des analyses génomiques comparatives et l’expression différentielle des gènes et des protéines dans ces amibes libres ont permis des percées pour identifier des gènes uniques à N. fowleri, des gènes avec une activité transcriptionnelle active et leur expression différentielle dans des conditions de virulence modifiée. En outre, les gènes orthologues des divers génomes nucléaires des genres Naegleria et Acanthamoeba ont été regroupés. Le protéome de B. mandrillaris a été reconstruit grâce aux données du transcriptome, et la structure de son génome mitochondrial décrite de manière détaillée, mettant ainsi une caractéristique unique à jour : un intron de type I avec la capacité d’interrompre les gènes par son activité d’auto-épissage des ribozymes. Avec l’intégration des données issues des diverses sciences omiques, il existe une approximation potentielle qui reflète la complexité moléculaire requise pour l’identification des facteurs de virulence, ainsi que des informations cruciales concernant la compréhension des mécanismes moléculaires avec lesquels ceux-ci interagissent. Dans l’ensemble, ces percées pourraient contribuer à des progrès notables à la fois dans les domaines de la conception de la thérapie et du diagnostic médical dans un avenir proche.

Avian malaria and related haemosporidians (Plasmodium, [Para]Haemoproteus and Leucocytoozoon) represent an exciting multihost, multiparasite system in ecology and evolution. Global research in this field accelerated after the publication in 2000 of PCR protocols to sequence a haemosporidian mitochondrial (mtDNA) barcode and the development in 2009 of an open-access database to document the geographic and host ranges of parasite mtDNA haplotypes. Isolating haemosporidian nuclear DNA from bird hosts, however, has been technically challenging, slowing the transition to genomic-scale sequencing techniques. We extend a recently developed sequence capture method to obtain hundreds of haemosporidian nuclear loci from wild bird samples, which typically have low levels of infection, or parasitemia. We tested 51 infected birds from Peru and New Mexico and evaluated locus recovery in light of variation in parasitemia, divergence from reference sequences and pooling strategies. Our method was successful for samples with parasitemia as low as ~0.02% (2 of 10,000 blood cells infected) and mtDNA divergence as high as 15.9% (one Leucocytozoonsample), and using the most cost-effective pooling strategy tested. Phylogenetic relationships estimated with >300 nuclear loci were well resolved, providing substantial improvement over the mtDNA barcode. We provide protocols for sample preparation and sequence capture including custom probe sequences and describe our bioinformatics pipeline using atram 2.0, phyluce and custom Perl/Python scripts. This approach can be applied to thousands of avian samples that have already been found to have haemosporidian infections of at least moderate intensity, greatly improving our understanding of parasite speciation, biogeography and evolutionary dynamics.

Antimalarial resistance is rapidly spreading across parts of southeast Asia where dihydroartemisinin-piperaquine is used as first-line treatment for Plasmodium falciparum malaria. The first published reports about resistance to antimalarial drugs came from western Cambodia in 2013. Here, we analyse genetic changes in the P falciparum population of western Cambodia in the 6 years before those reports.
We analysed genome sequence data on 1492 P falciparum samples from 11 locations across southeast Asia, including 464 samples collected in western Cambodia between 2007 and 2013. Different epidemiological origins of resistance were identified by haplotypic analysis of the kelch13 artemisinin resistance locus and the plasmepsin 2-3 piperaquine resistance locus.
We identified more than 30 independent origins of artemisinin resistance, of which the KEL1 lineage accounted for 140 (91%) of 154 parasites resistant to dihydroartemisinin-piperaquine. In 2008, KEL1 combined with PLA1, the major lineage associated with piperaquine resistance. By 2013, the KEL1/PLA1 co-lineage had reached a frequency of 63% (24/38) in western Cambodia and had spread to northern Cambodia.
The KEL1/PLA1 co-lineage emerged in the same year that dihydroartemisinin-piperaquine became the first-line antimalarial drug in western Cambodia and spread rapidly thereafter, displacing other artemisinin-resistant parasite lineages. These findings have important implications for management of the global health risk associated with the current outbreak of multidrug-resistant malaria in southeast Asia.
Wellcome Trust, Bill & Melinda Gates Foundation, Medical Research Council, UK Department for International Development, and the Intramural Research Program of the National Institute of Allergy and Infectious Diseases.

As a group of unicellular eukaryotes, ciliates offer a unique system to explore epigenetic regulation, mostly due to their nuclear dualism. Ciliates launched a successful radiation after their early evolutionary branching, therefore harboring an unexpectedly rich pool of diverse biological functions and mechanisms. In this review, we compare distinct features of different ciliates in mating type determination, genome organization, DNA methylation, and removal of internal eliminated sequences (IES), with emphasis on Tetrahymena, Paramecium and Oxytricha. Firstly, we review studies on mating type determination in Paramecium, one of the foundational phenomena that defined the field of epigenetics, and compare this process with that in Tetrahymena. Secondly, we showcase the high diversity in genome structure of several ciliates, such as genome size, gene copy numbers, genome rearrangement, etc. Thirdly, we present a brief description of features and potential functions of 5-methylcytosine (5mC) and N6-methyladenine (6mA) in ciliates so far studied. Fourthly, we describe both the initial and the continuously optimized scan RNA (scnRNA) model for IES elimination in Tetrahymena and contrast it with that in Paramecium and Oxytricha. Finally, we discuss the importance of integrative approaches to the study of epigenetic diversity in ciliates and provide possible directions for future research.

Chagas disease is an endemic zoonosis in Latin America and caused by the parasite Trypanosoma cruzi. This kinetoplastid displays remarkable genetic variability, allowing its classification into six Discrete Typing Units (DTUs) from TcI to TcVI. T. cruzi I presents the broadest geographical distribution in the continent and has been associated to severe forms of cardiomyopathies. Recently, a particular genotype associated to human infections has been reported and named as TcIDOM (previously named TcIa-b). This genotype shows to be clonal and adapted to the domestic cycle but so far no studies have determined the biological properties of domestic (TcIDOM) and sylvatic TcI strains (previously named TcIc-e). Hence, the aim of this study was to untangle the biological features of these genotypes in murine models. We infected ICR-CD1 mice with five TcI strains (two domestic, two sylvatic and one natural mixture) and determined the course of infection during 91 days (acute and chronic phase of the disease) in terms of parasitemia, tissue tropism, immune response (IgG titers) and tissue invasion by means of histopathology studies. Statistically significant differences were observed in terms of parasitemia curves and prepatent period between domestic (TcIDOM) and sylvatic strains. There were no differences in terms of IgG antibodies response across the mice infected with the five strains. Regarding the histopathology, our results indicate that domestic strains present higher parasitemias and low levels of histopathological damage. In contrast, sylvatic strains showed lower parasitemias and high levels of histopathological damage. These results highlight the sympatric and behavioral differences of domestic and sylvatic TcI strains; the clinical and epidemiological implications are herein discussed.

Plasmodium vivax infects a hundred million people annually and endangers 40% of the world\'s population. Unlike Plasmodium falciparum, P. vivax parasites can persist as a dormant stage in the liver, known as the hypnozoite, and these dormant forms can cause malaria relapses months or years after the initial mosquito bite. Here we analyze whole genome sequencing data from parasites in the blood of a patient who experienced consecutive P. vivax relapses over 33 months in a non-endemic country. By analyzing patterns of identity, read coverage, and the presence or absence of minor alleles in the initial polyclonal and subsequent monoclonal infections, we show that the parasites in the three infections are likely meiotic siblings. We infer that these siblings are descended from a single tetrad-like form that developed in the infecting mosquito midgut shortly after fertilization. In this natural cross we find the recombination rate for P. vivax to be 10 kb per centimorgan and we further observe areas of disequilibrium surrounding major drug resistance genes. Our data provide new strategies for studying multiclonal infections, which are common in all types of infectious diseases, and for distinguishing P. vivax relapses from reinfections in malaria endemic regions. This work provides a theoretical foundation for studies that aim to determine if new or existing drugs can provide a radical cure of P. vivax malaria.

Malaria is a major public health problem in India and one which contributes significantly to the overall malaria burden in Southeast Asia. The National Vector Borne Disease Control Program of India reported ∼1.6 million cases and ∼1100 malaria deaths in 2009. Some experts argue that this is a serious underestimation and that the actual number of malaria cases per year is likely between 9 and 50 times greater, with an approximate 13-fold underestimation of malaria-related mortality. The difficulty in making these estimations is further exacerbated by (i) highly variable malaria eco-epidemiological profiles, (ii) the transmission and overlap of multiple Plasmodium species and Anopheles vectors, (iii) increasing antimalarial drug resistance and insecticide resistance, and (iv) the impact of climate change on each of these variables. Simply stated, the burden of malaria in India is complex. Here we describe plans for a Center for the Study of Complex Malaria in India (CSCMi), one of ten International Centers of Excellence in Malaria Research (ICEMRs) located in malarious regions of the world recently funded by the National Institute of Allergy and Infectious Diseases, National Institutes of Health. The CSCMi is a close partnership between Indian and United States scientists, and aims to address major gaps in our understanding of the complexity of malaria in India, including changing patterns of epidemiology, vector biology and control, drug resistance, and parasite genomics. We hope that such a multidisciplinary approach that integrates clinical and field studies with laboratory, molecular, and genomic methods will provide a powerful combination for malaria control and prevention in India.

BACKGROUND: The reservoir of Plasmodium infection in humans has traditionally been defined by blood slide positivity. This study was designed to characterize the local reservoir of infection in relation to the diverse var genes that encode the major surface antigen of Plasmodium falciparum blood stages and underlie the parasite\'s ability to establish chronic infection and transmit from human to mosquito.
RESULTS: We investigated the molecular epidemiology of the var multigene family at local sites in Gabon, Senegal and Kenya which differ in parasite prevalence and transmission intensity. 1839 distinct var gene types were defined by sequencing DBLα domains in the three sites. Only 76 (4.1%) var types were found in more than one population indicating spatial heterogeneity in var types across the African continent. The majority of var types appeared only once in the population sample. Non-parametric statistical estimators predict in each population at minimum five to seven thousand distinct var types. Similar diversity of var types was seen in sites with different parasite prevalences.
CONCLUSIONS: Var population genomics provides new insights into the epidemiology of P. falciparum in Africa where malaria has never been conquered. In particular, we have described the extensive reservoir of infection in local African sites and discovered a unique var population structure that can facilitate superinfection through minimal overlap in var repertoires among parasite genomes. Our findings show that var typing as a molecular surveillance system defines the extent of genetic complexity in the reservoir of infection to complement measures of malaria prevalence. The observed small scale spatial diversity of var genes suggests that var genetics could greatly inform current malaria mapping approaches and predict complex malaria population dynamics due to the import of var types to areas where no widespread pre-existing immunity in the population exists.

BACKGROUND: The sub-membranous skeleton of the ciliate Paramecium, the epiplasm, is composed of hundreds of epiplasmic scales centered on basal bodies, and presents a complex set of proteins, epiplasmins, which belong to a multigenic family. The repeated duplications observed in the P. tetraurelia genome present an interesting model of the organization and evolution of a multigenic family within a single cell.
RESULTS: To study this multigenic family, we used phylogenetic, structural, and analytical transcriptional approaches. The phylogenetic method defines 5 groups of epiplasmins in the multigenic family. A refined analysis by Hydrophobic Cluster Analysis (HCA) identifies structural characteristics of 51 epiplasmins, defining five separate groups, and three classes. Depending on the sequential arrangement of their structural domains, the epiplasmins are defined as symmetric, asymmetric or atypical. The EST data aid in this classification, in the identification of putative regulating sequences such as TATA or CAAT boxes. When specific RNAi experiments were conducted using sequences from either symmetric or asymmetric classes, phenotypes were drastic. Local effects show either disrupted or ill-shaped epiplasmic scales. In either case, this results in aborted cell division. Using structural features, we show that 4 epiplasmins are also present in another ciliate, Tetrahymena thermophila. Their affiliation with the distinctive structural groups of Paramecium epiplasmins demonstrates an interspecific multigenic family.
CONCLUSIONS: The epiplasmin multigenic family illustrates the history of genomic duplication in Paramecium. This study provides a framework which can guide functional analysis of epiplasmins, the major components of the membrane skeleton in ciliates. We show that this set of proteins handles an important developmental information in Paramecium since maintenance of epiplasm organization is crucial for cell morphogenesis.

In medical sciences, a target is a broad concept to qualify a biological entity and/or a biological phenomenon, on which one aims to act as part of a therapy. It follows that a target can be defined as a phenotype, a biological process, a subcellular organelle, a protein or a protein domain. It also follows that a target cannot be defined independently of the type of intervention one considers implementing. In this brief review, we describe how in silico organization of genomic and post-genomic information of all partners involved in malaria (human patient, Plasmodium parasite and Anopheles vector), complying with knowledge of the disease in etiologic terms, appears as an efficient source of information not only to help selecting but also discarding target candidates. Some limitations in our capacity to explore the stored biological information, due to the current quality of genomic annotation, level of database integration, or to the performances of existing analytic and mining tools, are discussed. In silico strategies to assess the feasibility of bringing a target to a therapeutic development pipeline, in terms of target \"druggability\", are introduced.