Logsdon John M
Full Text Available Abstract Background Comparative genomic studies of the mitochondrion-lacking protist group Diplomonadida (diplomonads has been lacking, although Giardia lamblia has been intensively studied. We have performed a sequence survey project resulting in 2341 expressed sequence tags (EST corresponding to 853 unique clones, 5275 genome survey sequences (GSS, and eleven finished contigs from the diplomonad fish parasite Spironucleus salmonicida (previously described as S. barkhanus. Results The analyses revealed a compact genome with few, if any, introns and very short 3' untranslated regions. Strikingly different patterns of codon usage were observed in genes corresponding to frequently sampled ESTs versus genes poorly sampled, indicating that translational selection is influencing the codon usage of highly expressed genes. Rigorous phylogenomic analyses identified 84 genes – mostly encoding metabolic proteins – that have been acquired by diplomonads or their relatively close ancestors via lateral gene transfer (LGT. Although most acquisitions were from prokaryotes, more than a dozen represent likely transfers of genes between eukaryotic lineages. Many genes that provide novel insights into the genetic basis of the biology and pathogenicity of this parasitic protist were identified including 149 that putatively encode variant-surface cysteine-rich proteins which are candidate virulence factors. A number of genomic properties that distinguish S. salmonicida from its human parasitic relative G. lamblia were identified such as nineteen putative lineage-specific gene acquisitions, distinct mutational biases and codon usage and distinct polyadenylation signals. Conclusion Our results highlight the power of comparative genomic studies to yield insights into the biology of parasitic protists and the evolution of their genomes, and suggest that genetic exchange between distantly-related protist lineages may be occurring at an appreciable rate in eukaryote
Sheppard, Barbara J; Stockdale Walden, Heather D; Kondo, Hirotaka
A commercial facility producing hamsters with a history of infection by dwarf tapeworm (Hymenolepis nana) submitted 15 animals for necropsy and postmortem parasitological and microscopic examination. No tapeworms were detected grossly or microscopically. Fecal examination including gastrointestinal mucosal smears demonstrated mixed intestinal bacteria and low numbers of Giardia sp. Histologic examination of small intestine demonstrated filling of the small intestinal crypts by large numbers of 7-9 µm × 3 µm, rod to crescent or teardrop-shaped flagellates consistent with Spironucleus sp. These organisms had two 1-µm, basophilic, oval nuclei and multiple superficial flagella-like structures. Much larger 10-15 µm × 8-10 µm, oval to pear-shaped organisms were also present in lower numbers and usually located with the crypts. These larger flagellates had multiple flagella and a basophilic rod-shaped nucleus. The larger flagellates included Giardia sp., which had an intimate interface with the surface of the mucosal epithelium, bilaterally symmetry, and binucleation. Lower numbers of trichomonads were also present and were distinguished by an undulating surface membrane and a single nucleus. The mucosa was hyperplastic and moderately inflamed. Although the tapeworm infection was resolved, diagnosis of multiple intestinal flagellates by fecal examination is complicated by the varying sensitivity and diagnostic accuracy of different types of fecal analysis for different flagellate types. Key differences in the morphology and location of the different types of flagellates as observed by histology of intestinal tissues provide important additional diagnostic information to distinguish trichomonads, Spironucleus sp., and Giardia sp.
Full Text Available Abstract Background Diplomonads are common free-living inhabitants of anoxic aquatic environments and are also found as intestinal commensals or parasites of a wide variety of animals. Spironucleus vortens is a putatively commensal diplomonad of angelfish that grows to high cell densities in axenic culture. Genomic sequencing of S. vortens is in progress, yet little information is available regarding molecular and cellular aspects of S. vortens biology beyond descriptive ultrastructural studies. To facilitate the development of S. vortens as an additional diplomonad experimental model, we have constructed and stably transformed an episomal plasmid containing an enhanced green fluorescent protein (GFP tag, an AU1 epitope tag, and a tandem affinity purification (TAP tag. This construct also contains selectable antibiotic resistance markers for both S. vortens and E. coli. Results Stable transformants of S. vortens grew relatively rapidly (within 7 days after electroporation and were maintained under puromycin selection for over 6 months. We expressed the enhanced GFP variant, eGFP, under transcriptional control of the S. vortens histone H3 promoter, and visually confirmed diffuse GFP expression in over 50% of transformants. Next, we generated a histone H3::GFP fusion using the S. vortens conventional histone H3 gene and its native promoter. This construct was also highly expressed in the majority of S. vortens transformants, in which the H3::GFP fusion localized to the chromatin in both nuclei. Finally, we used fluorescence in situ hybridization (FISH of the episomal plasmid to show that the transformed plasmid localized to only one nucleus/cell and was present at roughly 10–20 copies per nucleus. Because S. vortens grows to high densities in laboratory culture, it is a feasible diplomonad from which to purify native protein complexes. Thus, we also included a TAP tag in the plasmid constructs to permit future tagging and subsequent purification
O. V. Galzitskaya
Full Text Available How is it possible to find good traits for phylogenetic reconstructions? Here, we present a new phyloproteomic criterion that is an occurrence of simple motifs which can be imprints of evolution history. We studied the occurrences of 11780 six-residue-long motifs consisting of two randomly located amino acids in 97 eukaryotic and 25 bacterial proteomes. For all eukaryotic proteomes, with the exception of the Amoebozoa, Stramenopiles, and Diplomonadida kingdoms, the number of proteins containing the motifs from the first group (one of the two amino acids occurs once at the terminal position made about 20%; in the case of motifs from the second (one of two amino acids occurs one time within the pattern and third (the two amino acids occur randomly groups, 30% and 50%, respectively. For bacterial proteomes, this relationship was 10%, 27%, and 63%, respectively. The matrices of correlation coefficients between numbers of proteins where a motif from the set of 11780 motifs appears at least once in 9 kingdoms and 5 phyla of bacteria were calculated. Among the correlation coefficients for eukaryotic proteomes, the correlation between the animal and fungi kingdoms (0.62 is higher than between fungi and plants (0.54. Our study provides support that animals and fungi are sibling kingdoms. Comparison of the frequencies of six-residue-long motifs in different proteomes allows obtaining phylogenetic relationships based on similarities between these frequencies: the Diplomonadida kingdoms are more close to Bacteria than to Eukaryota; Stramenopiles and Amoebozoa are more close to each other than to other kingdoms of Eukaryota.
Sanitary profile in mice and rat colonies in laboratory animal houses in Minas Gerais: I - Endo and ectoparasites Perfil sanitário de colônias de camundongos e ratos de biotérios de Minas Gerais: I - Endo e ectoparasitos
Full Text Available The sanitary conditions of 13 animal houses in nine public institutions in Minas Gerais, and the presence of endo and ectoparasites of mice and rats colonies kept in these facilities were evaluated. Data about barriers to prevent the transmission of diseases and a program of sanitary monitoring were obtained through a questionnaire and local visit. Parasitological methods were performed for diagnosing mite, lice, helminthes, and protozoa parasites in 344 mice and 111 rats. Data have shown that the majority of the animal houses had neither proper physical environment nor protection barriers to prevent the transmission of infections. Parasitological results have shown that only one animal house (7.7% had parasite free animals, whereas the others have presented infected animals and the prevalences of parasites in the mice colonies were: Myobia musculi (23.1%; Myocoptes musculinus (38.5%; Radfordia affinis (15.4%; Syphacia obvelata (92.3%; Aspiculuris tetraptera (23.1%; Hymenolepis nana (15.4%; Spironucleus muris (46.2%; Giardia muris (46.2%; Tritrichomonas muris (53.8%; Trichomonas minuta (61.5%; Hexamastix muris (7.7%; and Entamoeba muris (84.6%. As for the rat colonies, the prevalences were: Poliplax spinulosa (8.1%; Syphacia muris (46.2%; Trichosomoides crassicauda (28.6%; Spironucleus muris (85.7%; Tritrichomonas muris (85.7%; Trichomonas minuta (85.7%; Hexamastix muris (14.3% and Entamoeba muris (85.7%.Avaliaram-se as condições sanitárias de 13 biotérios de nove instituições públicas do estado de Minas Gerais, bem como a presença de endo e ectoparasitos nos camundongos e ratos criados nesses biotérios. Os dados sobre barreiras contra infecções e sobre o programa de monitoramento sanitário dos animais foram obtidos por meio de um questionário e de visitas aos biotérios. Métodos parasitológicos foram utilizados para o diagnóstico de ácaros, piolhos, helmintos e protozoários em 344 camundongos e 111 ratos. A maioria dos biot
Raissa Alves Gonçalves
Full Text Available Os parasitos têm sido reconhecidos como importantes componentes dos ecossistemas. Esses usam o ambiente para a sua transmissão e podem infectar diversas espécies de peixe no ecossistema. O presente estudo investigou a ecologia parasitária em Squaliforma emarginata e Harttia duriventris da bacia Igarapé Fortaleza, região de Macapá, estado do Amapá, Norte do Brasil. Para ambos os peixes houve similar nível de infecção por Ichthyophthirius multifiliis (Ciliophora, parasito dominante e com dispersão agregada. Spironucleus sp. (Hexamitidae ocorreu somente nas brânquias de H. duriventris, enquanto Trinigyrus mourei (Monogenoidea e Procamallanus inopinatus (Camallanidae foram encontrados somente S. emarginata. O índice de Brillouin, riqueza de espécies de parasitos, uniformidade e dominância de Berger-Parker foram maiores em S. emarginata, mas paraambos os hospedeiros a comunidade parasitária foi caracterizada por baixa diversidade e baixa uniformidade. Em H. duriventris e S. emarginata, o baixo parasitismo não afetou o fator de condição relativo (Kn dos hospedeiros. Em S. emarginata houve correlação positiva da abundância de I. multifiliis com o comprimento, peso e Kn dos hospedeiros,enquanto que em H. druventris somente o peso e Kn mostram correlação com a abundância desse ciliado. Este foi o primeiro registro de I. multifiliis, Spironucleus sp. e P.inopinatus para H. duriventris e S. emarginata. Palavras-chave: Amazônia, Diversidade, Parasitos, Peixes de água doce, Metazoários. DOI: http://dx.doi.org/10.18561/2179-5746/biotaamazonia.v4n1p15-21
Langdon-Jones, Emily E; Lloyd, David; Hayes, Anthony J; Wainwright, Shane D; Mottram, Huw J; Coles, Simon J; Horton, Peter N; Pope, Simon J A
A range of fluorescent alkynyl-naphthalimide fluorophores has been synthesized and their photophysical properties examined. The fluorescent ligands are based upon a 4-substituted 1,8-naphthalimide core and incorporate structural variations (at the 4-position) to tune the amphiphilic character: chloro (L1), 4-[2-(2-aminoethoxy)ethanol] (L2), 4-[2-(2-methoxyethoxy)ethylamino] (L3), piperidine (L4), morpholine (L5), 4-methylpiperidine (L6), and 4-piperidone ethylene ketal (L7) variants. The amino-substituted species (L2-L7) are fluorescent in the visible region at around 517-535 nm through a naphthalimide-localized intramolecular charge transfer (ICT), with appreciable Stokes' shifts of ca. 6500 cm(-1) and lifetimes up to 10.4 ns. Corresponding two-coordinate Au(I) complexes [Au(L)(PPh3)] were isolated, with X-ray structural studies revealing the expected coordination mode via the alkyne donor. The Au(I) complexes retain the visible fluorescence associated with the coordinated alkynyl-naphthalimide ligand. The ligands and complexes were investigated for their cytotoxicity across a range of cell lines (LOVO, MCF-7, A549, PC3, HEK) and their potential as cell imaging agents for HEK (human embryonic kidney) cells and Spironucleus vortens using confocal fluorescence microscopy. The images reveal that these fluorophores are highly compatible with fluorescence microscopy and show some clear intracellular localization patterns that are dependent upon the specific nature of the naphthalimide substituent.
Parasite survey in mouse and rat colonies of Brazilian laboratory animal houses kept under differents sanitary barrier conditions Estudo de parasitos em colônias de ratos e de camundongos em biotérios brasileiros mantidos sob diferentes condições de barreiras sanitárias
Full Text Available A parasitological study was undertaken to determine the health status of 15 mouse and 10 rat colonies bred in 18 Brazilian laboratory animal houses maintained under different sanitary barrier conditions which supply animals for teaching, research purposes and manufacture of biological products for medical or veterinary use. Parasitological methods were used for diagnosis of mites, lices, helminthes and protozoan parasites. A questionnaire was answered by institutions with the intention to obtain information about the existence of barriers against infections and of regular sanitary monitoring program of their colonies. The questionnaire data show that the majority of the animal houses investigated do not possess an efficient sanitary barrier system able to keep animals under controlled health sanitary conditions. Ecto and endoparasite infections are widespread in the colonies and multiple infections were common in animals from most facilities investigated. The prevalences of parasites detected among the mouse and rat colonies of the laboratory animal houses investigated were: Myocoptes musculinus (46.6%, Myobia musculi (26.6%, Radfordia ensifera (13.3%, Syphacia obvelata (86.6%, Aspiculuris tetraptera (60.0%, Hymenolepis nana (53.3%, Spironucleus muris (80.0%, Tritrichomonas muris (80.0%, Giardia muris (66.0%, Entamoeba muris (20.0%, Eimeria sp. (13.3%, Hexamastix muris (26.6%, Poliplax spinulosa (30.0%, Poliplax serrata (10.0%, Radfordia ensifera (30.0%, Syphacia muris (80.0%, Hymenolepis nana (40.0%, Trichosomoides crassicauda (55.5%, Spironucleus muris (90.0%, Tritrichomonas muris (80.0%, Giardia muris (60.0%, Entamoeba muris (80.0%, Eimeria sp. (60.0% and Hexamastix muris (60.0%.Um estudo parasitológico foi realizado para verificar as condições de saúde de 15 colônias de camundongos e 10 colônias de ratos produzidos em 18 biotérios de instituições brasileiras que fornecem animais para ensino, pesquisa e produção de imunobiol
Aquaculture, including both freshwater and marine production, has on a world scale exhibited one of the highest growth rates within animal protein production during recent decades and is expected to expand further at the same rate within the next 10 years. Control of diseases is one of the most prominent challenges if this production goal is to be reached. Apart from viral, bacterial, fungal and metazoan infections it has been documented that protozoan parasites affect health and welfare and thereby production of fish in marine aquaculture. Representatives within the main protozoan groups such as amoebae, dinoflagellates, kinetoplastid flagellates, diplomonadid flagellates, apicomplexans, microsporidians and ciliates have been shown to cause severe morbidity and mortality among farmed fish. Well studied examples are Neoparamoeba perurans, Amyloodinium ocellatum, Spironucleus salmonicida, Ichthyobodo necator, Cryptobia salmositica, Loma salmonae, Cryptocaryon irritans, Miamiensis avidus and Trichodina jadranica. The present report provides details on the parasites' biology and impact on productivity and evaluates tools for diagnosis, control and management. Special emphasis is placed on antiprotozoan immune responses in fish and a strategy for development of vaccines is presented.
eukaryotes, and duplicated into HSP90AA and HSP90AB in vertebrates. Diplomonadida was identified as the most basal organism in the eukaryote lineage. Conclusion The present study presents the first comparative genomic study and evolutionary analysis of the HSP90 family of genes across all kingdoms of organisms. HSP90 family members underwent multiple duplications and also subsequent losses during their evolution. This study established an overall framework of information for the family of genes, which may facilitate and stimulate the study of this gene family across all organisms.
Williams, Catrin F; Lloyd, David; Kombrabail, M; Vijayalakshmi, K; Krishnamoorthy, G; White, Nick
The ability to resolve the spatio-temporal complexity of intracellular O 2 distribution is the ‘Holy Grail’ of cellular physiology. In an effort to obtain a non-invasive approach of mapping intracellular O 2 tensions, two methods of phosphorescent lifetime imaging microscopy were examined in the current study. These were picosecond time-resolved epiphosphorescence microscopy (single 0.5 µm focused spot) and two-photon confocal laser scanning microscopy with pinhole shifting. Both methods utilized nanoparticle-embedded Ru complex (45 nm diameter) as the phosphorescent probe, excited using pulsed outputs of Ti–sapphire Tsunami lasers (710–1050 nm). The former method used a 1 ps pulse width excitation beam with vertical polarization via a dichroic mirror (610 nm, XF43) and a 20× objective (NA 0.55, Nikon). Transmitted luminescence (1–2 × 10 4 counts s −1 ) was collected and time-correlated single photon counted decay times measured. Alternatively, an unmodified Zeiss LSM510 Confocal NLO microscope with 40× objective (NA 1.3) used successively shifted pinhole positions to collect image data from the lagging trail of the raster scan. Images obtained from two-photon excitation of a yeast (Schizosaccharomyces pombe) and a flagellate fish parasite (Spironucleus vortens), electroporated with Ru complex, indicated the intracellular location and magnitude of O 2 gradients, thus confirming the feasibility of optical mapping under different external O 2 concentrations. Both methods gave similar lifetimes for Ru complex phosphorescence under aerobic and anaerobic gas phases. Estimation of O 2 tensions within individual fibroblasts (human dermal fibroblast (HDF)) and mammary adenocarcinoma (MCF-7) cells was possible using epiphosphorescence microscopy. MCF-7 cells showed lower intracellular O 2 concentrations than HDF cells, possibly due to higher metabolic rates in the former. Future work should involve construction of higher resolution 3D maps of Ru coordinate
Williams, Catrin F.; Kombrabail, M.; Vijayalakshmi, K.; White, Nick; Krishnamoorthy, G.; Lloyd, David
The ability to resolve the spatio-temporal complexity of intracellular O2 distribution is the ‘Holy Grail’ of cellular physiology. In an effort to obtain a non-invasive approach of mapping intracellular O2 tensions, two methods of phosphorescent lifetime imaging microscopy were examined in the current study. These were picosecond time-resolved epiphosphorescence microscopy (single 0.5 µm focused spot) and two-photon confocal laser scanning microscopy with pinhole shifting. Both methods utilized nanoparticle-embedded Ru complex (45 nm diameter) as the phosphorescent probe, excited using pulsed outputs of Ti-sapphire Tsunami lasers (710-1050 nm). The former method used a 1 ps pulse width excitation beam with vertical polarization via a dichroic mirror (610 nm, XF43) and a 20× objective (NA 0.55, Nikon). Transmitted luminescence (1-2 × 104 counts s-1) was collected and time-correlated single photon counted decay times measured. Alternatively, an unmodified Zeiss LSM510 Confocal NLO microscope with 40× objective (NA 1.3) used successively shifted pinhole positions to collect image data from the lagging trail of the raster scan. Images obtained from two-photon excitation of a yeast (Schizosaccharomyces pombe) and a flagellate fish parasite (Spironucleus vortens), electroporated with Ru complex, indicated the intracellular location and magnitude of O2 gradients, thus confirming the feasibility of optical mapping under different external O2 concentrations. Both methods gave similar lifetimes for Ru complex phosphorescence under aerobic and anaerobic gas phases. Estimation of O2 tensions within individual fibroblasts (human dermal fibroblast (HDF)) and mammary adenocarcinoma (MCF-7) cells was possible using epiphosphorescence microscopy. MCF-7 cells showed lower intracellular O2 concentrations than HDF cells, possibly due to higher metabolic rates in the former. Future work should involve construction of higher resolution 3D maps of Ru coordinate complex lifetime
McKeon, T; Dunsmore, J; Raidal, S R
To estimate the prevalence of infection with Trichomonas gallinae and other parasites of the alimentary tract in psittacine and columbid birds in Perth and to determine in vitro the effectiveness of drugs commonly recommended for treating trichomoniasis. Samples of crop contents were collected from aviary flocks of budgerigars (Melopsittacus undulatus) and other psittacine and columbid birds in both private and commercial collections in Perth. Similar samples from wild Senegal doves (Streptopelia senegalensis) also were collected. Crop contents were examined and cultured for Trichomonas gallinae and in vitro studies were conducted on the susceptibility of isolates to several drugs used commonly. Other parasites also were detected by faecal examination and/or necropsy. T gallinae was recovered from birds in 1 of 13 private collections of budgerigars (2/289 birds in total). Direct wet-mount examination of crop fluid identified 36.4% of samples at four commercial bird dealers which were later determined by culture to contain T gallinae. The prevalence of T gallinae infection range from 0 to 11.4% in budgerigars. The prevalence of T gallinae infection of wild Senegal doves was 46% and from one flock of racing pigeons was 59%. The in vitro minimum lethal concentrations of metronidazole, dimetridazole and ronidazole ranged from 40 to 96, 30 to 80 and 40 to 92 micrograms/mL respectively for six isolates of T gallinae. Other alimentary parasites detected during the survey included Spironucleus sp (syn. Hexamita sp), coccidia, Ascaridia platycerci and Raillietina sp. Thirteen budgerigar flocks belonging to members of avicultural societies in Perth had a low prevalence of trichomoniasis and other parasitic infections. The dose rate currently recommended for ronidazole may not result in complete protozoacidal activity against T gallinae infection.