Improvement of digital Gabor holographic microscopy using a lens in plankton studies.

This work utilizes a Gabor Holographic Optical Scheme integrated with a microscope objective and a thin convex plane lens. This bi-telecentric lens system corrects spherical aberration from the objective, maintains consistent magnification across various reconstruction distances, and ensures a plane incidence on CMOS. Depending on the focal lengths of the objective and lens, the final image can be enlarged or reduced compared to the classic Gabor system, resulting in high-quality reconstructed phase images without spherical aberration. This setup was employed to capture phase distribution and intensity images of planktonic objects, such as copepods, achieving superior image quality.

Teamwork in the viscous oceanic microscale.

Nutrient acquisition is crucial for oceanic microbes, and competitive solutions to solve this challenge have evolved among a range of unicellular protists. However, solitary solutions are not the only approach found in natural populations. A diverse array of oceanic protists form temporary or even long-lasting attachments to other protists and marine aggregates. Do these planktonic consortia provide benefits to their members? Here, we use empirical and modeling approaches to evaluate whether the relationship between a large centric diatom, Coscinodiscus wailesii, and a ciliate epibiont, Pseudovorticella coscinodisci, provides nutrient flux benefits to the host diatom. We find that fluid flows generated by ciliary beating can increase nutrient flux to a diatom cell surface four to 10 times that of a still cell without ciliate epibionts. This cosmopolitan species of diatom does not form consortia in all environments but frequently joins such consortia in nutrient-depleted waters. Our results demonstrate that symbiotic consortia provide a cooperative alternative of comparable or greater magnitude to sinking for enhancement of nutrient acquisition in challenging environments.

The epibiotic life of the cosmopolitan diatom Fragilariopsis doliolus on heterotrophic ciliates in the open ocean.

Diatoms are a diverse and ecologically important group of phytoplankton. Although most species are considered free living, several are known to interact with other organisms within the plankton. Detailed imaging and molecular characterization of any such partnership is, however, limited, and an appraisal of the large-scale distribution and ecology of such consortia was never attempted. Here, observation of Tara Oceans samples from the Benguela Current led to the detection of an epibiotic association between a pennate diatom and a tintinnid ciliate. We identified the diatom as Fragilariopsis doliolus that possesses a unique feature to form barrel-shaped chains, associated with seven different genera of tintinnids including five previously undescribed associations. The organisms were commonly found together in the Atlantic and Pacific Ocean basins, and live observations of the interaction have been recorded for the first time. By combining confocal and scanning electron microscopy of individual consortia with the sequencing of high-resolution molecular markers, we analyzed their distribution in the global ocean, revealing morpho-genetically distinct tintinnid haplotypes and biogeographically structured diatom haplotypes. The diatom was among the most abundant in the global ocean. We show that the consortia were particularly prevalent in nutrient-replete conditions, rich in potential predators. These observations support the hypothesis of a mutualistic symbiosis, wherein diatoms acquire increased motility and tintinnids benefit from silicification through increased protection, and highlight that such associations may be more prevalent than currently appreciated.

Morphological and molecular characterization of the toxic dinoflagellate Ostreopsis cf. ovata (Gonyaulacales: Dinophyceae) from Brazil (South Atlantic Ocean) / Caracterización morfológica y molecular del dinoflagelado tóxico Ostreopsis cf. ovata (Gonyaulacales: Dinophyceae) en Brasil (Océano Atlántico Sur)

AbstractOstreopsis cf. ovata is a toxic epiphytic dinoflagellate widely distributed in warm waters that often co-occur with species of the genera Coolia, Fukuyoa, Gambierdiscus and Prorocentrum. We investigated a strain isolated from the coast of Ubatuba, Brazil (Southwest Atlantic Ocean) by light and epifluorescence microscopies; we also report molecular data based on the LSU rDNA and ITS markers. Cells were 35-65 µm in the dorso-ventral diameter and 20-40 µm wide. We obtained the sequence of a ~1 900 base pair region of the rRNA gene cistron. In the LSU rDNA phylogeny, the sequences under the names O. ovata and O. cf. ovata branched into three clades. The ITS marker showed greater resolving power and the sequences of O. ovata/O. cf. ovata split into five clades. Our ITS sequence branched in a clade with sequences of strains from the Mediterranean Sea, European Atlantic coasts, subtropical NE Atlantic, other sequences from Brazil at Rio de Janeiro, and a few sequences from Japan. The cell dimensions and thecal plate arrangement were under the variability range reported in other ocean regions. Our observations confirm O. cf. ovata as the most commonly recorded species of Ostreopsis in the SW Atlantic Ocean. Ostreopsis cf. ovata co-occurred with Coolia malayensis in Brazil and Asia, but it has been commonly reported from the Mediterranean Sea, where C. malayensis has not yet been recorded; while Coolia malayensis has been reported from the Caribbean Sea, but not O. ovata. With the current knowledge, it is difficult to understand the factors that determine the biogeography of the tropical epiphytic dinoflagellates. Rev. Biol. Trop. 65 (3): 1022-1032. Epub 2017 September 01.

ResumenOstreopsis cf. ovata es un dinoflagelado tóxico epifítico de amplia distribución en aguas cálidas, que a menudo coincide con especies de los géneros Coolia, Fukuyoa, Gambierdiscus y Prorocentrum. Investigamos una cepa aislada en la costa de Ubatuba, Brasil (Atlántico sudoccidental) mediante microscopía óptica y de epifluorescencia. Obtuvimos una secuencia de una región de unos 1 900 pares de bases del cistrón del gen del ARN ribosómico. Las células tenían 35-65 µm de diámetro dorso-ventral y 20-40 µm de ancho. En la filogenia del marcador LSU rADN, las secuencias con los nombres O. ovata and O. cf. ovata se sitúan en tres grupos. El marcador ITS mostraba un mayor poder resolutivo y las secuencias de O. ovata/O. cf. ovata se separan en cinco grupos. Nuestra secuencia ITS se sitúa en un grupo con secuencias de cepas procedentes del Mar Mediterráneo, costas europeas Atlánticas, Atlántico subtropical nororiental, otras secuencias procedentes de Río de Janeiro en Brasil, y algunas secuencias de Japón. Las dimensiones celulares y la disposición de las placas tecales se sitúan en el rango de variabilidad descrito en otras regiones oceánicas. Nuestras observaciones confirman a O. cf. ovata como la especie más comúnmente registrada de Ostreopsis en el Atlántico sudoccidental. Ostreopsis cf. ovata coindice con Coolia malayensis en Brasil y Asia. Ostreopsis cf. ovata ha sido comúnmente encontrada en el Mar Mediterráneo, donde C. malayensis aún no ha sido registrada. Coolia malayensis has sido registrada en el Mar Caribe, donde O. ovata aún no ha sido encontrada. Es difícil comprender los factores que determinan la biogeografía de los dinoflagelados epífitos tropicales, a partir del conocimiento actual.

Morphological and molecular characterization of Ptychodiscus noctiluca revealed the polyphyletic nature of the order Ptychodiscales (Dinophyceae).

The planktonic dinoflagellate Ptychodiscus noctiluca combined distinctive morphological features such as a disk-shaped anteroposteriorly compressed cell body and an apical carina, together with a flexible and tough cell covering, suggesting intermediate characteristics between thecate and naked dinoflagellates. Ptychodiscus noctiluca was examined by light, epifluorescence, and scanning electron microscopy from specimens collected in the Mediterranean Sea and the North and South Atlantic Ocean. Ptychodiscus noctiluca showed a straight apical groove that bisected the carina, a cell covering with a polygonal surface reticulum, nucleus without capsule, sulcal intrusion in the episome, sulcal ventral flange, and yellowish-green chloroplasts that are shared characters with Brachidinium/Karenia. The cell division was the typical binary fission of gymnodinioid dinoflagellates, although exceptionally in an oblique transversal axis. We examined the intraspecific variability during incubation experiments. In the fattened cells, termed as Ptychodiscus carinatus, chloroplasts transformed into dark granules, and the cell acquired the swollen and smaller stage, termed as P. inflatus. Ptychodiscus carinatus, P. inflatus, and Diplocystis antarctica are synonyms of P. noctiluca. Molecular phylogeny based on the SSU rDNA sequence revealed that Ptychodiscus branched within the short-branching dinokaryotic dinoflagellates as an independent lineage with affinity to Brachidinium/Karenia and Karlodinium/Takayama in a generally poorly resolved clade. Our results indicated that the order Ptychodiscales, established for unarmored dinoflagellates with a strongly developed pellicle, has artificially grouped thecate dinoflagellates (Kolkwitziella, Herdmania), naked dinoflagellates with thick cell covering (Balechina, Cucumeridinium) and other insufficiently known unarmored genera with typical cell coverings (Brachidinium, Ceratoperidinium).

Sample size for estimating the mean concentration of organisms in ballast water.

We consider the computation of sample sizes for estimating the mean concentration of organisms in ballast water. Given the possible heterogeneity of their distribution in the tank, we adopt a negative binomial model to obtain confidence intervals for the mean concentration. We show that the results obtained by Chen and Chen (2012) in a different set-up hold for the proposed model and use them to develop algorithms to compute sample sizes both in cases where the mean concentration is known to lie in some bounded interval or where there is no information about its range. We also construct simple diagrams that may be easily employed to decide for compliance with the D-2 regulation of the International Maritime Organization (IMO).

Fukuyoa paulensis gen. et sp. nov., a new genus for the globular species of the dinoflagellate Gambierdiscus (Dinophyceae).

The marine epiphytic dinoflagellate Gambierdiscus is a toxicologically important genus responsible for ciguatera fish poisoning, the principal cause of non-bacterial illness associated with fish consumption. The genus currently contains species exhibiting either globular or anterior-posteriorly compressed morphologies with marked differences in cell shape and plate arrangement. Here we report a third globular, epiphytic and tychoplanktonic species from the coasts of Ubatuba, Brazil. The new species can be distinguished from G. yasumotoi and G. ruetzleri by its broader first apical plate that occupies a larger portion of the epitheca. Accordingly, phylogenetic trees from small subunit (SSU) and large subunit (LSU) ribosomal DNA sequences also showed strongly supported separation of the new species from the G. yasumotoi/G. ruetzleri group albeit with short distance. The molecular phylogenies, which included new sequences of the planktonic species Goniodoma polyedricum, further indicated that the globular species of Gambierdiscus formed a tight clade, clearly separated (with strong bootstrap support) from the clade of lenticular species including the type for Gambierdiscus. The morphological and molecular data in concert support the split of Gambierdiscus sensu lato into two genera. Gambierdiscus sensu stricto should be reserved for the species with lenticular shapes, highly compressed anterioposteriorly, with short-shank fishhook apical pore plate, large 2' plate, low and ascending cingular displacement, and pouch-like sulcal morphology. The new genus name Fukuyoa gen. nov. should be applied to the globular species, slightly laterally compressed, with long-shank fishhook apical pore plate, large 1' plate, greater and descending cingular displacement, and not pouch-like vertically-oriented sulcal morphology. Fukuyoa contains the new species Fukuyoa paulensis gen. et sp. nov., and F. yasumotoi comb. nov. and F. ruetzleri comb. nov.

Diversity and ecological structure of vibrios in benthic and pelagic habitats along a latitudinal gradient in the Southwest Atlantic Ocean.

We analyzed the diversity and population structure of the 775 Vibrio isolates from different locations of the southwestern Atlantic Ocean (SAO), including St. Peter and St. Paul Archipelago (SPSPA), Abrolhos Bank (AB) and the St. Sebastian region (SS), between 2005 and 2010. In this study, 195 novel isolates, obtained from seawater and major benthic organisms (rhodoliths and corals), were compared with a collection of 580 isolates previously characterized (available at www.taxvibrio.lncc.br). The isolates were distributed in 8 major habitat spectra according to AdaptML analysis on the basis of pyrH phylogenetic reconstruction and ecological information, such as isolation source (i.e., corals: Madracis decactis, Mussismilia braziliensis, M. hispida, Phyllogorgia dilatata, Scolymia wellsi; zoanthids: Palythoa caribaeorum, P. variabilis and Zoanthus solanderi; fireworm: Hermodice carunculata; rhodolith; water and sediment) and sampling site regions (SPSPA, AB and SS). Ecologically distinct groups were discerned through AdaptML, which finds phylogenetic groups that are significantly different in their spectra of habitat preferences. Some habitat spectra suggested ecological specialization, with habitat spectra 2, 3, and 4 corresponding to specialization on SPSPA, AB, and SS, respectively. This match between habitat and location may reflect a minor exchange of Vibrio populations between geographically isolated benthic systems. Moreover, we found several widespread Vibrio species predominantly from water column, and different populations of a single Vibrio species from H. carunculata in ecologically distinct groups (H-1 and H-8 respectively). On the other hand, AdaptML detected phylogenetic groups that are found in both the benthos and in open water. The ecological grouping observed suggests dispersal and connectivity between the benthic and pelagic systems in AB. This study is a first attempt to characterize the biogeographic distribution of vibrios in both seawater and several benthic hosts in the SAO. The benthopelagic coupling observed here stands out the importance of vibrios in the global ocean health.

Implications of heterogeneous distributions of organisms on ballast water sampling.

Ballast water sampling is one of the problems still needing investigation in order to enforce the D-2 Regulation of the International Convention for the Control and Management of Ship Ballast Water and Sediments. Although statistical "representativeness" of the sample is an issue usually discussed in the literature, neither a definition nor a clear description of its implications are presented. In this context, we relate it to the heterogeneity of the distribution of organisms in ballast water and show how to specify compliance tests under different models based on the Poisson and negative binomial distributions. We provide algorithms to obtain minimum sample volumes required to satisfy fixed limits on the probabilities of Type I and II errors. We show that when the sample consists of a large number of aliquots, the Poisson model may be employed even under moderate heterogeneity of the distribution of the organisms in the ballast water tank.

Genome sequence of the bacterioplanktonic, mixotrophic Vibrio campbellii strain PEL22A, isolated in the Abrolhos Bank.

Vibrio campbellii PEL22A was isolated from open ocean water in the Abrolhos Bank. The genome of PEL22A consists of 6,788,038 bp (the GC content is 45%). The number of coding sequences (CDS) is 6,359, as determined according to the Rapid Annotation using Subsystem Technology (RAST) server. The number of ribosomal genes is 80, of which 68 are tRNAs and 12 are rRNAs. V. campbellii PEL22A contains genes related to virulence and fitness, including a complete proteorhodopsin cluster, complete type II and III secretion systems, incomplete type I, IV, and VI secretion systems, a hemolysin, and CTXΦ.

Free-living and plankton-associated vibrios: assessment in ballast water, harbor areas, and coastal ecosystems in Brazil.

Ballast water (BW) is a major transport vector of exotic aquatic species and pathogenic microorganisms. The wide-ranging spread of toxigenic Vibrio cholerae O1 from harbor areas has been frequently ascribed to discharge of contaminated BW into eutrophic coastal environments, such as during the onset of the seventh cholera pandemic in South America in the early 1990s. To determine the microbiological hazards of BWs transported to Brazilian ports, we evaluated water and plankton samples taken from (i) BW tanks of recently arrived ships, (ii) port areas along the Brazilian coastline from ∼1 to 32°S and (iii) three coastal areas in São Paulo State. Vibrio concentration and toxigenic V. cholerae O1 occurrence were analyzed. Plankton-associated vibrios were more abundant than free-living vibrios in all studied environments. V. cholerae was found in 9.5% of ballast tanks and 24.2% of port samples, both as free-living and attached forms and, apart from the Santos harbor, was absent off São Paulo State. Toxigenic V. cholerae O1 isolates (ctxA(+), tcpA(+)), involved in cholera disease, were found in BW (2%) and harbor (2%) samples. These results confirm that BW is an important carrier of pathogenic organisms, and that monitoring of vibrios and other plankton-attached bacteria is of paramount importance in BW management programs.

Marine zooplankton studies in Brazil: a brief evaluation and perspectives.

Marine zooplankton research in Brazil has been primarily descriptive, with most studies focusing on community structure analysis and related issues. The composition and spatial distribution of several taxonomic groups are currently well known, although less-abundant and small-sized taxa as well as initial stages of almost all species have received little attention. Some numerically important taxa such as heterotrophic protists, ctenophores, acoel turbellarians and ostracods remain virtually unstudied. Large sectors of the continental shelf have not been sampled in detail, particularly those areas influenced by the North Brazil Current (5 degrees N-15 degrees S). Zooplankton abundance and biomass in offshore waters have seldom been quantified, and information on the distribution and vertical migration of meso- and bathypelagic species are lacking. Additional faunistic assessments must target those less-studied taxa and geographical locations. However, priority in ecological studies should be given to process-oriented investigations aimed at understanding the mechanisms controlling zooplankton distribution, trophic interactions within pelagic food webs and production cycles in relation to the physical environment. An effort should be made to incorporate state-of-the-art sampling technology and analytical methods into future research projects.

Marine zooplankton studies in Brazil: a brief evaluation and perspectives

Marine zooplankton research in Brazil has been primarily descriptive, with most studies focusing on community structure analysis and related issues. The composition and spatial distribution of several taxonomic groups are currently well known, although less-abundant and small-sized taxa as well as initial stages of almost all species have received little attention. Some numerically important taxa such as heterotrophic protists, ctenophores, acoel turbellarians and ostracods remain virtually unstudied. Large sectors of the continental shelf have not been sampled in detail, particularly those areas influenced by the North Brazil Current (5°N-15°S). Zooplankton abundance and biomass in offshore waters have seldom been quantified, and information on the distribution and vertical migration of meso- and bathypelagic species are lacking. Additional faunistic assessments must target those less-studied taxa and geographical locations. However, priority in ecological studies should be given to process-oriented investigations aimed at understanding the mechanisms controlling zooplankton distribution, trophic interactions within pelagic food webs and production cycles in relation to the physical environment. An effort should be made to incorporate state-of-the-art sampling technology and analytical methods into future research projects.

As pesquisas sobre o zooplâncton marinho no Brasil têm sido primariamente descritivas, com a maioria dos estudos enfocando a análise da estrutura da comunidade e assuntos relacionados. A composição e a distribuição espacial de muitos grupos taxonômicos encontram-se bem estudadas, embora os táxons menos abundantes e de menores dimensões, assimcomo os estágios iniciais do ciclo de vida da maioria das espécies, tenham recebido pouca atenção. Alguns táxons numericamenteimportantes encontram-se pouco estudados, como no caso dos protistas heterotróficos, ctenóforos, turbelários acelos e ostrácodes. Amplos setores da plataforma continental não têm sido suficientemente amostrados, em particular nas áreas influenciadas pela Corrente Norte do Brasil (5°N-15°S). As áreas oceânicas têm sido também pouco estudadas e praticamente inexistem dados sobre a distribuição espacial e vertical das espécies meso- e batipelágicas. Levantamentos faunísticos adicionais devem focalizar os táxons e locais menos conhecidos. No entanto, sob o ponto de vista ecológico é necessário darprioridade a estudos de processos voltados ao entendimentodos mecanismos que governam a distribuição, as interações tróficas nas teias alimentares pelágicas e os ciclos de produção do zooplâncton em relação ao ambiente físico. Deve ser feito um esforço para incorporar novas tecnologias de amostragem e métodos analíticos em futuros projetos de pesquisa.