Eric Viscogliosi

Molecular Phylogeny of Parabasalids Based on Small Subunit rRNA Sequences, with Emphasis on the Trichomonadinae Subfamily

Abstract We determined small subunit ribosomal DNA sequences from three parabasalid species, Trichomitus batrachorum strain R105, Tetratrichomonas gallinarum, and Pentatrichomonas hominis belonging to the Trichomonadinae subfamily. Unrooted molecular phylogenetic trees inferred by distance, parsimony, and likelihood methods reveal four discrete clades among the parabasalids. TheTrichomonadinae form a robust monophyletic group. Within this subfamily T. gallinarum is closely related to Trichomonas species as supported by morphological data, with P. hominis and Pseudotrypanosoma giganteum occupying basal positions. Our analysis does not place T. batrachorum within the Trichomonadinae. Trichomitus batrachorum (strains R105 and BUB) and Hypotrichomonas acosta form a well-separated cluster, suggesting the genus Trichomitus is polyphyletic. The emergence of T. batrachorum precedes the Trichomonadinae-Tritrichomonadinae dichotomy, emphasizing its pivotal evolutionary position among the Trichomonadidae. A third cluster unites the Devescovinidae and the Calonymphidae. The fourth clade contains the three hypermastigid sequences from the genus Trichonympha, which exhibit the earliest emergence among the parabasalids. The addition of these three new parabasalid species did not however resolve ambiguities regarding the relative branching order of the parabasalid clades. The phylogenetic positions of Tritrichomonas fœtus, Monocercomonas sp., Dientamoeba fragilis, and the unidentified Reticulitermes flavipes gut symbiont 1 remain unclear.

Tubulin post‐translational modifications in the primitive protist Trichomonas vaginalis

Using several specific monoclonal antibodies, we investigated the occurrence and distribution of different post-translationally modified tubulin during interphase and division of the primitive flagellated protist Trichomonas vaginalis. Immunoblotting and immunofluorescence experiments revealed that interphasic microtubular structures of T. vaginalis contained acetylated and glutamylated but non-tyrosinated and non-glycylated [Brugerolle and Adoutte, 1988: Bio Systems 21: 255-268] tubulin. Immunofluorescence studies performed on dividing cells showed that the extranuclear mitotic spindle (or paradesmosis) was acetylated and glutamylated, which contrast with the ephemeral nature of this structure. Newly formed short axostyles also contained acetylated and glutamylated tubulin suggesting that both post-translational modifications might take place very early after assembly of microtubular structures. Our results indicate that acetylation and glutamylation of tubulin appeared early in the history of eukaryotes and could reflect the occurrence of post-translational modifications of tubulin in the primitive eukaryotic cells. These cells probably had a highly ordered cross-linked microtubular cytoskeleton in which microtubules showed a low level of subunit exchange dynamics.

New Insights into the Phylogeny of Trichomonads Inferred from Small Subunit rRNA Sequences

Small subunit ribosomal DNA sequences were obtained by polymerase chain reaction from four trichomonad species: a frog endosymbiont Trichomitus batrachorum, an intestinal endosymbiont of a squamate reptile, Hypotrichomonas acosta and two free-living isolates, Monotrichomonas carabina and Monotrichomonas sp. Molecular trees inferred by distance, parsimony and likelihood techniques identify three well-resolved clusters within the trichomonads, however bootstrap values do not strongly support a particular branching order for these lineages. The first cluster includes the Devescovinidae and the Calonymphidae. The second clade unites Trichomitus batrachorum and Hypotrichomonas acosta. The third cluster embraces all known free-living genera, including Monotrichomonas, and various members of the Trichomonadinae subfamily such as Trichomonas vaginalis, and Pentatrichomonoides scroa. Neither Monocercomonadidae nor the Trichomonadidae as envisaged are monophyletic. Most of the monocercomonads, which possess a rudimentary cytoskeleton, were likely descendants of more complex forms. The study also suggests that the genus Trichomitus is currently polyphyletic, partly explaining the discordant positions of this genus in previous molecular analyses.

Cloning and characterization of iron-containing superoxide dismutase from the human malaria species Plasmodium ovale, P. malariae and P. vivax

Abstract The iron-containing superoxide dismutase (FeSOD) gene from three human malaria species, namely Plasmodium ovale, P. malariae and P. vivax, was amplified by polymerase chain reaction, cloned and then sequenced. Comparisons of their deduced amino acid sequences with that of the FeSOD from P. falciparum revealed a very low polymorphism at the FeSOD locus in human malaria species. One P. ovale and the P. vivax FeSOD genes presented the same nucleotide sequence as that of the P. falciparum strain HB3 whereas the second P. ovale and the P. malariae genes exhibited two punctual mutations. These mutations did not affect the function and structure of the enzyme. The FeSOD polymorphism was so low that no phylogenetic relationship among human malaria species could be proposed, but this conservative structure strengthened the potentiality of this enzyme as a possible target for anti-malarial drugs.

Cytoskeleton and morphogenesis in opalinid protozoa

Opa l ln ids a r e m u l t l n u c l e a t e d astom Protozoa . The c e l l su r f ace i s covered by c i l i a ar ranged in l o n g l t u d l n a l somatic k l n e t l e s . In the f r o n t a l r eg ion , t h e r e i s a second group of l e s s arranged c i l i a forming the f a l x zone s. E l e c t r o n mic roscop ica l s t u d i e s show tha t 2 types of c y t o s k e l e t a l s t r u c t u r e s o r i g i n a t e from t h i s p a r t l c u l a r r eg ion : ( I ) I o n g l t u d i n a l mic ro tubu les tha t emerge from a f l b r i l l a r ma t r ix embedding the klnetosomes of the f a l x zone and (2) m l c r o f i b r i l l a r s t r i n g s tha t l l n e the somat ic k l n e t l e s on t h e i r l e f t s i de . In the somatic k i n e t l e s basa l bodies a re l inked t o g e t h e r by a forked desmosls . Closer to the c e l l su r f ace t he re i s a t r a n s v e r s a l m i c r o f l b r i l l a r network. S i l v e r s t a l n l n g and l l g h t microscopy show t h a t new k i n e t l e s o r i g i n a t e around the f a lx zone and I n t e r c a l a t e between the o ld ones , sugges t ing an a n t e r o ~ p o s t e r i o r morphogenetic g r a d i e n t . Cons ider ing the d l r e c t l o n n a l arrangement of somatic basa l bodies along the m i c r o f l b r i l l a r s t r i n g s , the connec t ion between these s t r u c t u r e s , the v a r i a t i o n of i n t e r k l n e t o s o m a l spaces and f l n a l l y the absence of some basa l bodies along the m i c r o f l b r i l l a r s t r i p s , a r o l e of the microf i b r i l s i n the p o s l t l o n n l n g of the somatlc basa l bodies may be reasonnably env i saged . The space between the I o n g l t u d l n a l m lc ro tubu le s and between the k l n e t l e s could be c o n t r o l l e d by the t r a n s v e r s a l network. A p r e l i m i n a r y i n v e s t l g h t l o n demons t ra tes t ha t an aa t lbody r a i s e d aga ins t c e n t r i n l l k e p ro t e ln s r ecogn izes the f i l amentous s t r i p s in Immunofluorescence and a p r o t e l n double t band of 19-20 kd in Inununoblots. Thus, m l c r o f l b r i l l a r s t r l p s of Opallnld~ might be remin i scen t of c e n t r i n based f i l amen t systems whose r o l e in basa l body I e e n t r i o l e p o s i t l o n n l n g has been p r e v i o u s l y repor ted in o the r p r o t l s t s and in p l u r l c e l l u l a r organisms.