DOCSLIB.ORG

Mapping the Single Origin of Replication in the Naegleria Gruberi

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Mapping the Single Origin of Replication in the Naegleria Gruberi Protist, Vol. 170, 141–152, April 2019 http://www.elsevier.de/protis Published online date 13 February 2019 ORIGINAL PAPER Mapping the Single Origin of Replication in the Naegleria gruberi Extrachromosomal DNA Element 1,2 John C. Mullican , Nora M. Chapman, and Steven Tracy Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska 68198, USA Submitted October 25, 2018; Accepted February 6, 2019 Monitoring Editor: C. Graham Clark The genes encoding the ribosomal RNA (rRNA) subunits of the amoeba Naegleria gruberi are encoded in a relatively uncommon arrangement: on a circular extrachromosomal DNA element with each organism carrying about 4,000 copies of the element. As complete sequence analysis of the N. gruberi chromo- somal DNA revealed no copy of the rRNA genes, these extrachromosomal elements must therefore replicate autonomously. We reported elsewhere the molecular cloning and the complete sequence analysis of the entire rRNA gene-containing element of N. gruberi (strain EGB). Using neutral/neutral two-dimensional agarose electrophoresis, the region in the element enclosing the single replication origin using DNA from asynchronous and axenically propagated N. gruberi populations was localized within a 2.1 kbp fragment located approximately 2,300 bp from the 18S rRNA gene and 3,700 bp from the 28S rRNA gene. The results indicate that replication occurs from a single origin via a theta-type mode of replication rather than by a rolling circle mode. Further, G-quadruplex elements, often located near DNA replication origins, occur in and near this fragment in a repeated sequence. © 2019 Elsevier GmbH. All rights reserved. Key words: Amoeba; Naegleria gruberi; ribosomal RNA; extrachromosomal DNA element; origin of replication; G-quadruplex. 1 Corresponding author. 2 Current address: Department of Biology, Washburn University, Topeka, Kansas 66621, USA. e-mail [email protected] (J.C. Mullican). Abbreviations: TBE, Tris-borate-EDTA; ARS, Autonomous replication sequence; ORC, Origin Replication Complex; ori, origin; mtDNA, mitochondrial DNA; NRS, Non-Ribosomal Sequence; G4, G-quadruplex; pG4, Potential G-quadruplex; rDNA, ribosomal DNA; RDE, Ribosomal DNA-containing Element; rRNA, ribosomal RNA. https://doi.org/10.1016/j.protis.2019.02.001 1434-4610/© 2019 Elsevier GmbH. All rights reserved. 142 J.C. Mullican et al. Introduction chromosomal, RDE and mtDNA) of the N. gruberi organism. This is considerably higher than the ear- Free-living amoebae, such as the 47 characterized lier estimation of 17% (Clark and Cross 1987), a species of the Naegleria genus (De Jonckheere discrepancy that can be attributed to more precise 2014), are ubiquitous in soil and fresh water world- measurements of both the genome and element wide (Denet et al. 2017; Rodriguez-Zaragoza 1994; sizes (Fritz-Laylin et al. 2010). As the mass of Tyml et al. 2016). Naegleria fowleri is the sole the RDE population’s DNA represents a significant known Naegleria species that is pathogenic for fraction of the total DNA in a Naegleria organism, it humans and is infamous due to its ability, fol- will be important to understand the mechanism by lowing ingestion into the nasal cavity, to induce which this extrachromosomal element replicates. the nearly universally fatal disease, primary amoe- With thousands of copies of the RDE per cell bic meningoencephalitis (Grace et al. 2015). The (Clark and Cross 1987) and no ‘master’ copy of non-pathogenic Naegleria gruberi, on the other rDNA or the RDE itself in the chromosomal DNA hand, represents a useful research model organism (Clark and Cross 1987; Fritz-Laylin et al. 2010), (Ahmed Khan et al. 2015; Opperdoes et al. 2011). Clark and Cross (1987) inferred that the RDE must From bacteria to humans, all cells contain replicate autonomously. There have been no pub- repetitive rRNA genes (rDNA) (Long and Dawid lished studies to date designed to map the origin(s) 1980). Examining the NEG-M strain of N. gruberi, of replication (ori) of the RDE in Naegleria. Stud- Clark and Cross (1987) first demonstrated that ies of eukaryotic DNA replication ori have focused this Naegleria species carries its ribosomal RNA on chromosomes in the yeast, human and murine (rRNA) genes on an extrachromosomal element; genomes (Parker et al. 2017). It is noteworthy later, Maruyama and Nozaki (2007) sequenced that the primary structure requirements of eukary- the extrachromosomal element of this particu- otic DNA ori are not well understood, save for lar strain. Complete sequence analysis of the yeast; indeed, there is a better understanding of N. gruberi organism’s nuclear DNA (Fritz-Laylin the proteins involved in the ori replication complex et al. 2010) confirmed earlier work based on (ORC) than of the requirements of the DNA site(s) Southern blot hybridization analysis (Clark and that is/are recognized [reviewed in (Leonard and Cross 1987) that there is no copy of the ribo- Mechali 2013; O’Donnell et al. 2013)]. somal DNA in the chromosomal DNA. While Nonetheless, the search for specific primary DNA this arrangement is uncommon, the placement structure correlates of DNA ori has identified certain of rDNA only on an extrachromosomal element genetic elements often associated with replication has also been demonstrated for other members initiation. The discovery that GC enrichment of of the order Schizopyrenida (Clark and Cross regions where origins are found in metazoan DNA 1988). Entamoeba histolytica and E. invadens also has led to the recognition of a role for a structural lodge the rDNA on an extrachromosomal element component called the G-quadruplex (G4) (Prioleau (Bhattacharya et al. 1989, 1998) as do some other 2017). The G4s derive from primary sequences eukaryotic microbial species, several of which pos- containing four separated repeats of two or more sess circular rDNA (Torres-Machorro et al. 2010). G nucleotides. They can form structures, based Earlier work estimated the number of the ribo- on the non-canonical interactions [Hoogsteen base somal DNA-containing extrachromosomal element pairing; (Hoogsteen 1963)] of the G nucleotides, to (RDE) molecules at about 4,000 per cell based on form 2 or more planar structures. These are termed visual analysis of slot-blot data (Clark and Cross G tetrads [(Sen and Gilbert 1988); reviewed and 1987). Complete sequence analysis of the entire depicted in (Maizels 2006)]. Such tetrads can stack genome of N. gruberi (strain NEG-M) (Fritz-Laylin upon each other to form the G-quadruplex (Sup- et al. 2010) revealed the mass of the nuclear DNA plementary Material Fig. S1). Recognition of these 6 × per organism to be 0.045 pg DNA (41 10 bp sequence motifs associated with DNA ori (Cayrou per genome, calculated at 660 Da/bp). Each organ- et al. 2012) has prompted mutational analyses, ism (Clark 1990) also contains approximately 420 which demonstrate that while these components copies of mitochondrial DNA (mtDNA) which, at are not sufficient for an origin of DNA replication, 50 kbp per copy (Fritz-Laylin et al. 2010), yields they are necessary at least for some defined sites about 0.023 pg mtDNA per cell. Thus, at about (Valton et al. 2014). 4 × 4,000 copies of the 1.4 10 bp RDE per cell (Clark Autonomous replication sequences (ARS) with and Cross 1987; Maruyama and Nozaki 2007) or conserved origin sequence motifs have been iden- about 0.061 pg per cell, the RDE population repre- tified in yeast species. And, just as for the G4s, sents about 47% of the total DNA mass (nuclear these appear to be necessary for ori function (Peng N. gruberi Extrachromosomal rDNA Origin 143 et al. 2015). Interestingly, two sequences with clone revealed the location of the 5.8S, 18S homology to the conserved ori sequence motifs and 28S rRNA genes as well as numerous of Saccharomyces cerevisiae were noted in the repeat sequences [(Mullican et al. 2018); anno- N. gruberi ribosomal DNA extrachromosomal ele- tated in GenBank Accession MG699123]. We found ment (Mullican et al. 2018). The nonribosomal (Mullican et al. 2018), as had Maruyama and region of the RDE is largely composed of directly Nozaki (2007) earlier, that the 18S rDNA of the repeated DNA sequences ranging in size up to two different strains of N. gruberi lacked the type 315 nucleotides and include both interspersed and I intron reported to be in most Naegleria species’ tandemly repeated sequences. Due to the high RDEs (Wikmark et al. 2006). The two sequences copy number of these RDEs in Naegleria cells as derived from two closely related N. gruberi strains well as the considerable evolutionary divergence of are very similar with most differences contained these Excavata protists from metazoans and yeast, within a region of 120 bp tandem repeats in the and the repeated observations that DNA primary non-rDNA coding region. The RDEs of the 2 dif- structure in ori is quite variable, it is possible that ferent amoebic strains differ at only 4 sites (with the primary (as well as higher order) structure of nucleotide numbering referring to the pNgrubEGB the ori in the Naegleria RDE may differ from other sequence positions; GenBank MG699123): a sin- eukaryotic origins studied (Leonard and Mechali gle nucleotide (nt) deletion (nt8581-8582), a single 2013; Parker et al. 2017). nucleotide variation (nt8703) and a 119 bp deletion As the mechanism of the Naegleria RDE replica- (relative to the sequence for strain NEG-M; Gen- tion and regulation remains unexplored, we initiated Bank AB298288) of part of the 4th and 5th copy of studies toward defining the RDE ori. We hypothe- the 120 bp tandem repeat (nt8748-8749) of the non- sized that each RDE possesses at least one ori and ribosomal sequence (NRS). There is also a single we sought to map the location of such a domain nt deletion in the EGB strain’s RDE at nt13736- within a Naegleria RDE.
Load more

Recommended publications
  • UNIVERSITE DES ANTILLES Faculté Des Sciences Exactes Et Naturelles École Doctorale Pluridisciplinaire
    UNIVERSITE DES ANTILLES Faculté des Sciences Exactes et Naturelles École Doctorale pluridisciplinaire Thèse de Doctorat En Physiologie-biologie des organismes-populations-interactions Mirna MOUSSA Les amibes libres pathogènes des eaux chaudes de la Guadeloupe, étude écologique, caractérisation moléculaire et prophylaxie des zones de baignade Sous la direction de : Jérôme GUERLOTTÉ, Professeur, Université des Antilles Antoine TALARMIN, Docteur, Institut Pasteur de la Guadeloupe Soutenue le 25 Septembre 2015 à l’Institut Pasteur de Guadeloupe N: Jury: GROS Olivier, Pr. Université des Antilles Président HÉCHARD Yann, Pr. Université de Poitiers Rapporteur YERA Hélène , MCU-PH Institut Cochin, Paris Rapporteur ALBINA Emmanuel, Dr. CIRAD Antilles Examinateur DESBOIS Nicole, Dr. CHU de Fort-de-France Examinateur GUERLOTTÉ Jérôme, Pr. Université des Antilles Examinateur 2 A ma famille et mes amis pour leur Amour et leur soutien au quotidien. Vous avez toujours su trouver les mots pour m’encourager dans les moments les plus difficiles. Je ne vous remercierai jamais assez pour tout ce que vous avez fait. Je pense à ma grand-mère en Syrie qui m’a appris la joie de vivre malgré les difficultés. 3 4 Remerciements Je souhaite tout d’abord exprimer ma profonde gratitude au Docteur Antoine Talarmin qui m’a permis de réaliser cette thèse au sein de l’Institut Pasteur de la Guadeloupe ainsi qu’au Professeur Jérôme Guerlotté qui m’a fait l’honneur d’être mon Directeur de thèse durant ces trois années. Je leur suis extrêmement reconnaissante de m’avoir fait confiance et de m’avoir donné des opportunités pour réussir dans ce domaine. Je les remercie pour leur patience, leurs conseils, leur soutien et leur disponibilité.
    [Show full text]
  • Mitochondrial DNA
    GBE Missing Genes, Multiple ORFs, and C-to-U Type RNA Editing in Acrasis kona (Heterolobosea, Excavata) Mitochondrial DNA Cheng-Jie Fu1,*, Sanea Sheikh1, Wei Miao2, Siv G.E. Andersson3, and Sandra L. Baldauf1,* 1Program in Systematic Biology, Department of Organismal Biology, Evolutionary Biology Centre, Uppsala University, Sweden 2Key Laboratory of Aquatic Biodiversity and Conservation, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, China 3Department of Molecular Evolution, Cell and Molecular Biology, Science for Life Laboratory, Biomedical Centre, Uppsala University, Sweden *Corresponding author: E-mail: [email protected], [email protected]; [email protected]. Accepted: August 18, 2014 Data deposition: The Acrasis kona mitochondrial genome sequence and annotation have been deposited at GenBank under the accession KJ679272. Downloaded from Abstract Discoba(Excavata)isanancientgroupof eukaryotes withgreatmorphological andecologicaldiversity.Unlikethe other major divisions http://gbe.oxfordjournals.org/ of Discoba (Jakobida and Euglenozoa), little is known about the mitochondrial DNAs (mtDNAs) of Heterolobosea. We have assembled a complete mtDNA genome from the aggregating heterolobosean amoeba, Acrasis kona, which consists of a single circular highly AT- rich (83.3%) molecule of 51.5 kb. Unexpectedly, A. kona mtDNA is missing roughly 40% of the protein-coding genes and nearly half of the transfer RNAs found in the only other sequenced heterolobosean mtDNAs, those of Naegleria spp. Instead, over a quarter of A. kona mtDNA consists of novel open reading frames. Eleven of the 16 protein-coding genes missing from A. kona mtDNA were identified in its nuclear DNA and polyA RNA, and phylogenetic analyses indicate that at least 10 of these 11 putative nuclear-encoded mitochondrial (NcMt) proteins arose by direct transfer from the mitochondrion.
    [Show full text]
  • The Genome of Naegleria Lovaniensis, the Basis for a Comparative Approach to Unravel Pathogenicity Factors of the Human Pathogenic Amoeba N
    Liechti et al. BMC Genomics (2018) 19:654 https://doi.org/10.1186/s12864-018-4994-1 RESEARCHARTICLE Open Access The genome of Naegleria lovaniensis, the basis for a comparative approach to unravel pathogenicity factors of the human pathogenic amoeba N. fowleri Nicole Liechti1,2,3, Nadia Schürch2, Rémy Bruggmann1 and Matthias Wittwer2* Abstract Background: Members of the genus Naegleria are free-living eukaryotes with the capability to transform from the amoeboid form into resting cysts or moving flagellates in response to environmental conditions. More than 40 species have been characterized, but only Naegleria fowleri (N. fowleri) is known as a human pathogen causing primary amoebic meningoencephalitis (PAM), a fast progressing and mostly fatal disease of the central nervous system. Several studies report an involvement of phospholipases and other molecular factors, but the mechanisms involved in pathogenesis are still poorly understood. To gain a better understanding of the relationships within the genus of Naegleria and to investigate pathogenicity factors of N. fowleri, we characterized the genome of its closest non-pathogenic relative N. lovaniensis. Results: To gain insights into the taxonomy of Naegleria, we sequenced the genome of N. lovaniensis using long read sequencing technology. The assembly of the data resulted in a 30 Mb genome including the circular mitochondrial sequence. Unravelling the phylogenetic relationship using OrthoMCL protein clustering and maximum likelihood methods confirms the close relationship of N. lovaniensis and N. fowleri. To achieve an overview of the diversity of Naegleria proteins and to assess characteristics of the human pathogen N. fowleri, OrthoMCL protein clustering including data of N.
    [Show full text]
  • Lais Helena Teixeira.Pdf
    1 Universidade Católica de Santos Mestrado em Saúde Coletiva OCORRÊNCIA DE AMEBAS DE VIDA-LIVRE, DOS GÊNEROS Acanthamoeba E Naegleria, EM PISOS DE AMBIENTES INTERNOS, NA UNIVERSIDADE CATÓLICA DE SANTOS, SP, BRASIL. LAIS HELENA TEIXEIRA Santos 2008 2 Universidade Católica de Santos Mestrado em Saúde Coletiva OCORRÊNCIA DE AMEBAS DE VIDA-LIVRE, DOS GÊNEROS Acanthamoeba E Naegleria, EM PISOS DE AMBIENTES INTERNOS, NA UNIVERSIDADE CATÓLICA DE SANTOS, SP, BRASIL. LAIS HELENA TEIXEIRA Dissertação apresentada ao Programa de Mestrado em Saúde Coletiva da Universidade Católica de Santos para obtenção do grau de Mestre em Saúde Coletiva. Área de concentração: Meio Ambiente e Saúde Orientador: Profº. Dr. Sérgio Olavo Pinto da Costa Co ! Orientador: Profº Dr. Marcos Montani Caseiro Santos 2008 3 Dados Internacionais de Catalogação Sistema de Bibliotecas da Universidade Católica de Santos SIBIU _______________________________________________________________ T266o Teixeira, Lais Helena Ocorrência de amebas de vida-livre, dos gêneros Acanthamoeba e Naegleria, em pisos de ambientes internos, na Universidade Católica de Santos, SP, Brasil. / Lais Helena Teixeira ! Santos: [s.n.], 2008. 78 f.; 30 cm. (Dissertação de Mestrado - Universidade Católica de Santos, Programa em Saúde Coletiva) I. Teixeira, Lais Helena. II. Título. CDU 614(043.3) _______________________________________________________________ 4 Agradecimentos A minha mãe, W anda Silva Rodrigues, que teve grande importância em toda a minha formação, por todo apoio, pela paciência, por todas as noites mal dormidas, e por ter sido a maior incentivadora para eu cursar este programa de Mestrado. Ao Prof. Dr. Sergio Olavo Pinto da Costa, pela sua enorme confiança, desde o início do projeto, pela sua compreensão e paciência, e pelo incentivo para a realização e conclusão desta dissertação.
    [Show full text]
  • Free-Living Protozoa in Drinking Water Supplies: Community Composition and Role As Hosts for Legionella Pneumophila
    Free-living protozoa in drinking water supplies: community composition and role as hosts for Legionella pneumophila Rinske Marleen Valster Thesis committee Thesis supervisor Prof. dr. ir. D. van der Kooij Professor of Environmental Microbiology Wageningen University Principal Microbiologist KWR Watercycle Institute, Nieuwegein Thesis co-supervisor Prof. dr. H. Smidt Personal chair at the Laboratory of Microbiology Wageningen University Other members Dr. J. F. Loret, CIRSEE-Suez Environnement, Le Pecq, France Prof. dr. T. A. Stenstrom,¨ SIIDC, Stockholm, Sweden Dr. W. Hoogenboezem, The Water Laboratory, Haarlem Prof. dr. ir. M. H. Zwietering, Wageningen University This research was conducted under the auspices of the Graduate School VLAG. Free-living protozoa in drinking water supplies: community composition and role as hosts for Legionella pneumophila Rinske Marleen Valster Thesis submitted in fulfilment of the requirements for the degree of doctor at Wageningen University by the authority of the Rector Magnificus Prof. dr. M.J. Kropff, in the presence of the Thesis Committee appointed by the Academic Board to be defended in public on Monday 20 June 2011 at 11 a.m. in the Aula Rinske Marleen Valster Free-living protozoa in drinking water supplies: community composition and role as hosts for Legionella pneumophila, viii+186 pages. Thesis, Wageningen University, Wageningen, NL (2011) With references, with summaries in Dutch and English ISBN 978-90-8585-884-3 Abstract Free-living protozoa, which feed on bacteria, play an important role in the communities of microor- ganisms and invertebrates in drinking water supplies and in (warm) tap water installations. Several bacteria, including opportunistic human pathogens such as Legionella pneumophila, are able to sur- vive and replicate within protozoan hosts, and certain free-living protozoa are opportunistic human pathogens as well.
    [Show full text]
  • Actions of Cannabinoids on Amoebae
    Actions of Cannabinoids on Amoebae By Israa Al-hammadi This thesis is submitted for the degree of Doctor of Philosophy April 2020 Department of Biomedical and Life Sciences 2nd April 2020 To Whom It May Concern: Declaration Actions of cannabinoids on amoebae. This thesis has not been submitted in support of an application for another degree at this or any other university. It is the result of my own work and includes nothing that is the outcome of work done in collaboration except where specifically indicated. Many of the ideas in this thesis were the product of discussion with my supervisor Dr. Jackie Parry and Dr. Karen Wright. Israa Al-hammadi MSc. BSc. Table of contents Abstract 1 Chapter 1: General Introduction 3 1.1 Introduction 3 1.2 The Endocannabinoid System (ECS) in humans 3 1.2.1 General overview 3 1.2.2 Main ligands of the endocannabinoid system 3 1.2.2.1 Endocannabinoids 4 1.2.2.2 Phytocannabinoids 7 1.2.3 Main cannabinoid receptors 10 1.2.3.1. CB1 and CB2 receptors 10 1.2.3.2 The transient receptor potential vanilloid type 1 (TRPV1) 11 1.2.3.3 G-protein coupled receptor 55 (GPR55) 11 1.2.4 Other receptors 12 1.2.4.1 Peroxisome Proliferator-activated Receptors (PPARs) 12 1.2.4.2 Dopamine Receptors 14 1.2.4.3 Serotonin receptors (5-HT) 16 1.3. Existence of an endocannabinoid system in single-celled eukaryotes 17 1.3.1. Endocannabinoids in Tetrahymena 18 1.3.2. Enzymes in Tetrahymena 19 1.3.3.
    [Show full text]
  • Diversity, Phylogeny and Phylogeography of Free-Living Amoebae
    School of Doctoral Studies in Biological Sciences University of South Bohemia in České Budějovice Faculty of Science Diversity, phylogeny and phylogeography of free-living amoebae Ph.D. Thesis RNDr. Tomáš Tyml Supervisor: Mgr. Martin Kostka, Ph.D. Department of Parasitology, Faculty of Science, University of South Bohemia in České Budějovice Specialist adviser: Prof. MVDr. Iva Dyková, Dr.Sc. Department of Botany and Zoology, Faculty of Science, Masaryk University České Budějovice 2016 This thesis should be cited as: Tyml, T. 2016. Diversity, phylogeny and phylogeography of free living amoebae. Ph.D. Thesis Series, No. 13. University of South Bohemia, Faculty of Science, School of Doctoral Studies in Biological Sciences, České Budějovice, Czech Republic, 135 pp. Annotation This thesis consists of seven published papers on free-living amoebae (FLA), members of Amoebozoa, Excavata: Heterolobosea, and Cercozoa, and covers three main topics: (i) FLA as potential fish pathogens, (ii) diversity and phylogeography of FLA, and (iii) FLA as hosts of prokaryotic organisms. Diverse methodological approaches were used including culture-dependent techniques for isolation and identification of free-living amoebae, molecular phylogenetics, fluorescent in situ hybridization, and transmission electron microscopy. Declaration [in Czech] Prohlašuji, že svoji disertační práci jsem vypracoval samostatně pouze s použitím pramenů a literatury uvedených v seznamu citované literatury. Prohlašuji, že v souladu s § 47b zákona č. 111/1998 Sb. v platném znění souhlasím se zveřejněním své disertační práce, a to v úpravě vzniklé vypuštěním vyznačených částí archivovaných Přírodovědeckou fakultou elektronickou cestou ve veřejně přístupné části databáze STAG provozované Jihočeskou univerzitou v Českých Budějovicích na jejích internetových stránkách, a to se zachováním mého autorského práva k odevzdanému textu této kvalifikační práce.
    [Show full text]
  • Downloaded from the Virulence Factor of Pathogenic Bacteria Database (VFDB)70
    www.nature.com/scientificreports OPEN Investigation of potential pathogenicity of Willaertia magna by investigating the transfer of bacteria pathogenicity genes into its genome Issam Hasni1,2, Nisrine Chelkha1, Emeline Baptiste1, Mouh Rayane Mameri2, Joel Lachuer3,4,5, Fabrice Plasson2, Philippe Colson 1 & Bernard La Scola1* Willaertia magna c2c maky is a thermophilic amoeba closely related to the genus Naegleria. This free-living amoeba has the ability to eliminate Legionella pneumophila, which is an amoeba-resisting bacterium living in an aquatic environment. To prevent the proliferation of L. pneumophila in cooling towers, the use of W. magna as natural biocide has been proposed. To provide a better understanding of the W. magna genome, whole-genome sequencing was performed through the study of virulence factors and lateral gene transfers. This amoeba harbors a genome of 36.5 megabases with 18,519 predicted genes. BLASTp analyses reported protein homology between 136 W. magna sequences and amoeba-resistant microorganisms. Horizontal gene transfers were observed based on the basis of the phylogenetic reconstruction hypothesis. We detected 15 homologs of N. fowleri genes related to virulence, although these latter were also found in the genome of N. gruberi, which is a non-pathogenic amoeba. Furthermore, the cytotoxicity test performed on human cells supports the hypothesis that the strain c2c maky is a non-pathogenic amoeba. This work explores the genomic repertory for the frst draft genome of genus Willaertia and provides genomic data for further comparative studies on virulence of related pathogenic amoeba, N. fowleri. Heterolobosea is a rich class of species of heterotrophs, almost all free-living, containing pathogenic (e.g.
    [Show full text]
  • Phylogeny of Deep-Level Relationships Within Euglenozoa Based on Combined Small Subunit and Large Subunit Ribosomal DNA Sequence
    PHYLOGENY OF DEEP-LEVEL RELATIONSHIPS WITHIN EUGLENOZOA BASED ON COMBINED SMALL SUBUNIT AND LARGE SUBUNIT RIBOSOMAL DNA SEQUENCES by BING MA (Under the Direction of Mark A. Farmer) ABSTRACT My master’s research addressed questions about the evolutionary histories of Euglenozoa, with special attention given to deep-level relationships among taxa. The Euglenozoa are a putative early-branching assemblage of flagellated Eukaryotes, comprised primarily by three subgroups: euglenids, kinetoplastids and diplonemids. The goals of my research were to 1) evaluate the phylogenetic potential of large subunit ribosomal DNA (LSU rDNA) gene sequences as a molecular marker; 2) construct a phylogeny for the Euglenozoa to address their deep-level relationships; 3) provide morphological data of the flagellate Petalomonas cantuscygni to infer its evolutionary position in Euglenozoa. A dataset based on LSU rDNA sequences combined with SSU rDNA from thirty-nine taxa representing every subgroup of Euglenozoa and outgroup species was used for testing relationships within the Euglenozoa. Our results indicate that a) LSU rDNA is a useful marker to infer phylogeny, b) euglenids and diplonemdis are more closely related to one another than either is to the kinetoplatids and c) Petalomonas cantuscygni is closely related to the diplonemids. INDEX WORDS: Euglenozoa, phylogenetic inference, molecular evolution, 28S rDNA, LSU rDNA, 18S rDNA, SSU rDNA, euglenids, diplonemids, kinetoplastids, conserved core regions, expansion segments, TOL PHYLOGENY OF DEEP-LEVEL RELATIONSHIPS WITHIN EUGLENOZOA BASED ON COMBINED SMALL SUBUNIT AND LARGE SUBUNIT RIBOSOMAL DNA SEQUENCES by BING MA B. Med., Zhengzhou University, P. R. China, 2002 A Thesis Submitted to the Graduate Faculty of The University of Georgia in Partial Fulfillment of the Requirements for the Degree MASTER OF SCIENCE ATHENS, GEORGIA 2005 © 2005 Bing Ma All Rights Reserved PHYLOGENY OF DEEP-LEVEL RELATIONSHIPS WITHIN EUGLENOZOA BASED ON COMBINED SMALL SUBUNIT AND LARGE SUBUNIT RIBOSOMAL DNA SEQUENCES by BING MA Major Professor: Mark A.
    [Show full text]
  • The Hydrogenosomes of Psalteriomonas Lanterna
    BMC Evolutionary Biology BioMed Central Research article Open Access The hydrogenosomes of Psalteriomonas lanterna Rob M de Graaf†1, Isabel Duarte†2, Theo A van Alen1, Jan WP Kuiper1,4, Klaas Schotanus1,5, Jörg Rosenberg3, Martijn A Huynen2,6 and Johannes HP Hackstein*1 Address: 1Department of Evolutionary Microbiology, IWWR, Radboud University Nijmegen, Heyendaalseweg 135, 6525AJ Nijmegen, The Netherlands, 2Center for Molecular and Biomolecular Informatics, Nijmegen Center for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Geert Grooteplein 28, 6525GA Nijmegen, The Netherlands, 3Sommerhaus 45, D-50129 Bergheim, Germany, 4CIHR Group in Matrix Dynamics, University of Toronto, 150 College Street, Toronto, Ontario, Canada M5S 3E2, 5Department of Ecology and Evolutionary Biology, 106A Guyot Hall, Princeton University, Princeton NJ 08554-2016, USA and 6Netherlands Bioinformatic Centre, Geert Grooteplein 28, 6525 GA Nijmegen, The Netherlands Email: Rob M de Graaf - [email protected]; Isabel Duarte - [email protected]; Theo A van Alen - [email protected]; Jan WP Kuiper - [email protected]; Klaas Schotanus - [email protected]; Jörg Rosenberg - [email protected]; Martijn A Huynen - [email protected]; Johannes HP Hackstein* - [email protected] * Corresponding author †Equal contributors Published: 9 December 2009 Received: 20 July 2009 Accepted: 9 December 2009 BMC Evolutionary Biology 2009, 9:287 doi:10.1186/1471-2148-9-287 This article is available from: http://www.biomedcentral.com/1471-2148/9/287 © 2009 de Graaf et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
    [Show full text]
  • Multi-Gene Phylogenetic Analysis of the Supergroup Excavata
    MULTI-GENE PHYLOGENETIC ANALYSIS OF THE SUPERGROUP EXCAVATA By CHRISTINA CASTLEJOHN (Under the Direction of Mark A. Farmer) ABSTRACT The supergroup Excavata, one of six supergroups of eukaryotes, has been a controversial supergroup within the Eukaryotic Tree of Life. Excavata was originally based largely on morphological data and to date has not been well supported by molecular studies. The goals of this research were to test the monophyly of Excavata and to observe relationships among the nine subgroups of excavates included in this study. Several different types of phylogenetic analyses were performed on a data set consisting of sequences from nine reasonably conserved genes. Analyses of this data set recovered monophyly of Excavata with moderate to strong support. Topology tests rejected all but two topologies: one with a monophyletic Excavata and one with Excavata split into two major clades. Simple gap coding, which was performed on the ribosomal DNA alignments, was found to be more useful for species-level analyses than deeper relationships with the eukaryotes. INDEX WORDS: Excavata, excavates, monophyly, phylogenetic analysis, gap coding MULTI-GENE PHYLOGENETIC ANALYSIS OF THE SUPERGROUP EXCAVATA By CHRISTINA CASTLEJOHN B.S., Georgia Institute of Technology, 2002 A Thesis Submitted to the Graduate Faculty of The University of Georgia in Partial Fulfillment of the Requirements for the Degree MASTER OF SCIENCE ATHENS, GEORGIA 2009 © 2009 Christina Castlejohn All Rights Reserved MULTI-GENE PHYLOGENETIC ANALYSIS OF THE SUPERGROUP EXCAVATA By CHRISTINA CASTLEJOHN Major Professor: Mark A. Farmer Committee: James Leebens-Mack Joseph McHugh Electronic Version Approved: Maureen Grasso Dean of the Graduate School The University of Georgia August 2009 iv DEDICATION To my family, who have supported me in my journey v ACKNOWLEDGEMENTS I would like to thank Mark Farmer for helping me so much in my pursuit of higher education and my plans for the future.
    [Show full text]
  • Identity of Naegleria Strains Isolated from Organs of Freshwater Fishes
    DISEASES OF AQUATIC ORGANISMS Vol. 46: 115–121, 2001 Published September 12 Dis Aquat Org Identity of Naegleria strains isolated from organs of freshwater fishes I. Dyková1, 2,*, I. Kyselová1, 2, H. Pecková1, M. Oborník1, 2, J. Luke$1, 2 1Institute of Parasitology, Academy of Sciences of the Czech Republic, and 2Faculty of Biological Sciences, University of South Bohemia, Brani$ovská 31, 370 05 >eské Budˇejovice, Czech Republic ABSTRACT: Eighteen Naegleria strains were isolated from organs of freshwater fishes belonging to 5 species. Morphometric study allowed the separation of the Naegleria strains from the non- vahlkampfiid amoeboflagellates, but was inadequate for species determination. Six strains, repre- sentatives of groups that had a slightly different cyst size, were selected and corresponding derived clones were subjected to sequence analysis and riboprinting restriction fragment length polymor- phism (RFLP)-PCR analysis of the small subunit (SSU) rRNA genes. One strain isolated from the brain of a fish with systemic infection was characterised by an intronless 2 kb long SSU rRNA gene and was identified as N. australiensis. Another 5 strains had a 1.3 kb long group I intron in their SSU rRNA gene and, based on the SSU rRNA sequences and riboprints, RFLP-PCR patterns appeared in phylo- genetic trees to be closely related to Naegleria clarki. KEY WORDS: Naegleria spp. · Fish strains · Molecular taxonomy · SSU rRNA Resale or republication not permitted without written consent of the publisher INTRODUCTION hosts. Naegleria gruberi was isolated along with other species of free-living amoebae from the intestinal con- The genus and species composition of amoebae that tents of white suckers Catostomus commersoni and invade fish as well as their biology and host-parasite common shiners Notropis cornutus (Franke & Mack- relations are of fundamental interest to fish patho- iewicz 1982).
    [Show full text]