Distribution, Community Composition, and Potential Metabolic Activity of Bacterioplankton in an Urbanized Mediterranean Sea Coastal Zone.

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 composition was mainly due to salinity and changes in environmental conditions across sites, as defined by nutrient and chlorophyll a concentrations. Bacterioplankton communities were composed of a few dominant taxa and a large proportion (92%) of rare taxa (here defined as operational taxonomic units [OTUs] accounting for <0.1% of the total sequence abundance), the majority of which were unique to each site. The relationship between 16S rRNA and the 16S rRNA gene, i.e., between potential metabolic activity and abundance, was positive for the whole community. However, analysis of individual OTUs revealed high rRNA-to-rRNA gene ratios for most (71.6% ± 16.7%) of the rare taxa, suggesting that these low-abundance organisms were potentially active and hence might be playing an important role in ecosystem diversity and functioning in the GON.IMPORTANCE The study of bacterioplankton in coastal zones is of critical importance, considering that these areas are highly productive and anthropogenically impacted. Their richness and evenness, as well as their potential activity, are very important to assess ecosystem health and functioning. Here, we investigated bacterial distribution, community composition, and potential metabolic activity in the GON, which is an ideal test site due to its heterogeneous environment characterized by a complex hydrodynamics and terrestrial inputs of varied quantities and quality. Our study demonstrates that bacterioplankton communities in this region are highly diverse and strongly regulated by a combination of different environmental factors leading to their heterogeneous distribution, with the rare taxa contributing to a major proportion of diversity and shifts in community composition and potentially holding a key role in ecosystem functioning.

miR-338-3p is over-expressed in blood, CFS, serum and spinal cord from sporadic amyotrophic lateral sclerosis patients.

Amyotrophic lateral sclerosis (ALS) is a progressive and seriously disabling adult-onset neurological disease. Ninety percent of ALS patients are sporadic cases (sALS) with no clear genetic linkage. Accumulating evidence indicates that various microRNAs (miRNAs), expressed in a spatially and temporally controlled manner in the brain, play a key role in neuronal development. In addition, microRNA dysregulation contributes to some mental disorders and neurodegeneration diseases. In our research, the expression of one selected miRNA, miR-338-3p, which previously we have found over-expressed in blood leukocytes, was studied in several different tissues from sALS patients. For the first time, we detected a specific microRNA disease-related upregulation, miR-338-3p, in blood leukocytes as well in cerebrospinal fluid, serum, and spinal cord from sALS patients. Besides, staining of in situ hybridization showed that the signals of miR-338-3p were localized in the grey matter of spinal cord tissues from sALS autopsied patients. We propose that miRNA profiles found in tissue samples from sALS patients can be relevant to understand sALS pathogenesis and lead to set up effective biomarkers for sALS early diagnosis.

Telomerase expression in amyotrophic lateral sclerosis (ALS) patients.

Telomerase and telomeric complex have been linked to a variety of disease states related to neurological dysfunction. In amyotrophic lateral sclerosis (ALS) patients, telomerase activity, as human telomerase reverse transcriptase (hTERT) expression, has not been characterized yet. Here, for the first time, we characterized telomerase and related pathway in blood sample and spinal cord from ALS patients compared with healthy controls. We found that hTERT expression level was significantly lower in ALS patients and was correlated either to p53 mRNA expression or p21 expression, pointing out the hypothesis that telomerase inhibition could be a pathogenetic contributor to neurodegeneration in ALS. As a consequence of the reduced telomerase activity, we identified shorter telomeres in leukocytes from sporadic ALS patients compared with healthy control group.

Ontogenetic profile of innate immune related genes and their tissue-specific expression in brown trout, Salmo trutta (Linnaeus, 1758).

The innate immune system is a fundamental defense weapon of fish, especially during early stages of development when acquired immunity is still far from being completely developed. The present study aims at looking into ontogeny of innate immune system in the brown trout, Salmo trutta, using RT-PCR based approach. Total RNA extracted from unfertilized and fertilized eggs and hatchlings at 0, 1 h and 1, 2, 3, 4, 5, 6, 7 weeks post-fertilization was subjected to RT-PCR using self-designed primers to amplify some innate immune relevant genes (TNF-α, IL-1ß, TGF-ß and lysozyme c-type). The constitutive expression of ß-actin was detected in all developmental stages. IL-1ß and TNF-α transcripts were detected from 4 week post-fertilization onwards, whereas TGF-ß transcript was detected only from 7 week post-fertilization onwards. Lysozyme c-type transcript was detected early from unfertilized egg stage onwards. Similarly, tissues such as muscle, ovary, heart, brain, gill, testis, liver, intestine, spleen, skin, posterior kidney, anterior kidney and blood collected from adult brown trout were subjected to detection of all selected genes by RT-PCR. TNF-α and lysozyme c-type transcripts were expressed in all tissues. IL-1ß and TGF-ß transcripts were expressed in all tissues except for the brain and liver, respectively. Taken together, our results show a spatial-temporal expression of some key innate immune-related genes, improving the basic knowledge of the function of innate immune system at early stage of brown trout.

Association between exposure to dioxin-like polychlorinated biphenyls and miR-191 expression in human peripheral blood mononuclear cells.

Accumulating evidence shows an association between deregulation of miRNAs and exposure to environmental chemicals; miRNAs play a unique regulatory role in gene expression. Among environmental pollutants, dioxins are a family of compounds that are known to have multiple hazardous effects. Also, in utero exposure of the fetus to dioxins has been shown to cause impaired psychomotor development, decreased immune function and skin disease. miR-191 is a microRNA that has been found to be up-regulated by dioxin in hepatocellular carcinoma cells in vitro. Our study provides the first molecular evidence in vivo of a positive relationship between levels of polychlorinated biphenyls (PCBs) and miR-191 expression in human peripheral blood mononuclear cells. miR-191 expression was significantly correlated with blood concentrations of total PCB and, in particular, of 3,3',4,4',5,5'-hexachlorobiphenyl (PCB 169, a coplanar congener). Blood concentrations of PCB 169 correlated significantly with miR-191 expression in pregnant women living in a PCB-polluted area, who underwent therapeutic abortion due to fetal malformations. These data suggest that miRNAs could be potential biomarkers to clarify the mechanisms of environmental disease.

Analysis of the P. lividus sea urchin genome highlights contrasting trends of genomic and regulatory evolution in deuterostomes.

Sea urchins are emblematic models in developmental biology and display several characteristics that set them apart from other deuterostomes. To uncover the genomic cues that may underlie these specificities, we generated a chromosome-scale genome assembly for the sea urchin Paracentrotus lividus and an extensive gene expression and epigenetic profiles of its embryonic development. We found that, unlike vertebrates, sea urchins retained ancestral chromosomal linkages but underwent very fast intrachromosomal gene order mixing. We identified a burst of gene duplication in the echinoid lineage and showed that some of these expanded genes have been recruited in novel structures (water vascular system, Aristotle's lantern, and skeletogenic micromere lineage). Finally, we identified gene-regulatory modules conserved between sea urchins and chordates. Our results suggest that gene-regulatory networks controlling development can be conserved despite extensive gene order rearrangement.

TITF1 Screening in Human Congenital Diaphragmatic Hernia (CDH).

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 studies showed positive brown staining of fetal follicular thyroid epithelium, normal fetal trachea, and normal fetal lung bronchial epithelium. Fetal esophageal wall was immunohistochemically negative. Molecular genetic analysis showed complete identity between the sequences obtained and the Wild Type (WT) form of the gene in all cases. No mutation, insertion and/or deletion was detected. Although TITF1 is expressed in the human fetal lung and has been considered to have a role in the pathogenesis of PH in CDH, the results of our study do not support the hypothesis that TITF1 mutations play a key role in the etiopathogenesis of CDH.

First identification and characterization of detoxifying plastic-degrading DBP hydrolases in the marine diatom Cylindrotheca closterium.

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.

Microsatellite primers in the planktonic diatom Pseudo-nitzschia multistriata (Bacillariophyceae).

PREMISE OF THE STUDY: Seven microsatellite loci were characterized for the toxic diatom Pseudo-nitzschia multistriata Takano (Takano) to investigate intraspecific variability and estimate population genetic structure over blooms, seasons, and sexual and vegetative reproduction. METHODS AND RESULTS: Selected microsatellites consisted of di- and trinucleotide repeats in the core region, and showed four to twelve alleles per locus in strains of P. multistriata collected in the Gulf of Naples (Italy). Primer pairs were species-specific since they positively amplified against conspecific strains from Portugal and Spain but failed to generate PCR products from the diatoms Pseudo-nitzschia pseudodelicatissima (Hasle) Hasle and Leptocylindrus minimum Gran. CONCLUSIONS: The seven selected microsatellite markers will be useful in studying population dynamics of Pseudo-nitzschia multistriata in space and time.

Construction of an adult barnacle (Balanus amphitrite) cDNA library and selection of reference genes for quantitative RT-PCR studies.

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.

Selection and validation of a set of reliable reference genes for quantitative RT-PCR studies in the brain of the Cephalopod Mollusc Octopus vulgaris.

BACKGROUND: Quantitative real-time polymerase chain reaction (RT-qPCR) is valuable for studying the molecular events underlying physiological and behavioral phenomena. Normalization of real-time PCR data is critical for a reliable mRNA quantification. Here we identify reference genes to be utilized in RT-qPCR experiments to normalize and monitor the expression of target genes in the brain of the cephalopod mollusc Octopus vulgaris, an invertebrate. Such an approach is novel for this taxon and of advantage in future experiments given the complexity of the behavioral repertoire of this species when compared with its relatively simple neural organization. RESULTS: We chose 16S, and 18S rRNA, actB, EEF1A, tubA and ubi as candidate reference genes (housekeeping genes, HKG). The expression of 16S and 18S was highly variable and did not meet the requirements of candidate HKG. The expression of the other genes was almost stable and uniform among samples. We analyzed the expression of HKG into two different set of animals using tissues taken from the central nervous system (brain parts) and mantle (here considered as control tissue) by BestKeeper, geNorm and NormFinder. We found that HKG expressions differed considerably with respect to brain area and octopus samples in an HKG-specific manner. However, when the mantle is treated as control tissue and the entire central nervous system is considered, NormFinder revealed tubA and ubi as the most suitable HKG pair. These two genes were utilized to evaluate the relative expression of the genes FoxP, creb, dat and TH in O. vulgaris. CONCLUSION: We analyzed the expression profiles of some genes here identified for O. vulgaris by applying RT-qPCR analysis for the first time in cephalopods. We validated candidate reference genes and found the expression of ubi and tubA to be the most appropriate to evaluate the expression of target genes in the brain of different octopuses. Our results also underline the importance of choosing a proper normalization strategy when analyzing gene expression by qPCR taking into appropriate account the experimental setting and variability of the sample of animals (and tissues), thus providing a set of HGK which expression appears to be unaffected by the experimental factor(s).

SNPs and Hox gene mapping in Ciona intestinalis.

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.

De novo assembly of a transcriptome from the eggs and early embryos of Astropecten aranciacus.

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 through the stages from oocyte to early embryo, as judged by real-time PCR, the transcript encoding Mos, a negative regulator of mitotic cell cycle, was drastically reduced during the period of rapid cleavages. Molecular phylogenetic analysis using the homologous amino acid sequences of cytochrome oxidase subunit I from A. aranciacus and 30 other starfish species indicated that Paxillosida, to which A. aranciacus belongs, is not likely to be the most basal order in Asteroidea. Taken together, the first transcriptome we assembled in this species is expected to enable us to perform comparative studies and to design gene-specific molecular tools with which to tackle long-standing biological questions.

Tissue-specific transcriptomic profiling provides new insights into the reproductive ecology and biology of the iconic seagrass species Posidonia oceanica.

Seagrasses form extensive meadows in shallow coastal waters and are among the world's most productive ecosystems. Seagrasses can produce both clonally and sexually, and flowering has long been considered infrequent, but important for maintaining genetically diverse stands. Here we investigate the molecular mechanisms involved in flowering of the seagrass Posidonia oceanica, an iconic species endemic to the Mediterranean. We generated a de novo transcriptome of this non-model species for leaf, male and female flower tissue of three individuals, and present molecular evidence for genes that may be involved in the flowering process and on the reproductive biology of the species. We present evidence that suggests that P. oceanica exhibits a strategy of protogyny, where the female part of the hermaphroditic flower develops before the male part, in order to avoid self-fertilization. We found photosynthetic genes to be up-regulated in the female flower tissues, indicating that this may be capable of photosynthesis. Finally, we detected a number of interesting genes, previously known to be involved in flowering pathways responding to light and temperature cues and in pathways involved in anthocyanin and exine synthesis. This first comparative transcriptomic approach of leaf, male and female tissue provides a basis for functional genomics research on flower development in P. oceanica and other seagrass species.

Toxicity of diatom polyunsaturated aldehydes to marine bacterial isolates reveals their mode of action.

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's bases, and therefore, they act as very toxic and effective poison, probably accumulating in cytoplasmic membranes because of their high hydrophobicity.

Multiplex PCR approach to simultaneously identify several mutations in fine needle cytology thyroid samples.

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 point mutations, and, in parallel, RT-PCR is used to detect the chimeric RET/PTC1 and RET/PTC3 transcripts in RNA extracted from FNA.

Seasonal heterogeneity of ocean warming: a mortality sink for ectotherm colonizers.

Distribution shifts are a common adaptive response of marine ectotherms to climate change but the pace of redistribution depends on species-specific traits that may promote or hamper expansion to northern habitats. Here we show that recently, the loggerhead turtle (Caretta caretta) has begun to nest steadily beyond the northern edge of the species' range in the Mediterranean basin. This range expansion is associated with a significant warming of spring and summer sea surface temperature (SST) that offers a wider thermal window suitable for nesting. However, we found that post-hatchlings departing from this location experience low winter SST that may affect their survival and thus hamper the stabilization of the site by self-recruitment. The inspection of the Intergovernmental Panel on Climate Change model projections and observational data on SST trends shows that, despite the annual warming for this century, winter SST show little or no trends. Therefore, thermal constraints during the early developmental phase may limit the chance of population growth at this location also in the near future, despite increasingly favourable conditions at the nesting sites. Quantifying and understanding the interplay between dispersal and environmental changes at all life stages is critical for predicting ectotherm range expansion with climate warming.

A Rapid and Cheap Methodology for CRISPR/Cas9 Zebrafish Mutant Screening.

The introduction of new genome editing tools such as ZFNs, TALENs and, more recently, the CRISPR/Cas9 system, has greatly expanded the ability to knock-out genes in different animal models, including zebrafish. However, time and costs required for the screening of a huge number of animals, aimed to identify first founder fishes (F0), and then carriers (F1) are still a bottleneck. Currently, high-resolution melting (HRM) analysis is the most efficient technology for large-scale InDels detection, but the very expensive equipment demanded for its application may represent a limitation for research laboratories. Here, we propose a rapid and cheap method for high-throughput genotyping that displays efficiency rate similar to the HRM. In fact, using a common ViiA™7 real-time PCR system and optimizing the parameters of the melting analysis, we demonstrated that it is possible to discriminate between the mutant and the wild type melting curves. Due to its simplicity, rapidity and cheapness, our method can be used as a preliminary one-step approach for massive screening, in order to restrict the scope at a limited number of embryos and to focus merely on them for the next sequencing step, necessary for the exact sequence identification of the induced mutation. Moreover, thanks to its versatility, this simple approach can be readily adapted to the detection of any kind of genome editing approach directed to genes or regulatory regions and can be applied to many other animal models.

A Bacillus sp. isolated from sediments of the Sarno River mouth, Gulf of Naples (Italy) produces a biofilm biosorbing Pb(II).

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) confirm that the biofilm contains Pb, suggesting an active biosorption of this metal by the bacterial cells, sequestering 14% of inoculated Pb as evidenced by microscopic analyses. Altogether, these observations support evidence that strain Pb15 has potentials for being used in bioremediation of its native polluted sediments, with engineering solutions to be found in order to eliminate the adsorbed Pb before replacement of sediments in situ.

Effect of bisphenol A on P-glycoprotein-mediated efflux and ultrastructure of the sea urchin embryo.

Usage of bisphenol A (BPA) in production of polycarbonate plastics has resulted in global distribution of BPA in the environment. These high concentrations cause numerous negative effects to the aquatic biota, among which the most known is the induction of endocrine disruption. The focus of this research was to determine the effects of two experimentally determined concentrations of BPA (100nM and 4µM) on cellular detoxification mechanisms during the embryonic development (2-cell, pluteus) of the rocky sea urchin (Paracentrotus lividus), primarily the potential involvement of multidrug efflux transport in the BPA intercellular efflux. The results of transport assay, measurements of the intracellular BPA and gene expression surveys, for the first time indicate the importance of P-glycoprotein (P-gp/ABCB1) in defense against BPA. Cytotoxic effects of BPA, validated by the immunohistochemistry (IHC) and the transmission electron microscopy (TEM), induced the aberrant karyokinesis, and consequently, the impairment of embryo development through the first cell division and retardation.