The purpose of this study was to compare the distribution of dinoflagellates in the Strait of Georgia, British Columbia with a number of environmental parameters, and in particular with vitamin B₁₂ concentrations. The latter was determined by a modified bioassay technique, employing the marine dinoflagellate, Amphidinium carterae Hulburt. The conclusions are based on the analysis of 511 samples from 19 cruises. The annual cycle of vitamin B₁₂ found to be characterized by three major peaks of concentration: 1) a peak following the spring 'bloom' of phytoplankton; 2) a summer increase that was closely associated with silt particles contained in river runoff; and 3) a fall maximum that followed the breakdown of density gradients in the water column. Seventy-seven species of dinoflagellates were recorded from the Strait of Georgia. The temporal distribution of dinoflagellates was divided into two distinct periods of abundance: one occurring during the early spring months and another during the summer months. The results of multiple regression analyses between the dinoflagellates and the environmental parameters measured (vitamin B₁₂, nitrogen, phosphorus, zooplankton, temperature, salinity and sunlight) indicated that the size of the dinoflagellate community in the early spring was closely associated with vitamin B₁₂ concentration in the water column. Nitrogen concentration was apparently a significant factor in determining the size of the community during the late summer months. Phosphorus concentration did not appear to be closely related with total dinoflagellate numbers. The composition and seasonal distribution of the dinoflagellates was found to be essentially similar to that of other northern temperate neritic waters. The major exception was the presence of a distinct spring component in the Strait of Georgia, consisting largely of small nonthecate species. / Science, Faculty of / Botany, Department of / Graduate

Viable cysts collected from natural sediments were induced to excyst. Ten cyst-theca relationships, first established elsewhere, were confirmed for British Columbia (B.C.). These were: Gonyaulax tamarensis, Protoperidinium aspidotum, P. claudicans, P. conicoides, P. conicum, P. cf. denticulatum, P. leonis, P. oblongum, and P. punctulatum. Five cyst-theca relationships were established for the first time: Peridiniopsis cf. hainanensis, Protoperidinium sp. nov., P. thorianum, and two apparently new species of Gonyaulax. P. pentagonum was found to have a cyst different from the cyst of this species in the Atlantic. Forty-five samples from Recent sediments were collected along the coast of B.C. Twenty-three of the samples had very few cysts. Hidden Basin was the chief source of viable cysts for the excystment experiments. Ten cyst-based taxa were described from the sediment samples. These were: Operculodinium centrocarpum, the cyst of Scrippsiella faeroense (= Micrhystridium bifurcatum), Spiniferites belerius, S. bentori, S. bulloideus, S. elongatus, S. membranaceus, S. nodosum, and S. ramosus. Tanyosphaeridium sp. has been recorded previously as the cyst of Polykrikos schwarzi. Two new cyst-based taxa are described for the first time. These are a cyst of Protoperidinium sp., and Spiniferites "sp. A". Cyst assemblages in the Recent sediments of B.C. were similar to many temperate estuarine and neritic areas. Some cysts which are characteristic of these areas in other regions, have not yet been found in B.C. The relative importance of some cysts also varies from that found in similar sediments elsewhere. The dominance of Operculodinium centrocarpum in many of the cyst assemblages, including B.C., is a pattern typical of temperate estuarine conditions. Some cysts appear to be characteristically associated with fjord environments. Scrippsiella faeroense, for example, has been found in Norwegian fjords and Scottish sea lochs as well as some B.C. fjords and inlets. / Science, Faculty of / Botany, Department of / Zoology, Department of / Graduate

As proteínas que contem domínios ricos em leucina (LRR) mediam interações macromoleculares que estão envolvidas em muitos processos biológicos como infecção bacteriana em células hospedeiras e respostas imunológicas de plantas. Estudos anteriores em nosso laboratório identificaram um gene que codifica uma proteína contendo 6 LRRs (LaLRR17) em L. (L.) amazonensis. O LaLRR17 é um gene com expressão estágio regulada sendo abundantemente expresso na fase amastigota. Seqüências homólogas ao gene LaLRR17 foram encontradas em todas as espécies de Leishmania analisadas. Esse trabalho tem como objetivo a caracterização da proteína homóloga em L. (L.) major (LmLRR17). Anticorpos obtidos contra seqüências conservadas das proteínas LaLRR17 e LmLRR17 permitiram o estudo da abundância protéica em diferentes estágios do parasita. Curiosamente, a proteína LmLRR17 foi encontrada em maior abundância em promastigotas procíclicos em vez de amastigotas. Linhagens hiperexpressoras da proteína LmLRR17 ou expressoras da proteína LaLRR17 em fusão com o epitopo viral myc foram obtidas. As proteínas quiméricas foram expressas seguindo o mesmo padrão observado na cepa selvagem. O fenótipo desses mutantes foi avaliado mediante infecções de macrófagos in vitro. A hiperexpressão da proteína LmLRR17 em L. (L.) major não alterou o fenótipo da infecção in vitro. Por outro lado, a expressão da proteína heteróloga, LaLRR17, em promastigotas de L. (L.) major levou a incremento na virulência com maior número de células infectadas e de parasitas por célula. Esses resultados indicam que a expressão da proteína LmLRR17 em L. (L.) major é fortemente regulada. Esse trabalho também mostra que a expressão da proteína LaLRR17 em L. (L.) major leva a um aumento na infectividade. / Proteins containing leucine rich repeats (LRR) are known to be involved in macromolecular interactions in many processes such as signal transduction, cell-adhesion, RNA processing, apoptosis, disease resistance and immune response. A previous study in our laboratory identified a L. (L.) amazonensis gene encoding a protein containing 6 LRRs (LaLRR17). LaLRR17 is a stage-regulated gene expressed with increased abundance in the amastigote stage. Highly conserved homologues of LaLRR17 were found in all Leishmania species analyzed. Therefore, the aim of this study was to characterize the homologous protein of L. major (LmLRR17). Antibodies raised against peptide sequences common to LaLRR17 and LmLRR17 allowed the study of the steady-state protein abundance. Interestingly, LmLRR17 protein was found to be up-regulated in procyclic promastigotes, instead of amastigotes. Mutants of L. (L.) major overexpressing a myc-tagged version of LmLRR17 or of LaLRR17 protein were obtained. In these parasites, the chimeric proteins were expressed following the same pattern of expression observed in the wild type parasites. The phenotype of these mutants was assessed in vitro through macrophage infections. Overexpression of LmLRR17 protein in L. (L.) major resulted in an unaltered phenotype. On the other hand, overexpression of LaLRR17 in L. (L.) major induced an increase in virulence with a higher number of infected cells and intracellular parasites. These results indicate that the expression of LmLRR17 protein in L. major is tightly regulated and the expression of the heterologous LaLRR17 protein increased infectivity in vitro.

As proteínas que contem domínios ricos em leucina (LRR) mediam interações macromoleculares que estão envolvidas em muitos processos biológicos como infecção bacteriana em células hospedeiras e respostas imunológicas de plantas. Estudos anteriores em nosso laboratório identificaram um gene que codifica uma proteína contendo 6 LRRs (LaLRR17) em L. (L.) amazonensis. O LaLRR17 é um gene com expressão estágio regulada sendo abundantemente expresso na fase amastigota. Seqüências homólogas ao gene LaLRR17 foram encontradas em todas as espécies de Leishmania analisadas. Esse trabalho tem como objetivo a caracterização da proteína homóloga em L. (L.) major (LmLRR17). Anticorpos obtidos contra seqüências conservadas das proteínas LaLRR17 e LmLRR17 permitiram o estudo da abundância protéica em diferentes estágios do parasita. Curiosamente, a proteína LmLRR17 foi encontrada em maior abundância em promastigotas procíclicos em vez de amastigotas. Linhagens hiperexpressoras da proteína LmLRR17 ou expressoras da proteína LaLRR17 em fusão com o epitopo viral myc foram obtidas. As proteínas quiméricas foram expressas seguindo o mesmo padrão observado na cepa selvagem. O fenótipo desses mutantes foi avaliado mediante infecções de macrófagos in vitro. A hiperexpressão da proteína LmLRR17 em L. (L.) major não alterou o fenótipo da infecção in vitro. Por outro lado, a expressão da proteína heteróloga, LaLRR17, em promastigotas de L. (L.) major levou a incremento na virulência com maior número de células infectadas e de parasitas por célula. Esses resultados indicam que a expressão da proteína LmLRR17 em L. (L.) major é fortemente regulada. Esse trabalho também mostra que a expressão da proteína LaLRR17 em L. (L.) major leva a um aumento na infectividade. / Proteins containing leucine rich repeats (LRR) are known to be involved in macromolecular interactions in many processes such as signal transduction, cell-adhesion, RNA processing, apoptosis, disease resistance and immune response. A previous study in our laboratory identified a L. (L.) amazonensis gene encoding a protein containing 6 LRRs (LaLRR17). LaLRR17 is a stage-regulated gene expressed with increased abundance in the amastigote stage. Highly conserved homologues of LaLRR17 were found in all Leishmania species analyzed. Therefore, the aim of this study was to characterize the homologous protein of L. major (LmLRR17). Antibodies raised against peptide sequences common to LaLRR17 and LmLRR17 allowed the study of the steady-state protein abundance. Interestingly, LmLRR17 protein was found to be up-regulated in procyclic promastigotes, instead of amastigotes. Mutants of L. (L.) major overexpressing a myc-tagged version of LmLRR17 or of LaLRR17 protein were obtained. In these parasites, the chimeric proteins were expressed following the same pattern of expression observed in the wild type parasites. The phenotype of these mutants was assessed in vitro through macrophage infections. Overexpression of LmLRR17 protein in L. (L.) major resulted in an unaltered phenotype. On the other hand, overexpression of LaLRR17 in L. (L.) major induced an increase in virulence with a higher number of infected cells and intracellular parasites. These results indicate that the expression of LmLRR17 protein in L. major is tightly regulated and the expression of the heterologous LaLRR17 protein increased infectivity in vitro.