Marco M B Borra | Stazione Zoologica Anton Dohrn Napoli (original) (raw)

Papers by Marco M B Borra

Journal of Experimental Marine Biology and Ecology, Feb 1, 2014

ABSTRACT Diatoms are eukaryotic microalgae broadly present in freshwater and marine ecosystems. T... more ABSTRACT Diatoms are eukaryotic microalgae broadly present in freshwater and marine ecosystems. The great ecological interest about diatoms has recently led to increased efforts towards the exploration of the molecular properties of these organisms and the development of molecular tools to study the function of genes. We are using as model system two pennate diatoms, Pseudo-nitzschia multistriata and Pseudo-nitzschia arenysensis. In order to enable molecular investigations in these two species, we have made use of sequence information from transcriptomic data to identify genes that can be used as reference genes in quantitative PCR (qPCR). We have analyzed the expression of a set of genes commonly used as reference genes: histone H4, TBP (TATA binding protein), RPS (30 S ribosomal protein), GAPDH (glyceraldehyde 3-phosphate dehydrogenase), TUB A and TUB B (tubulin α and tubulin β), ACT (actin), CDK A (cyclin dependent kinase A) and COPA (coatomer protein complex, subunit α). The suitability of these genes as references in qPCR has been tested in different conditions and among different strains. We have found that only TUB A, TUB B and CDK A are stable in all the conditions analyzed. These three genes, in addition to ACT and COPA, are good reference genes for P. multistriata, while GAPDH appears to be differentially expressed in the tested conditions in this species. In P. arenysensis, instead, TUB A, TUB B, CDK A, GAPDH, H4 and RPS show the highest levels of stability and can be considered reliable reference genes.

BMC Genomics, 2008

Background: The tunicate Ciona intestinalis (Enterogona, Ascidiacea), a major model system for ev... more Background: The tunicate Ciona intestinalis (Enterogona, Ascidiacea), a major model system for evolutionary and developmental genetics of chordates, harbours two cryptic species. To assess the degree of intra-and inter-specific genetic variability, we report the identification and analysis of C. intestinalis SNP (Single Nucleotide Polymorphism) markers. A SNP subset was used to determine the genetic distance between Hox-5 and-10 genes. Results: DNA fragments were amplified from 12 regions of C. intestinalis sp. A. In total, 128 SNPs and 32 one bp indels have been identified within 8 Kb DNA. SNPs in coding regions cause 4 synonymous and 12 non-synonymous substitutions. The highest SNP frequency was detected in the Hox5 and Hox10 intragenic regions. In C. intestinalis, these two genes have lost their archetypal topology within the cluster, such that Hox10 is located between Hox4 and Hox5. A subset of the above primers was used to perform successful amplification in C. intestinalis sp. B. In this cryptic species, 62 SNPs were identified within 3614 bp: 41 in non-coding and 21 in coding regions. The genetic distance of the Hox-5 and-10 loci, computed combining a classical backcross approach with the application of SNP markers, was found to be 8.4 cM (Haldane's function). Based on the physical distance, 1 cM corresponds to 39.5 Kb. Linkage disequilibrium between the aforementioned loci was calculated in the backcross generation. Conclusion: SNPs here described allow analysis and comparisons within and between C. intestinalis cryptic species. We provide the first reliable computation of genetic distance in this important model chordate. This latter result represents an important platform for future studies on Hox genes showing deviations from the archetypal topology.

Frontiers in Marine Science, Jun 29, 2022

The Ocean Decade (OD) is the name of a United Nations (UN) initiative devoted to ocean science fo... more The Ocean Decade (OD) is the name of a United Nations (UN) initiative devoted to ocean science for sustainable development. It started in 2021 and will provide an opportunity to create a new foundation, across the science-policy interface, to strengthen the sustainable management of oceans and coasts and, at same time, bring benefits for future generations. The OD will establish a common strategy to achieve the objectives of the 2030 Agenda for Sustainable Development and the other European Union (EU) and international agreements, such as the EU Marine Strategy Framework Directive (MSFD), the Marine Biodiversity Strategy and the UN World Assessment I and II aimed at preserving ocean health. Furthermore, the OD has several expected outcomes that concern different aspects of marine environment, including its enormous values for humans. Several entities will combine efforts to achieve these goals and science is at the forefront of the sustainable blue economy. Marine research is solving complex challenges through interdisciplinary approaches, revolutionizing our life and our interaction with the ocean. This review discusses recent advancements in science related to the OD outcomes. The role of new technology for ocean exploration and monitoring, the importance of omics science and biotechnology to deal with ocean pollution, and other innovative solutions are discussed. All of these are inspired by the idea of using marine resources in a sustainable way and without impacting in a negative way on marine ecosystems. The role of science communication is therefore considered a crucial issue to spread the OD messages and to reach the general public and stakeholders. Only by the integration of science, governance, industry and public, the OD will have any chance to succeed.

Science of The Total Environment, Mar 1, 2022

Diatoms are photosynthetic organisms with potential biotechnological applications in the bioremed... more Diatoms are photosynthetic organisms with potential biotechnological applications in the bioremediation sector, having shown the capacity to reduce environmental concentrations of different pollutants. The diatom Cylindrotheca closterium is known to degrade di-n-butyl phthalate (DBP), one of the most abundant phthalate esters in aquatic environments and a known endocrine-disrupting chemical. In this study, we present for the first time the in silico identification of two putative DBP hydrolases (provisionally called DBPH1 and DBPH2) in the transcriptome of C. closterium. We modeled the structure of both DBPH1-2 and their proposed interactions with the substrate to gain insights into their mechanism of action. Finally, we analyzed the expression levels of the two putative hydrolases upon exposure of C. closterium to different concentrations of DBP (5 and 10 mg/L) for 24 and 48 h. The data showed a DBP concentration-dependent increase in expression levels of both dbph1 and 2 genes, further highlighting their potential involvement in phthalates degradation. This is the first identification of phthalate-degrading enzymes in microalgae, providing new insights into the possible use of diatoms in bioremediation strategies targeting phthalates.

Dottorato di Ricerca in Biologia Animale, XX Ciclo,a.a.2006-2007In this study, I searched and was... more Dottorato di Ricerca in Biologia Animale, XX Ciclo,a.a.2006-2007In this study, I searched and was able to identify FoxP in the transcriptome of the cephalopod mollusc Octopus vulgaris, an invertebrate. In addition, I attempted to analyze the expression of Ov-FoxP in the brain of this animal. The results of this analysis are preliminary at this stage. Fox proteins are a set of transcription factors highly conserved in metazoans. They are characterized by a typical DNA binding domain (Forkhead) that, among others, allows to identify 15 different classes of Fox genes. Fox proteins are reported to act as activators/repressors of transcription during both development (including differentiation) and the adult life (e.g. lung, brain, etc.). In vertebrates, FoxP2 (together with FoxP1), in particular, are known to be involved in the development of the neural circuit controlling bird-song and human speech. Our interest for the octopus derives from the fact that this animal, together with other cephalopods, is considered as the most evolved among molluscs. The complexity of the architecture and wiring of the cephalopod nervous system stems from the simpler nervous systems of other taxa belonging to the phylum. In addition, cephalopods show a highly rich behavioral repertoire including the unique capability of changing the appearance of their body (through body patterning) in fractions of seconds and for both mimetic and communicative purposes. Taken all together, these features allow these animals to be considered analogous to higher vertebrates. In the first part of my project, a detailed analysis of the aminoacidic and nucleotidic sequences available for FoxP2 (vertebrates) and FoxP (invertebrates), allowed us to design FoxP in Octopus vulgaris 1 appropriate oligos that were utilized in subsequent PCR experiments to identify the gene of interest in the transcriptome of the brain of O. vulgaris. FoxP resulted in a fragment of 220 bp that corresponded to the Forkhead domain. Further efforts allowed us to identify a 1111bp mRNA sequence of Ov-FoxP corresponding to almost the entire part of the mature mRNA codifying for this protein (the 5’ extremity of the gene results unidentified at this stage). During the second part of my project, I attempted to analyze the expression pattern of Ov- FoxP in the octopus brain using Real Time qPCR and in-situ hybridization. This was carried out with the aim of investigating the possible variability of expression of the gene in different parts of the brain (i.e. supra-, sub-esophageal masses and optic lobes) relative to another tissue (muscular tissue of the mantle) here considered as control. Other genes (16S, tubulin, actin) were also cloned for the aims of this project and their expression was taken as reference; an analysis that is carried out for the first time in O. vulgaris. By Real-Time qPCR I was able to recognize a different pattern of expression in different parts of the brain (N = 10). The data allowed to identify a gradient in the expression levels of FoxP (relative to reference genes) in the subesophageal mass, when the smallest individual of my sample (30 g body weight) was compared with the others (150-2100 g body weight). In situ hybridization (N=6) allowed to localize the expression of FoxP in the lobes of the octopus brain. Ov-FoxP transcripts were identified in neurons of: i. the optic lobes (several sparse cells possibly related with visual input processing); ii. the superior buccal and the lateral part of the basal lobes (high-order motor centers of the supraesophageal mass), and iii. the pedal tracts and anterior and posterior chromatophore lobes (subesophageal mass). FoxP in Octopus vulgaris 2 An elevated number of cells was revealed through in-situ hybridization in the last two lobes. It is noteworthy to mention that these structures are known to play a key role in the neural control of the chromatic expression of the skin of O. vulgaris (and other cephalopods): namely the animal’s body pattern. Our data seems to suggest that Ov-FoxP is expressed during different phases of the life of the octopus. In addition the localized expression in definite lobes and the variability among individuals of its expression in the same brain parts allows us to formulate the working hypothesis of the role of Ov-FoxP in the plasticity and/or maintainance of neural networks. My project in O. vulgaris confirms similar results deduced from other studies in both invertebrates (i.e. motor neurons in C. elegans) and vertebrates (i.e. song-birds, mouse, etc).Università della Calabri

BMC Molecular Biology, Jun 24, 2009

Background: Balanus amphitrite is a barnacle commonly used in biofouling research. Although many ... more Background: Balanus amphitrite is a barnacle commonly used in biofouling research. Although many aspects of its biology have been elucidated, the lack of genetic information is impeding a molecular understanding of its life cycle. As part of a wider multidisciplinary approach to reveal the biogenic cues influencing barnacle settlement and metamorphosis, we have sequenced and annotated the first cDNA library for B. amphitrite. We also present a systematic validation of potential reference genes for normalization of quantitative real-time PCR (qRT-PCR) data obtained from different developmental stages of this animal. Results: We generated a cDNA library containing expressed sequence tags (ESTs) from adult B. amphitrite. A total of 609 unique sequences (comprising 79 assembled clusters and 530 singlets) were derived from 905 reliable unidirectionally sequenced ESTs. Bioinformatics tools such as BLAST, HMMer and InterPro were employed to allow functional annotation of the ESTs. Based on these analyses, we selected 11 genes to study their ability to normalize qRT-PCR data. Total RNA extracted from 7 developmental stages was reverse transcribed and the expression stability of the selected genes was compared using geNorm, BestKeeper and NormFinder. These software programs produced highly comparable results, with the most stable gene being mt-cyb, while tuba, tubb and cp1 were clearly unsuitable for data normalization. Conclusion: The collection of B. amphitrite ESTs and their annotation has been made publically available representing an important resource for both basic and applied research on this species. We developed a qRT-PCR assay to determine the most reliable reference genes. Transcripts encoding cytochrome b and NADH dehydrogenase subunit 1 were expressed most stably, although other genes also performed well and could prove useful to normalize gene expression studies.

Environmental Science and Pollution Research, May 21, 2013

A multixenobiotic resistance (MXR) mechanism mediated by ABC binding cassette (ABC) transport pro... more A multixenobiotic resistance (MXR) mechanism mediated by ABC binding cassette (ABC) transport proteins is an efficient chemical defence mechanism in sea urchin embryos. The aim of our work was to evidence whether exposure to sub-lethal doses of specific contaminants (oxybenzone (OXI), mercuric chloride (HgCl 2) and trybutiltin (TBT)) would induce MXR transporter activity during embryonic development (from zygote to blastula stage) in purple sea urchin (Paracentrotus lividus) embryos. Further, we present data on molecular identification, transport function, expression levels and gene localisation of two ABC efflux transporters-P-glycoprotein (ABCB1/P-gp) and sulfonylurea-receptor-like protein (ABCC9/SUR-like). Partial cDNA sequences of abcb1 and abcc9 were identified and quantitative PCR (qPCR) evidenced an increase in mRNA transcript levels of both ABC transporters during the two-cell, as well as an overall decrease during the blastulae stage. Calcein-AM efflux activity assay indicated the activation of multidrug resistance-associated protein/ABCC-like transport in the presence of HgCl 2 and TBT in exposed blastulae. The in situ hybridisation of the two-cell and blastula stages showed ubiquitous localisation of both transcripts within cells, supporting qPCR data. In conclusion, ABCB1 and ABCC9 are constitutive, as are HgCl 2 , TBT and OXI-inducible ABC membrane transporters, coexpressed in the zygote, two-cell and blastula stages of the P. lividus. Their ubiquitous cell localisation further fortifies their protective role in early embryonic development. Keywords In situ hybridisation. P-glycoprotein (ABCB1/ P-gp). Sulfonylurea receptor like protein (ABCC9/SUR). Multixenobiotic resistance. qPCR. Sea urchin embryos. Paracentrotus lividus Background, aim and scope One important cellular defence of aquatic embryos against a wide range of toxic chemicals in the water is based on efflux transporter proteins belonging to the ATP-binding cassette (ABC) subfamily, including the ABCC/multidrug-resistance-associated proteins (MRPs) and the ABCB/P-glycoprotein (P-gp) (Kurelec 1992; Hamdoun et al. 2004; Epel et al. 2008). These ABC efflux transporters hydrolyse ATP and use the energy to actively transport various endogenous and exogenous toxic compounds across cell membranes against their concentration gradient (Higgins 1992; Dassa and Bouige 2001; Dean et al. 2001). In a clinical context, such ABC efflux transporter activity is associated with a multidrug resistance (MDR) phenotype of cancer cells, which shows resistance against a wide range of chemotherapeutic drugs (Cole et al. 1992; Leslie et al. 2005; Deeley et al. 2006). In the human genome, the members of ABCC subfamily responsible for the efflux of various compounds are multidrug resistance-associated proteins (MRPs/ABCClike members) (Borst et al. 2000; Dean et al. 2001).

Journal of Experimental Marine Biology and Ecology, Jul 1, 2011

In this study we develop gene expression tools in Calanus helgolandicus to study the effects of t... more In this study we develop gene expression tools in Calanus helgolandicus to study the effects of toxic diatom diets on copepod fitness and survival. We demonstrate that when adult females are fed on the control dinoflagellate Prorocentrum minimum and the flagellate Rhodomonas baltica, which are not known to produce toxic oxylipins, there are no significant changes in alpha-and beta-tubulin (microtubule subunits) gene expression levels. By contrast, the oxylipin-producing diatom Skeletonema marinoi influences tubulin expression levels which were markedly down-regulated. We scored a panel of putative reference genes (ACT, EFA, GAPDH, 18S, S7, S20, ATPs, UBI and IST) and found that two (S20 and S7) were highly stable in the tested conditions and can be used for further experiments. To normalize reverse transcription-quantitative real time polymerase chain reaction (RT-qPCR) data we also used the third best reference gene, GAPDH, considering the possibility that the two ribosomal proteins could be co-regulated. This pilot study will pave the way for further investigations on which genes are affected by diatom diets and clarify when and if a stress response or a detoxification mechanism becomes visible in C. helgolandicus. Moreover, the RT-qPCR analysis presented here may also be useful to study the effects of other diets and/or environmental factors such as salinity, temperature, pollution and other toxic compounds, on gene expression levels in this copepod species.

PLoS ONE, 2011

Background: Diatoms are dominant photosynthetic organisms in the world's oceans and are considere... more Background: Diatoms are dominant photosynthetic organisms in the world's oceans and are considered essential in the transfer of energy through marine food chains. However, these unicellular plants at times produce secondary metabolites such as polyunsaturated aldehydes and other products deriving from the oxidation of fatty acids that are collectively termed oxylipins. These cytotoxic compounds are responsible for growth inhibition and teratogenic activity, potentially sabotaging future generations of grazers by inducing poor recruitment in marine organisms such as crustacean copepods.

Journal of Experimental Marine Biology and Ecology, 2011

Children

TITF1 (Thyroid Transcription Factor-1) is a homeodomain-containing transcription factor. Previous... more TITF1 (Thyroid Transcription Factor-1) is a homeodomain-containing transcription factor. Previous studies showed that Titf1 null mice are characterized by failure of tracheo-oesophageal separation and impaired lung morphogenesis resulting in Pulmonary Hypoplasia (PH). In this study, we aim to evaluate the role of TITF1 in the pathogenesis of congenital diaphragmatic hernia (CDH) in humans. We investigated TITF1 expression in human trachea and lungs and performed direct mutation analysis in a CDH population. We studied 13 human fetuses at 14 to 24 weeks of gestation. Five μm sections were fixed in paraformaldehyde and incubated with anti-TITF1 primary antibody. Positive staining was visualized by biotinylated secondary antibody. We also performed TITF1 screening on genomic DNA extracted from peripheral blood of 16 patients affected by CDH and different degrees of PH, searching for mutations, insertions, and/or deletions, by sequencing the exonic regions of the gene. Histochemical stu...

Applied and Environmental Microbiology

Bacterioplankton are fundamental components of marine ecosystems and influence the entire biosphe... more Bacterioplankton are fundamental components of marine ecosystems and influence the entire biosphere by contributing to the global biogeochemical cycles of key elements. Yet, there is a significant gap in knowledge about their diversity and specific activities, as well as environmental factors that shape their community composition and function. Here, the distribution and diversity of surface bacterioplankton along the coastline of the Gulf of Naples (GON; Italy) were investigated using flow cytometry coupled with high-throughput sequencing of the 16S rRNA gene. Heterotrophic bacteria numerically dominated the bacterioplankton and comprised mainly Alphaproteobacteria , Gammaproteobacteria , and Bacteroidetes . Distinct communities occupied river-influenced, coastal, and offshore sites, as indicated by Bray-Curtis dissimilarity, distance metric (UniFrac), linear discriminant analysis effect size (LEfSe), and multivariate analyses. The heterogeneity in diversity and community compositi...

PloS one, 2017

Starfish have been instrumental in many fields of biological and ecological research. Oocytes of ... more Starfish have been instrumental in many fields of biological and ecological research. Oocytes of Astropecten aranciacus, a common species native to the Mediterranean Sea and the East Atlantic, have long been used as an experimental model to study meiotic maturation, fertilization, intracellular Ca2+ signaling, and cell cycle controls. However, investigation of the underlying molecular mechanisms has often been hampered by the overall lack of DNA or protein sequences for the species. In this study, we have assembled a transcriptome for this species from the oocytes, eggs, zygotes, and early embryos, which are known to have the highest RNA sequence complexity. Annotation of the transcriptome identified over 32,000 transcripts including the ones that encode 13 distinct cyclins and as many cyclin-dependent kinases (CDK), as well as the expected components of intracellular Ca2+ signaling toolkit. Although the mRNAs of cyclin and CDK families did not undergo significant abundance changes ...

Oncotarget, Jan 25, 2017

The most frequent initial manifestation of thyroid cancer is the appearance of a nodule. More tha... more The most frequent initial manifestation of thyroid cancer is the appearance of a nodule. More than 20% of the general population has a palpable thyroid nodule and the percentage rises to 70% based on ultrasound identification. In 95% of cases the nodule is simply a hyperplastic or benign lesion. The most reliable diagnostic test for thyroid nodules is fine needle aspiration (FNA), but cytological discrimination between malignant and benign follicular neoplasms remains difficult. Cytological analysis is now, almost routinely, being combined with molecular genetics to enable the pathologist to make a more objective diagnosis. In this study, we performed the molecular analysis using a new simplified procedure that involves a panel of BRAF, RAS, RET and RET/PTC gene mutations in easily obtainable FNA samples, in the attempt to improve the efficacy of the FNA diagnosis of thyroid nodules and thus patient management. In this new procedure, PCR and sequencing analysis are used to detect po...

Chemosphere, 2017

Diatoms produce and release polyunsaturated aldehydes (PUAs) during senescence in culture and at ... more Diatoms produce and release polyunsaturated aldehydes (PUAs) during senescence in culture and at the end of blooms in nature and these compounds play different ecological roles, as infochemicals, allelochemicals and pheromones In order to elucidate the toxic effects of PUAs, we isolated six bacterial strains from the Mediterranean Sea during a diatom bloom and tested their tolerance to PUA in terms of growth and cell membrane properties. Based upon 16S rRNA sequencing, these bacteria were assigned to the genera Pseudomonas, Sufflavibacter, Halomonas, Vibrio, Idiomarina, and Labrenzia. Growth of these strains was reduced by 50% (EC50) at PUA concentrations ranging from 600 to 1700 μM of 2E,4E/Z-heptadienal (HEPTA), 400-800 μM of 2E, 4E/Z-octadienal (OCTA), and 70-400 μM of 2E, 4E/Z-decadienal (DECA). Two of these strains, Vibrio sp. and Halomonas, sp. were also investigated for membrane fatty acid composition in terms of adaptive modifications of their degree of saturation (ratio between saturated and unsaturated fatty acids) by GC-FID. A direct correlation between hydrophobicity and PUA toxicity was observed, and these bacteria were also found to react to PUAs by increasing the degree of saturation of their membranes fatty acids. Tested PUAs were 4-fold more toxic than the well-investigated n-alkanols, most probably due to their additional chemical aldehyde toxicity to disrupting proteins by the formation of Schiff&amp;amp;amp;amp;amp;amp;amp;#39;s bases, and therefore, they act as very toxic and effective poison, probably accumulating in cytoplasmic membranes because of their high hydrophobicity.

The Science of the total environment, Aug 15, 2016

A Pb-resistant bacterial strain (named hereinafter Pb15) has been isolated from highly polluted m... more A Pb-resistant bacterial strain (named hereinafter Pb15) has been isolated from highly polluted marine sediments at the Sarno River mouth, Italy, using an enrichment culture to which Pb(II) 0.48mmoll(-1) were added. 16S rRNA gene sequencing (Sanger) allowed assignment of the isolate to the genus Bacillus, with Bacillus pumilus as the closest species. The isolate is resistant to Pb(II) with a minimum inhibitory concentration (MIC) of 4.8mmoll(-1) and is also resistant to Cd(II) and Mn(II) with MIC of 2.22mmoll(-1) and 18.20mmoll(-1), respectively. Inductively coupled plasma atomic emission spectrometry (ICP-AES) showed that Pb inoculated in the growth medium is absorbed by the bacterial cells at removal efficiencies of 31.02% and 28.21% in the presence of 0.48mmoll(-1) or 1.20mmoll(-1) Pb(II), respectively. Strain Pb15 forms a brown and compact biofilm when grown in presence of Pb(II). Scanning Electron Microscopy (SEM) coupled with Energy Dispersive X-ray Spectroscopy (SEM-EDS) conf...