Acidotropic probes and flow cytometry: a powerful combination for detecting phagotrophy in mixotrophic and heterotrophic protists

Carvalho, Wanderson Fernandes de; Granéli, Edna

doi:10.3354/ame044085

Cited by 33 publications

(30 citation statements)

References 52 publications

(79 reference statements)

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“…By using a modern method that combines the fluorescence of an acidotropic probe and flow cytometry (Carvalho and Graneli, 2006), we were able to analyze a greater number of D. norvegica cells in a shorter time than otherwise possible by using conventional microscopy. Besides, this technique allows the analysis of live cells from cultures or natural populations in real time excluding experimental artifacts and distortions due to fixation.…”

Section: Discussionmentioning

confidence: 99%

Dinophysis norvegica (Dinophyceae), more a predator than a producer?

Carvalho¹,

Minnhagen²,

Granéli³

2008

Harmful Algae

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Section: Discussionmentioning

confidence: 99%

Dinophysis norvegica (Dinophyceae), more a predator than a producer?

Carvalho¹,

Minnhagen²,

Granéli³

2008

Harmful Algae

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“…Given the challenge of obtaining pure Dinophysis samples from natural microbial assemblages for DNA and RNA extraction, isolation of these cells can be made more efficient with the use of flow cytometry (6). Furthermore, dinoflagellate-specific primers can significantly reduce the amplification of gene homologs in other organisms still present in the sorted cell samples.…”

mentioning

confidence: 99%

Mitochondrial cob and cox1 Genes and Editing of the Corresponding mRNAs in Dinophysis acuminata from Narragansett Bay, with Special Reference to the Phylogenetic Position of the Genus Dinophysis

Zhang

Bhattacharya

Maranda

et al. 2008

Appl Environ Microbiol

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Dinophysis acuminata cells were isolated from Narragansett Bay water samples in June 2005 using flow cytometry. Dinoflagellate-specific PCR primers were used to isolate small-subunit rRNA (18S rRNA), mitochondrial cytochrome b (cob), and cytochrome c oxidase I (cox1) genes and the encoded cDNAs. Maximumlikelihood analysis of a concatenated data set of ribosomal DNA and cDNA sequences of cob and cox1 showed that D. acuminata was sister to Gonyaulacoids, but without strong bootstrap support. The approximately unbiased test could not reject alternative positions of D. acuminata. To gain better resolution, mRNA editing of cob and cox1 was inferred for D. acuminata and 13 other dinoflagellate species. The location and type of editing as well as the distribution pattern in D. acuminata were generally similar to those in other dinoflagellates except for two edited sites that are unique to this species. Bayesian analyses of a matrix that recorded the location and type of editing, and of a matrix that included the protein sequences of COB and COX1 with the editing data yielded tree topologies similar to the three-gene tree but again failed to resolve the phylogenetic position of D. acuminata. However, the density of edited sites in the D. acuminata mitochondrial genes, consistent with phylogenetic trees, indicated that Dinophysis is a derived dinoflagellate lineage, diverging after other lineages such as Oxyrrhis, Amphidinium, and Symbiodinium. We demonstrate that dinoflagellate-specific PCR coupled with flow cytometry can be a useful tool to analyze genes and their transcripts from a natural dinoflagellate population.

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“…To identify mixotrophic nutrition based on the observation of particles inside a protist cell by light microscopy is challenging (CARVALHO; GRANÉLI, 2006). Chloroplasts may mask food vacuoles and it can be problematic to distinguish food vacuoles from other types of inclusions such as gametes or parasites (GRANÉLI; LEGRAND et al, 1998;CARVALHO and GRANÉLI, 2006). In an early stage of the infection, parasites can be easily misidentified as prey.…”

Section: Discussionmentioning

confidence: 99%

Investigation of phagotrophy in natural assemblages of the benthic dinoflagellates Ostreopsis, Prorocentrum and Coolia

2017

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Almada et al.: Phagotrophy assessment in benthic dinoflagellates 392 Investigation of phagotrophy in natural assemblages of the benthic dinoflagellates Ostreopsis, Prorocentrum and CooliaMixotrophy has been shown to be a common trait among dinoflagellates and its importance in the nutritional ecology of harmful algae has been hypothesized. Benthic harmful species have not been extensively investigated as their planktonic counterparts and there are major gaps in the knowledge of their nutritional strategies. In this study the occurrence of phagotrophy was investigated in natural assemblages of benthic dinoflagellates using epi-fluorescence microscopy with DAPI and LysoSensor staining. The study was conducted at five sites along the coast of Rio de Janeiro that were visited in January, August and December 2010. In total, 1659 dinoflagellate cells were observed. From these, only 0.4% of 1195 Ostreopsis cf. ovata and 2.2% of 134 Coolia spp. cells presented evidence of phagotrophy with vacuoles stained by LysoSensor or a DAPI (4',6-diamidino-2-phenylindole) stained inclusion. Stained vacuoles were not registered in the 330 Prorocentrum spp. cells observed. Few O. cf. ovata cells contained round red inclusions ("red spots") that were not stained either by DAPI or LysoSensor, suggesting that these structures are not ingested prey. The results showed that phagotrophy was not a frequent nutritional strategy in benthic dinoflagellates during the study period. AbstrAct Descriptors:Dinoflagellates, LysoSensor, Mixotrophy, Phagotrophy.A mixotrofia tem se mostrado uma característica comum entre dinoflagelados e a sua importância na ecologia nutricional de algas nocivas vem sendo investigada. Espé-cies bentônicas nocivas não foram tão estudadas quanto as planctônicas, e existem grandes lacunas no conhecimento de suas estratégias nutricionais. A ocorrência de fagotrofia em comunidades naturais de dinoflagelados bentônicos foi investigada usando microscopia de epi--fluorescência e marcação por DAPI e LysoSensor. O estudo foi realizado em cinco locais ao longo da costa do Rio de Janeiro nos meses de janeiro, agosto e dezembro de 2010. No total, foram observadas 1659 cé-lulas de dinoflagelados. Destas, apenas 0,4% das 1195 células de Ostreopsis cf. ovata e 2,2% das 134 células de Coolia spp. observadas apresentaram evidências de fagotrofia com vacúolos marcados por LysoSensor ou inclusão celular marcada por DAPI (4', 6-diamidino-2--phenylindole). Não foram observados vacúolos marcados nas 330 células de Prorocentrum spp. observadas. Células de O. cf. ovata apresentaram inclusões esféricas avermelhadas ("red spots") que não foram coradas por DAPI e LysoSensor, sugerindo que estas estruturas não correspondem a presas ou a vacúolos digestivos. Os resultados mostraram que a fagotrofia não foi uma estraté-gia nutricional frequente nas espécies de dinoflagelados bentônicos observadas durante o período de estudo. resumoDescritores: Dinoflagelados, LysoSensor, Mixotrofia, Fagotrofia.

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Acidotropic probes and flow cytometry: a powerful combination for detecting phagotrophy in mixotrophic and heterotrophic protists

Cited by 33 publications

References 52 publications

Dinophysis norvegica (Dinophyceae), more a predator than a producer?

Dinophysis norvegica (Dinophyceae), more a predator than a producer?

Mitochondrial cob and cox1 Genes and Editing of the Corresponding mRNAs in Dinophysis acuminata from Narragansett Bay, with Special Reference to the Phylogenetic Position of the Genus Dinophysis

Investigation of phagotrophy in natural assemblages of the benthic dinoflagellates Ostreopsis, Prorocentrum and Coolia

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