<p>Blue, green and red fluence rates (µmol photon m<sup>−2</sup> s<sup>−1... more <p>Blue, green and red fluence rates (µmol photon m<sup>−2</sup> s<sup>−1</sup>) measured at light peak and red : blue ratio values for the different light conditions. <i>a</i>*×10<sup>–11</sup>, absorption coefficient (m<sup>2</sup> cell<sup>−1</sup>); PUR×10<sup>–6</sup>, photosynthetically usable radiation (µW cell<sup>–1</sup>); <sub>rel</sub>ETR<sub>max</sub>×10<sup>–6</sup>, (maximal relative rate of linear electron transport, nmol e<sup>−1</sup> s<sup>−1</sup> cell<sup>−1</sup>), α×10<sup>–9</sup> (maximum light use efficiency, nmol e<sup>−1</sup> s<sup>−1</sup> cell<sup>−1</sup>(µmol photon m<sup>−2</sup> s<sup>−1</sup>)<sup> −1</sup>), and E<i>k</i> (light intensity for reaching <sub>rel</sub>ETR<sub>max</sub>, µmol photon m<sup>−2</sup> s<sup>−1</sup>); POC, particulate organic carbon (pg cell<sup>−1</sup>); POC/PON, particulate organic carbon (POC) to particulate organic nitrogen (PON) ratio (pg/pg); Chl <i>a</i>/POC×10<sup>–3</sup>, Chlorophyll <i>a</i> to POC ratio (pg/pg). Data represent mean and standard deviation. For <i>a</i>* and PUR, <i>n</i> = 3; For <sub>rel</sub>ETR<sub>max</sub>, α and E<i>k</i>, <i>n</i> = 6 (mean of the two days light peak measurements); For POC, PON, POC/PON and Chl <i>a</i>/POC, <i>n</i> = 21.</p
The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species... more The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species that can produce domoic acid, a potent neurotoxin known to cause amnesic shellfish poisoning (ASP). An early identification and an adequate monitoring of the potential toxic Pseudo-nitzschia spp. are necessary. However, effective monitoring programs are time consuming due, in some cases, to the cell morphology similarities among species, determined with light microscopy, that can result in insufficient data to give a definitive species and toxins attribution. In this paper, Whole-Cell Fluorescent In Situ Hybridization (WC-FISH) has been evaluated as a powerful tool to detect and enumerate harmful cryptic and/or pseudo-cryptic Pseudo-nitzschia spp. collected in the Gulf of Naples. Fluorescently labelled probes directed against the ribosomal RNA (rRNA) of the 28S large subunit (LSU) were used. In particular, five probes detecting four cryptic species of Pseudo-nitzschia delicatissima complex and one specific for Pseudo-nitzschia multistriata gave good results for the molecular identification of potentially toxic target species in natural samples. Finally, we can state that the WC-FISH method, to identify Pseudo-nitzschia species, is faster and more cost-effective if compared with other rDNA-based methods.
Heavy metals (HMs) can induce both chronic and acute harmful effects on marine and freshwater bio... more Heavy metals (HMs) can induce both chronic and acute harmful effects on marine and freshwater biota. The environmental impact of HMs in freshwater, seawater, soil, and wastewater can be limited using microbes, including microalgae, that are able to remove metals from environmental matrices. Indeed, they can passively adsorb and actively accumulate these persistent pollutants within their organelles, limiting their detrimental effects on cellular metabolism. The Sarno River is a 30 km long freshwater stream located in Southern Italy, polluted by partially untreated municipal, agricultural, and industrial wastewaters. In spite of this, microalgal cultures from Sarno River or Sarno River Mouth have never been established. In the present study, we isolated a green algal strain from the Sarno River Mouth and determined its ability to grow in polluted seawater containing different concentrations of cadmium, lead, or zinc. This strain was found to be able to accumulate these elements withi...
Microalgae are increasingly recognised as suitable microorganisms for heavy metal (HM) removal, s... more Microalgae are increasingly recognised as suitable microorganisms for heavy metal (HM) removal, since they are able to adsorb them onto their cell wall and, in some cases, compartmentalise them inside organelles. However, at relatively high HM concentrations, they could also show signs of stress, such as organelle impairments and increased activities of antioxidant enzymes. The main aim of this review is to report on the mechanisms adopted by microalgae to counteract detrimental effects of high copper (Cu) concentrations, and on the microalgal potential for Cu bioremediation of aquatic environments. Studying the delicate balance between beneficial and detrimental effects of Cu on microalgae is of particular relevance as this metal is widely present in aquatic environments facing industrial discharges. This metal often induces chloroplast functioning impairment, generation of reactive oxygen species (ROS) and growth rate reduction in a dose-dependent manner. However, microalgae also ...
The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species... more The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species. Some of these can produce domoic acid, a potent neurotoxin. Thus, monitoring programs are needed to screen for the presence of these toxic species. Unfortunately, many are impossible to distinguish using light microscopy. Therefore, we assessed the applicability of microarray technology for detection of toxic and non-toxic Pseudo-nitzschia species in the Gulf of Naples (Mediterranean Sea). Here, 11 species have been detected, of which at least 5 are potentially toxic. A total of 49 genus-and species-specific DNA probes were designed in silico against the nuclear LSU and SSU rRNA of 19 species, and spotted on the microarray. The microarray was tested against total RNA of monoclonal cultures of eight species. Only three of the probes designed to be species-specific were indeed so within the limits of our experimental design. To assess the effectiveness of the microarray in detecting Pseu...
One of the key tasks in the project MIDTAL (MIcroarrays for the Detection of Toxic ALgae) is to d... more One of the key tasks in the project MIDTAL (MIcroarrays for the Detection of Toxic ALgae) is to demonstrate the applicability of microarrays to monitor harmful algae across a broad range of ecological niches and toxic species responsible for harmful algal events. Water samples are collected from a series of sites used in national phytoplankton and biotoxin monitoring programmes across Europe. The samples are filtered; the rRNA is extracted, labelled with a fluorescent dye and applied to a microarray chip. The signal intensity from >120 probes previously spotted on the chip is measured and analysed. Preliminary results comparing microarray signal intensities with actual field counts are presented.
Isolation of nuclei tagged in specific cell types (INTACT) is a method developed to isolate cell-... more Isolation of nuclei tagged in specific cell types (INTACT) is a method developed to isolate cell-type-specific nuclei that are tagged through in vivo biotin labeling of a nuclear targeting fusion (NTF) protein. In our work, INTACT was used to capture nuclei of meiocytes and to generate a meiotic transcriptome in Arabidopsis. Using the promoter of AtDMC1 recombinase to label meiotic nuclei, we generated transgenic plants carrying AtDMC1:NTF along with biotin ligase enzyme (BirA) under the constitutive ACTIN2 (ACT2) promoter. AtDMC1-driven expression of biotin-labeled NTF allowed us to collect nuclei of meiocytes by streptavidin-coated magnetic beads. The nuclear meiotic transcriptome was obtained by RNA-seq using low-quantity input RNA. Transcripts grouped into different categories according to their expression levels were investigated by gene ontology enrichment analysis (GOEA). The most enriched GO term “DNA demethylation” in mid/high-expression classes suggests that this biologica...
Diatoms are unicellular eukaryotic microalgae widely distributed in aquatic environments, possess... more Diatoms are unicellular eukaryotic microalgae widely distributed in aquatic environments, possessing a porous silica cell wall known as frustule. Diatom frustules are considered as a sustainable source for several industrial applications because of their high biocompatibility and the easiness of surface functionalisation, which make frustules suitable for regenerative medicine and as drug carriers. Frustules are made of hydrated silica, and can be extracted and purified both from living and fossil diatoms using acid treatments or high temperatures. Biosilica frustules have proved to be suitable for biomedical applications, but, unfortunately, they are not officially recognised as safe by governmental food and medical agencies yet. In the present review, we highlight the frustule formation process, the most common purification techniques, as well as advantages and bottlenecks related to the employment of diatom-derived silica for medical purposes, suggesting possible solutions for a ...
The biochemical profile and growth of the coastal diatom Skeletonema marinoi was investigated und... more The biochemical profile and growth of the coastal diatom Skeletonema marinoi was investigated under four different daily blue light doses (sinusoidal light peaking at 88, 130, 250 and 450μmol photons m(-2)s(-1), respectively). Ability of cells to regulate the light energy input caused alterations in growth and different biosynthetic pathways. The light saturation index for photosynthesis (Ek), which governs the photoacclimative processes, ranged between 250 and 300μmol photons m(-2)s(-1). Cells that were adapted to low light (&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;Ek) enhanced their carotenoid, lipid and protein contents and lowered carbohydrate content, and vice versa under high light (≥Ek). Variations in fatty acid, pigment and amino acid compositions were a result of light adaptation. Our data show that light is a potent factor for manipulating biomass synthesis in microalgae, such as diatoms for microalgal biotechnology.
ABSTRACT In diatoms, as in other organisms, many genetically distinct and reproductively isolated... more ABSTRACT In diatoms, as in other organisms, many genetically distinct and reproductively isolated species may show identical or highly similar morphological features. Such groups of species are defined as cryptic and pseudo-cryptic species, respectively. The difficulty of discriminating them with optical means impairs the study of their temporal patterns and geographic ranges. This is also the case for Pseudo-nitzschia, a worldwide distributed planktonic diatom genus which includes several toxigenic species. Using a Pseudo-nitzschia-specific pair of large sub-units ribosomal DNA (LSU rDNA) primers, we generated clone libraries from 19 samples collected at the Long Term Ecological Station MareChiara (LTER-MC) in the Gulf of Naples (GoN) from 2009 to 2010 and compared sequence records with light microscopy (LM) counts from the same samples. Our aim was to elucidate the diversity and the seasonal patterns of taxa within Pseudo-nitzschia. Most of the Pseudo-nitzschia species already known from the GoN were identified within the 1643 obtained sequences. In addition, two species known from elsewhere and three un-described ribotypes were detected. Several cryptic species showed distinct temporal patterns of occurrence, with most species confined to restricted periods and only a few present year-round. Microscopic and molecular results generally concurred for species recognizable using LM, while clone libraries tended to overestimate the relative abundance of some of the species. Due to its high resolution and detection power, the DNA-barcoding approach used in our study is an optimal tool to trace the distribution of cryptic and toxigenic Pseudo-nitzschia species and the diversity of this key diatom genus in the natural environment.
<p>Blue, green and red fluence rates (µmol photon m<sup>−2</sup> s<sup>−1... more <p>Blue, green and red fluence rates (µmol photon m<sup>−2</sup> s<sup>−1</sup>) measured at light peak and red : blue ratio values for the different light conditions. <i>a</i>*×10<sup>–11</sup>, absorption coefficient (m<sup>2</sup> cell<sup>−1</sup>); PUR×10<sup>–6</sup>, photosynthetically usable radiation (µW cell<sup>–1</sup>); <sub>rel</sub>ETR<sub>max</sub>×10<sup>–6</sup>, (maximal relative rate of linear electron transport, nmol e<sup>−1</sup> s<sup>−1</sup> cell<sup>−1</sup>), α×10<sup>–9</sup> (maximum light use efficiency, nmol e<sup>−1</sup> s<sup>−1</sup> cell<sup>−1</sup>(µmol photon m<sup>−2</sup> s<sup>−1</sup>)<sup> −1</sup>), and E<i>k</i> (light intensity for reaching <sub>rel</sub>ETR<sub>max</sub>, µmol photon m<sup>−2</sup> s<sup>−1</sup>); POC, particulate organic carbon (pg cell<sup>−1</sup>); POC/PON, particulate organic carbon (POC) to particulate organic nitrogen (PON) ratio (pg/pg); Chl <i>a</i>/POC×10<sup>–3</sup>, Chlorophyll <i>a</i> to POC ratio (pg/pg). Data represent mean and standard deviation. For <i>a</i>* and PUR, <i>n</i> = 3; For <sub>rel</sub>ETR<sub>max</sub>, α and E<i>k</i>, <i>n</i> = 6 (mean of the two days light peak measurements); For POC, PON, POC/PON and Chl <i>a</i>/POC, <i>n</i> = 21.</p
The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species... more The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species that can produce domoic acid, a potent neurotoxin known to cause amnesic shellfish poisoning (ASP). An early identification and an adequate monitoring of the potential toxic Pseudo-nitzschia spp. are necessary. However, effective monitoring programs are time consuming due, in some cases, to the cell morphology similarities among species, determined with light microscopy, that can result in insufficient data to give a definitive species and toxins attribution. In this paper, Whole-Cell Fluorescent In Situ Hybridization (WC-FISH) has been evaluated as a powerful tool to detect and enumerate harmful cryptic and/or pseudo-cryptic Pseudo-nitzschia spp. collected in the Gulf of Naples. Fluorescently labelled probes directed against the ribosomal RNA (rRNA) of the 28S large subunit (LSU) were used. In particular, five probes detecting four cryptic species of Pseudo-nitzschia delicatissima complex and one specific for Pseudo-nitzschia multistriata gave good results for the molecular identification of potentially toxic target species in natural samples. Finally, we can state that the WC-FISH method, to identify Pseudo-nitzschia species, is faster and more cost-effective if compared with other rDNA-based methods.
Heavy metals (HMs) can induce both chronic and acute harmful effects on marine and freshwater bio... more Heavy metals (HMs) can induce both chronic and acute harmful effects on marine and freshwater biota. The environmental impact of HMs in freshwater, seawater, soil, and wastewater can be limited using microbes, including microalgae, that are able to remove metals from environmental matrices. Indeed, they can passively adsorb and actively accumulate these persistent pollutants within their organelles, limiting their detrimental effects on cellular metabolism. The Sarno River is a 30 km long freshwater stream located in Southern Italy, polluted by partially untreated municipal, agricultural, and industrial wastewaters. In spite of this, microalgal cultures from Sarno River or Sarno River Mouth have never been established. In the present study, we isolated a green algal strain from the Sarno River Mouth and determined its ability to grow in polluted seawater containing different concentrations of cadmium, lead, or zinc. This strain was found to be able to accumulate these elements withi...
Microalgae are increasingly recognised as suitable microorganisms for heavy metal (HM) removal, s... more Microalgae are increasingly recognised as suitable microorganisms for heavy metal (HM) removal, since they are able to adsorb them onto their cell wall and, in some cases, compartmentalise them inside organelles. However, at relatively high HM concentrations, they could also show signs of stress, such as organelle impairments and increased activities of antioxidant enzymes. The main aim of this review is to report on the mechanisms adopted by microalgae to counteract detrimental effects of high copper (Cu) concentrations, and on the microalgal potential for Cu bioremediation of aquatic environments. Studying the delicate balance between beneficial and detrimental effects of Cu on microalgae is of particular relevance as this metal is widely present in aquatic environments facing industrial discharges. This metal often induces chloroplast functioning impairment, generation of reactive oxygen species (ROS) and growth rate reduction in a dose-dependent manner. However, microalgae also ...
The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species... more The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species. Some of these can produce domoic acid, a potent neurotoxin. Thus, monitoring programs are needed to screen for the presence of these toxic species. Unfortunately, many are impossible to distinguish using light microscopy. Therefore, we assessed the applicability of microarray technology for detection of toxic and non-toxic Pseudo-nitzschia species in the Gulf of Naples (Mediterranean Sea). Here, 11 species have been detected, of which at least 5 are potentially toxic. A total of 49 genus-and species-specific DNA probes were designed in silico against the nuclear LSU and SSU rRNA of 19 species, and spotted on the microarray. The microarray was tested against total RNA of monoclonal cultures of eight species. Only three of the probes designed to be species-specific were indeed so within the limits of our experimental design. To assess the effectiveness of the microarray in detecting Pseu...
One of the key tasks in the project MIDTAL (MIcroarrays for the Detection of Toxic ALgae) is to d... more One of the key tasks in the project MIDTAL (MIcroarrays for the Detection of Toxic ALgae) is to demonstrate the applicability of microarrays to monitor harmful algae across a broad range of ecological niches and toxic species responsible for harmful algal events. Water samples are collected from a series of sites used in national phytoplankton and biotoxin monitoring programmes across Europe. The samples are filtered; the rRNA is extracted, labelled with a fluorescent dye and applied to a microarray chip. The signal intensity from >120 probes previously spotted on the chip is measured and analysed. Preliminary results comparing microarray signal intensities with actual field counts are presented.
Isolation of nuclei tagged in specific cell types (INTACT) is a method developed to isolate cell-... more Isolation of nuclei tagged in specific cell types (INTACT) is a method developed to isolate cell-type-specific nuclei that are tagged through in vivo biotin labeling of a nuclear targeting fusion (NTF) protein. In our work, INTACT was used to capture nuclei of meiocytes and to generate a meiotic transcriptome in Arabidopsis. Using the promoter of AtDMC1 recombinase to label meiotic nuclei, we generated transgenic plants carrying AtDMC1:NTF along with biotin ligase enzyme (BirA) under the constitutive ACTIN2 (ACT2) promoter. AtDMC1-driven expression of biotin-labeled NTF allowed us to collect nuclei of meiocytes by streptavidin-coated magnetic beads. The nuclear meiotic transcriptome was obtained by RNA-seq using low-quantity input RNA. Transcripts grouped into different categories according to their expression levels were investigated by gene ontology enrichment analysis (GOEA). The most enriched GO term “DNA demethylation” in mid/high-expression classes suggests that this biologica...
Diatoms are unicellular eukaryotic microalgae widely distributed in aquatic environments, possess... more Diatoms are unicellular eukaryotic microalgae widely distributed in aquatic environments, possessing a porous silica cell wall known as frustule. Diatom frustules are considered as a sustainable source for several industrial applications because of their high biocompatibility and the easiness of surface functionalisation, which make frustules suitable for regenerative medicine and as drug carriers. Frustules are made of hydrated silica, and can be extracted and purified both from living and fossil diatoms using acid treatments or high temperatures. Biosilica frustules have proved to be suitable for biomedical applications, but, unfortunately, they are not officially recognised as safe by governmental food and medical agencies yet. In the present review, we highlight the frustule formation process, the most common purification techniques, as well as advantages and bottlenecks related to the employment of diatom-derived silica for medical purposes, suggesting possible solutions for a ...
The biochemical profile and growth of the coastal diatom Skeletonema marinoi was investigated und... more The biochemical profile and growth of the coastal diatom Skeletonema marinoi was investigated under four different daily blue light doses (sinusoidal light peaking at 88, 130, 250 and 450μmol photons m(-2)s(-1), respectively). Ability of cells to regulate the light energy input caused alterations in growth and different biosynthetic pathways. The light saturation index for photosynthesis (Ek), which governs the photoacclimative processes, ranged between 250 and 300μmol photons m(-2)s(-1). Cells that were adapted to low light (&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;Ek) enhanced their carotenoid, lipid and protein contents and lowered carbohydrate content, and vice versa under high light (≥Ek). Variations in fatty acid, pigment and amino acid compositions were a result of light adaptation. Our data show that light is a potent factor for manipulating biomass synthesis in microalgae, such as diatoms for microalgal biotechnology.
ABSTRACT In diatoms, as in other organisms, many genetically distinct and reproductively isolated... more ABSTRACT In diatoms, as in other organisms, many genetically distinct and reproductively isolated species may show identical or highly similar morphological features. Such groups of species are defined as cryptic and pseudo-cryptic species, respectively. The difficulty of discriminating them with optical means impairs the study of their temporal patterns and geographic ranges. This is also the case for Pseudo-nitzschia, a worldwide distributed planktonic diatom genus which includes several toxigenic species. Using a Pseudo-nitzschia-specific pair of large sub-units ribosomal DNA (LSU rDNA) primers, we generated clone libraries from 19 samples collected at the Long Term Ecological Station MareChiara (LTER-MC) in the Gulf of Naples (GoN) from 2009 to 2010 and compared sequence records with light microscopy (LM) counts from the same samples. Our aim was to elucidate the diversity and the seasonal patterns of taxa within Pseudo-nitzschia. Most of the Pseudo-nitzschia species already known from the GoN were identified within the 1643 obtained sequences. In addition, two species known from elsewhere and three un-described ribotypes were detected. Several cryptic species showed distinct temporal patterns of occurrence, with most species confined to restricted periods and only a few present year-round. Microscopic and molecular results generally concurred for species recognizable using LM, while clone libraries tended to overestimate the relative abundance of some of the species. Due to its high resolution and detection power, the DNA-barcoding approach used in our study is an optimal tool to trace the distribution of cryptic and toxigenic Pseudo-nitzschia species and the diversity of this key diatom genus in the natural environment.
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