Natural Biofilms Research Articles

Assessing ecological status with diatoms DNA metabarcoding: Scaling-up on a WFD monitoring network (Mayotte island, France)

Diatoms are excellent ecological indicators of water quality because they are broadly distributed, they show high species diversity and they respond rapidly to human pressures. In Europe, the Water Framework Directive (WFD) gives the legal basis for the use of this indicator for water quality assessment and its management. Several quality indices, like the Specific Polluosensitivity Index (SPI), were developed to assess the ecological quality status of rivers based on diatom communities. It is based on morphological identifications and count of diatom species present in natural biofilms using a microscope. This methodology requires high taxonomic skills and several hours of analysis per sample as 400 individuals must be identified to species level. Since several years, a molecular approach based on DNA metabarcoding combined to High-Throughput Sequencing (HTS) is developed to characterize species assemblages in environmental samples which is potentially faster and cheaper. The ability of this approach to provide reliable diatom inventories has been demonstrated and its application to water quality assessment is currently being improved. Despite optimization of the DNA metabarcoding process with diatoms, few studies had yet extended it at the scale of a freshwater monitoring network and evaluated the reliability of its quality assessment compared to the classical morphological approach.In the present study we applied DNA metabarcoding to the river monitoring network of the tropical Island Mayotte. This island is a French département since 2011 and the WFD has to be applied. This offered the opportunity to scale up the comparison of molecular and morphological approaches and their ability to produce comparable community inventories and water quality assessments. Benthic diatoms were sampled following WFD standards in 45 river sites in 2014 and 2015 (80 samples). All samples were submitted in parallel to the molecular and the morphological approaches. DNA metabarcoding was carried out using Genelute DNA extraction method, rbcL DNA barcode and PGM sequencing, while microscopic counts were carried out for the classical methodology. Diatom community structures in terms of molecular (OTUs) and of morphological (species) were significantly correlated. However, only 13% of the species was shared by both approaches, with qualitative and quantitative variation due to i) the incompleteness of the reference library (82% of morphological species are not represented in the database), ii) limits in taxonomic knowledge and iii) biases in the estimation of relative abundances linked to diatom cell biovolume. However, ecological quality status assessed with the molecular and morphological SPI values were congruent, and little affected by sequencing depth. DNA metabarcoding of diatom communities allowed a reliable estimation of the quality status for most of the rivers at the scale of the full biomonitoring network of Mayotte Island.

Controlling growth and colour of phototrophs by using simple and inexpensive coloured lighting: A preliminary study in the Light4Heritage project towards future strategies for outdoor illumination

The quantity (duration and intensity) and quality (colour or wavelength) of light are key factors involved in regulating growth and physiological processes in photosynthetic organisms. As a preliminary approach to developing biocolonization control strategies based on urban ornamental lighting, this study aimed to evaluate how coloured cellophane film affects the quality and intensity of light and, therefore, the growth and colour of photosynthetic organisms. We used red, yellow, green and blue cellophane film (rather than coloured LEDs) to generate different light qualities (by cancelling the spectral components in certain bands of the visible electromagnetic spectrum) at different photon flux densities. We then used the coloured cellophane to cover phototrophic cultures, derived from natural biofilm growing on a historic granitic building and mainly comprising green algae and cyanobacteria, in order to promote specific physiological responses. The blue cellophane inhibited growth of the test culture. By contrast, the yellow cellophane did not significantly decrease the biomass, pigment or EPS content relative to uncovered, control cultures. The different coloured cellophane covers also generated colour changes in the culture; e.g. the red cellophane produced notable greening, whereas the green cellophane enhanced the redness of the culture.

Thermophilic Moorella thermoautotrophica-immobilized cathode enhanced microbial electrosynthesis of acetate and formate from CO2

Microbial electrosynthesis (MES) is a promising technique that converts electricity and CO2 to biofuels using microbes as the catalysts. However, most of previous MES are conducted at mesophilic temperatures and challenged by low performances. Here we report a significant electrosynthesis performance enhancement via immobilization of a thermophilic microbe to cathodes. A temperature-dependent electron uptake rate of Moorella thermoautotrophica was observed at a cathode potential of −0.4V (vs. SHE), with a maximum current density of 63.47mAm−2 at 55°C. Moreover, electrosynthesis rates of formate and acetate at 55°C were accelerated by 23.2 and 2.8 fold than those of 25°C, respectively. Compared with natural biofilms, immobilization of M. thermoautotrophica with carbon nanoparticles to electrodes further enhanced acetate and formate production rates (by 14 and 7.9 fold), reaching 58.2 and 63.2mmolm−2day−1 at a coulombic efficiency of 65%, respectively. To our best knowledge, these are the highest electrosynthesis rates obtained thus far for pure cultures under the conditions of −0.4V (vs. SHE) and 55°C. This study, for the first time, demonstrates that embedding microbes to electrodes by carbon nanoparticles is a facile and efficient method of improving MES performance.

Applications of microalgal biofilms for wastewater treatment and bioenergy production

BackgroundMicroalgae have shown clear advantages for the production of biofuels compared with energy crops. Apart from their high growth rates and substantial lipid/triacylglycerol yields, microalgae can grow in wastewaters (animal, municipal and mining wastewaters) efficiently removing their primary nutrients (C, N, and P), heavy metals and micropollutants, and they do not compete with crops for arable lands. However, fundamental barriers to the industrial application of microalgae for biofuel production still include high costs of removing the algae from the water and the water from the algae which can account for up to 30–40% of the total cost of biodiesel production. Algal biofilms are becoming increasingly popular as a strategy for the concentration of microalgae, making harvesting/dewatering easier and cheaper.ResultsWe have isolated and characterized a number of natural microalgal biofilms from freshwater, saline lakes and marine habitats. Structurally, these biofilms represent complex consortia of unicellular and multicellular, photosynthetic and heterotrophic inhabitants, such as cyanobacteria, microalgae, diatoms, bacteria, and fungi. Biofilm #52 was used as feedstock for bioenergy production. Dark fermentation of its biomass by Enterobacter cloacae DT-1 led to the production of 2.4 mol of H2/mol of reduced sugar. The levels and compositions of saturated, monosaturated and polyunsaturated fatty acids in Biofilm #52 were target-wise modified through the promotion of the growth of selected individual photosynthetic inhabitants. Photosynthetic components isolated from different biofilms were used for tailoring of novel biofilms designed for (i) treatment of specific types of wastewaters, such as reverse osmosis concentrate, (ii) compositions of total fatty acids with a new degree of unsaturation and (iii) bio-flocculation and concentration of commercial microalgal cells. Treatment of different types of wastewaters with biofilms showed a reduction in the concentrations of key nutrients, such as phosphates, ammonia, nitrates, selenium and heavy metals.ConclusionsThis multidisciplinary study showed the new potential of natural biofilms, their individual photosynthetic inhabitants and assembled new algal/cyanobacterial biofilms as the next generation of bioenergy feedstocks which can grow using wastewaters as a cheap source of key nutrients.

Quantification of the Metabolic Activities of Natural Biofilm of Different Microenvironments

Quantification of the Metabolic Activities of Natural Biofilm of Different Microenvironments

Changes in copper toxicity towards diatom communities with experimental warming

Biological communities in aquatic environments most commonly face multiple stress, where natural and anthropogenic stressors often act jointly. Their interactions are most easily assessed using short cycle organisms such as periphytic diatoms.In this experiment, we analyzed the combined effects of copper exposure and warming on diatom successions over 6 weeks. Natural biofilm collected in winter was left to grow in mesocosms exposed or unexposed to realistic Cu concentrations at four different temperatures. Separate and joint impacts of the two stressors were determined through structural and functional endpoints.Both temperature and copper influenced the biological responses; their interaction, when significant, was always antagonistic. Diatom communities gradually changed with rising temperature. Under copper exposure, the dominant Planothidium lanceolatum was superseded by Achnanthidium exiguum, which accounted for about 70% relative abundance in the warmest conditions (18–23°C). Tolerance to copper was derived from dose-response curves based on photosynthesis inhibition. Cu-induced community tolerance was always found, but it decreased with warming and time.Biodiversity loss associated with lower Cu tolerance under combined Cu exposure and increasing temperatures evidences the major influence of cumulative stressors on aquatic health. These results highlight the crucial interplay between environmental stressors, which are expected to intensify with climate change.

Molecular methods resolve the bacterial composition of natural marine biofilms on galvanically coupled stainless steel cathodes.

Navy vessels consist of various metal alloys and biofilm accumulation at the metal surface is thought to play a role in influencing metal deterioration. To develop better strategies to monitor and control metallic biofilms, it is necessary to resolve the bacterial composition within the biofilm. This study aimed to determine if differences in electrochemical current could influence the composition of dominant bacteria in a metallic biofilm, and if so, determine the level of resolution using metagenomic amplicon sequencing. Current was generated by creating galvanic couples between cathodes made from stainless steel and anodes made from carbon steel, aluminum, or copper nickel and exposing them in the Delaware Bay. Stainless steel cathodes (SSCs) coupled to aluminum or carbon steel generated a higher mean current (0.39mA) than that coupled to copper nickel (0.17mA). Following 3months of exposure, the bacterial composition of biofilms collected from the SSCs was determined and compared. Dominant bacterial taxa from the two higher current SSCs were different from that of the low-current SSC as determined by DGGE and verified by Illumina DNA-seq analysis. These results demonstrate that electrochemical current could influence the composition of dominant bacteria in metallic biofilms and that amplicon sequencing is sufficient to complement current methods used to study metallic biofilms in marine environments.

The role of extracellular polymeric substances on the sorption of pentachlorophenol onto natural biofilms in different incubation times: a fluorescence study

ABSTRACTExtracellular polymeric substances (EPS) have been regarded as the most significant components in sorbing organic contaminants onto natural biofilms. However, few investigations have reported the effects of EPS on the sorption of organic contaminants onto biofilms in different incubation times. Here, pentachlorophenol (PCP), selected as a model organic pollutant, was sorbed by biofilms and EPS-free biofilms cultured for 15, 30, and 45 days, to evaluate the role of EPS in the sorption process. EPS were extracted from biofilms to investigate the binding mechanism of PCP to the fluorescence matters in EPS by fluorescence quenching titration. Results showed that EPS, bound with biofilms or released to water, could increase or decrease the sorption amounts of PCP on biofilms in various incubation times. The presence of bound EPS in biofilms enhanced the sorption amounts of 30-day biofilms (6.0 ± 0.11 μmol g−1) because of the formation of the EPS-PCP complex between PCP and fluorescence matters in EPS, such as protein-like matters. In contrast, the release of PCP-bound EPS in 15- and 45-day biofilms decreased the sorption amounts of PCP on biofilms. All the results suggested that EPS dominated the sorption of PCP onto biofilms.

The effect of burdock leaf fraction on adhesion, biofilm formation, quorum sensing and virulence factors of Pseudomonas aeruginosa.

This study aimed to evaluate the effect of a fraction of burdock (Arctium lappa L.) leaf on the initial adhesion, biofilm formation, quorum sensing and virulence factors of Pseudomonas aeruginosa. Antibiofilm activity of the burdock leaf fraction was studied by the method of crystal violet staining. When the concentration of the burdock leaf fraction was 2·0mgml-1 , the inhibition rates on biofilm formation of P. aeruginosa were 100%. The burdock leaf fraction was found to inhibit the formation of biofilm by reducing bacterial surface hydrophobicity, decreasing bacterial aggregation ability and inhibiting swarming motility. Interestingly, the burdock leaf fraction inhibited the secretion of quorum-sensing (QS) signalling molecule 3-oxo-C12-HSL and interfered quorum sensing. Moreover, the QS-regulated pyocyanin and elastase were also inhibited. Chemical composition analysis by UPLC-MS showed 11 active compounds in the burdock leaf fraction. The burdock leaf fraction significantly inhibited the formation of biofilm and quorum sensing, as well as significantly decreased the content of virulence factors. This study introduces a natural and effective bacterial biofilm inhibitor, which could also significantly decrease the content of virulence factors and the drug resistance of P. aeruginosa.

Erratum to: Effect of the pollution level on the functional bacterial groups aiming at degrading bisphenol A and nonylphenol in natural biofilms of an urban river.

Erratum to: Effect of the pollution level on the functional bacterial groups aiming at degrading bisphenol A and nonylphenol in natural biofilms of an urban river.

Cyanobacteria as test organisms and biosorbents

Bioassay and biosorption potentials of different groups of cyanobacteria (CB)—Nostoc linckia (Roth.) Born. et Flah. No. 271, natural biofilms dominated by CB of Phormidium genus, and biofilms dominated by Nostoc commune (Vauch. Elenk)—were estimated. The physiological-biochemical response of CB to the influence of copper sulfate (II) (catalase activity with a gasometric method and dehydrogenase activity, lipid peroxidation, and chlorophyll ɑ and pheophytin contents with a spectrophotometric method) was studied; metal bioaccumulation was determined with a stripping voltammetry method. It was found that the communities dominated by Phormidium genus (CB biomass 0.2 g/dm3) removed copper compounds from the solutions with Cu2+ ion concentration of 20 mg/dm3 almost completely (by 99%); communities dominated by CB N. commune, by 87%; and pure culture of N. linckia, by 50%. Dehydrogenase and catalase activities and the intensity of bioluminescence proved to be sensitive indicators of the response of CB to Cu2+ ions. The impact of Cu2+ ions (20 mg/dm3) on a biofilm dominated by CB of Phormidium genus resulted in the fivefold decrease of catalase activity during 24 h; dehydrogenase activity decreased by nearly 357 times. The bioluminescence intensity during 24 h decreased by 1.3–100 times under the impact of Cu2+ (2 mg/dm3) and by 8.6–200 times in variants with a higher concentration of Cu2+ (20 mg/dm3). This regularity can be used as a test function in bioassay.

Spatial and temporal analogies in microbial communities in natural drinking water biofilms.

Biofilms are ubiquitous throughout drinking water distribution systems (DWDS), playing central roles in system performance and delivery of safe clean drinking water. However, little is known about how the interaction of abiotic and biotic factors influence the microbial communities of these biofilms in real systems. Results are presented here from a one-year study using in situ sampling devices installed in two operational systems supplied with different source waters. Independently of the characteristics of the incoming water and marked differences in hydraulic conditions between sites and over time, a core bacterial community was observed in all samples suggesting that internal factors (autogenic) are central in shaping biofilm formation and composition. From this it is apparent that future research and management strategies need to consider the specific microorganisms found to be able to colonise pipe surfaces and form biofilms, such that it might be possible to exclude these and hence protect the supply of safe clean drinking water.

Interactions between natural biofilm, substratum surface wettability, and mussel plantigrade settlement

Mytilus galloprovincialis is a major fouling organism in the inter-tidal zone. However, the interactions between M. galloprovincialis plantigrade settlement, biofilm characteristics, and surface wettability remains unknown. Here, we examined M. galloprovincialis plantigrade settlement responses to marine biofilms (BFs) on surfaces of varying wettability. No significant difference in mussel settlement was observed on young BFs (7 d) on surfaces of differing wettability; while settlement decreased on older BFs (14, 21, and 28 d) formed on low compared to high wettability surfaces. Surface wettability affected BF characteristics. The standardized harmonic mean and water contact angles values were not correlated with diatom density and chlorophyll a concentration, but were correlated with bacterial density, dry weight, and thickness. Denaturing gradient gel electrophoresis revealed that bacterial community structure differed on BFs on surfaces of varying wettability. Thus, surface wettability affects biofilm characteristics, and the subsequent changes in BF characteristics may be responsible for the variation in biofilm-inducing activity of M. galloprovincialis plantigrade settlement.

Laboratory grown subaerial biofilms on granite: application to the study of bioreceptivity

Simulated environmental colonisation of granite was induced under laboratory conditions in order to develop an experimental protocol for studying bioreceptivity. The experimental set-up proved suitable for producing subaerial biofilms by inoculating granite blocks with planktonic multi-species phototrophic cultures derived from natural biofilms. The ability of four different cultures to form biofilms was monitored over a three-month growth period via colour measurements, quantification of photosynthetic pigments and EPS, and CLSM observations. One of the cultures under study, which comprised several taxa including Bryophyta, Charophyta, Chlorophyta and Cyanobacteria, was particularly suitable as an inoculum, mainly because of its microbial richness, its rapid adaptability to the substratum and its high colonisation capacity. The use of this culture as an inoculum in the proposed experimental set-up to produce subaerial biofilms under laboratory conditions will contribute to standardising the protocols involved, thus enabling more objective assessment of the bioreceptivity of granite in further experiments.

Bacterial adhesion on direct and indirect dental restorative composite resins: An in vitro study on a natural biofilm

Statement of problemBoth direct and indirect techniques are used for dental restorations. Which technique should be preferred or whether they are equivalent with respect to bacterial adhesion is unclear. PurposeThe purpose of this in vitro study was to determine the affinity of bacterial biofilm to dental restorative composite resins placed directly and indirectly. Material and methodsFive direct composite resins for restorations (Venus Diamond, Adonis, Optifil, Enamel Plus HRi, Clearfil Majesty Esthetic) and 3 indirect composite resins (Gradia, Estenia, Signum) were selected. The materials were incubated in unstimulated whole saliva for 1 day. The biofilms grown were collected and their bacterial cells counted. In parallel, the composite resin surface morphology was analyzed with atomic force microscopy. Both bacterial cell count and surface topography parameters were subjected to statistical analysis (α=.05). ResultsIndirect composite resins showed significantly lower levels than direct composite resins for bacterial cell adhesion, (P<.001). No significant differences were observed within the direct composite resins (P>.05). However, within the indirect composite resins a significantly lower level was found for Gradia than Estenia or Signum (P<.01). A partial correlation was observed between composite resin roughness and bacterial adhesion when the second and particularly the third-order statistical moments of the composite resin height distributions were considered. ConclusionsIndirect dental restorative composite resins were found to be less prone to biofilm adhesion than direct composite resins. A correlation of bacterial adhesion to surface morphology exists that is described by kurtosis; thus, advanced data analysis is required to discover possible insights into the biologic effects of morphology.

Photoinhibition on natural ammonia oxidizers biofilm populations and implications for nitrogen uptake in stream biofilms

AbstractWe investigated photoinhibition on natural communities of ammonia oxidizing (AO) archaea (AOA) and bacteria (AOB) embedded in complex stream biofilms, and its implications on nitrogen uptake at biofilm scale. Based on the strong photoinhibition previously exhibited by free living and cultured AOA and AOB, we expected AO activity to decrease in biofilms experimentally exposed to light, reducing the contribution of microbial nitrification to ammonium uptake. To test it, we conducted light manipulation experiments in mesocosms using biofilms naturally developed on stream cobbles sides both exposed to light (light‐side) and facing the sediment (dark‐side). We observed a strong AO photoinhibition in dark‐side biofilms, accompanied by either biofilm‐scale decreases in nitrification or increases in nitrogen uptake likely by heterotrophic activity. Conversely, in light‐side biofilms photoinhibition was not observed suggesting that photoautotrophic layers may protect AO in situ by a sunshade effect. Experimental light and dark alternation cycles stimulated AO, enhancing both nitrification and ammonium uptake. These results support photoinhibition effects on natural AO communities, especially in biofilms developed under ambient dark conditions, whereas this effect seems to be buffered in biofilms developed under light conditions. Therefore, the contribution of nitrification to ammonium uptake at biofilm scale should consider not only the physiological study of AOA and AOB, but also the environmental conditions and community structure at the habitat microscale, since they may counterbalance, among others, the damaging light effects under natural conditions.

A Critical Role for Extracellular DNA in Dental Plaque Formation

Extracellular DNA (eDNA) has been identified in the matrix of many different monospecies biofilms in vitro, including some of those produced by oral bacteria. In many cases, eDNA stabilizes the structure of monospecies biofilms. Here, the authors aimed to determine whether eDNA is an important component of natural, mixed-species oral biofilms, such as plaque on natural teeth or dental implants. To visualize eDNA in oral biofilms, approaches for fluorescently stained eDNA with either anti-DNA antibodies or an ultrasensitive cell-impermeant dye, YOYO-1, were first developed using Enterococcus faecalis, an organism that has previously been shown to produce extensive eDNA structures within biofilms. Oral biofilms were modelled as in vitro “microcosms” on glass coverslips inoculated with the natural microbial population of human saliva and cultured statically in artificial saliva medium. Using antibodies and YOYO-1, eDNA was found to be distributed throughout microcosm biofilms, and was particularly abundant in the immediate vicinity of cells. Similar arrangements of eDNA were detected in biofilms on crowns and overdenture abutments of dental implants that had been recovered from patients during the restorative phase of treatment, and in subgingival dental plaque of periodontitis patients, indicating that eDNA is a common component of natural oral biofilms. In model oral biofilms, treatment with a DNA-degrading enzyme, NucB from Bacillus licheniformis, strongly inhibited the accumulation of biofilms. The bacterial species diversity was significantly reduced by treatment with NucB and particularly strong reductions were observed in the abundance of anaerobic, proteolytic bacteria such as Peptostreptococcus, Porphyromonas and Prevotella. Preformed biofilms were not significantly reduced by NucB treatment, indicating that eDNA is more important or more exposed during the early stages of biofilm formation. Overall, these data demonstrate that dental plaque eDNA is potentially an important target for oral biofilm control.

Analysis of the Start-Up of Natural Bio-Film Colonization of Constructed Rapid Infiltration System

Abstract this paper studied the start-up of natural bio-film colonization under the conditions where wet-to-dry ratio was 1:6, hydraulic load was 0.8m3/m2·d, and water temperature was 10°C-15°C, it took 25d in total to become mature, after the success of bio-film colonization, the removal rate of COD and NH4+-N can be stabilized at 80%, and the removal rate of NH4+-N can reach 90% above. The microorganism was divided into the following stages during bio-film colonization: 1.Reversible adhesion of cell on the surface of the carrier, 2.Irreversible adhesion of cell on the surface of the carrier, 3.Division of cell adhering to the surface of the carrier and formation of microcolony adhering to the surface of the carrier, 4.Growth of microcolony adhering to the surface of the carrier to be mature bio-film with three-dimensional structure. The process of natural bio-film colonization can be judged comprehensively according to removal rate of NH4+-N and COD, effluent pH as well as color of microbial film.

Trace levels of sewage effluent are sufficient to increase class 1 integron prevalence in freshwater biofilms without changing the core community

Most river systems are impacted by sewage effluent. It remains unclear if there is a lower threshold to the concentration of sewage effluent that can significantly change the structure of the microbial community and its mobile genetic elements in a natural river biofilm. We used novel in situ mesocosms to conduct replicated experiments to study how the addition of low-level concentrations of sewage effluent (nominally 2.5 ppm) affects river biofilms in two contrasting Chalk river systems, the Rivers Kennet and Lambourn (high/low sewage impact, respectively). 16S sequencing and qPCR showed that community composition was not significantly changed by the sewage effluent addition, but class 1 integron prevalence (Lambourn control 0.07% (SE ± 0.01), Lambourn sewage effluent 0.11% (SE ± 0.006), Kennet control 0.56% (SE ± 0.01), Kennet sewage effluent 1.28% (SE ± 0.16)) was significantly greater in the communities exposed to sewage effluent than in the control flumes (ANOVA, F = 5.11, p = 0.045) in both rivers. Furthermore, the difference in integron prevalence between the Kennet control (no sewage effluent addition) and Kennet sewage-treated samples was proportionally greater than the difference in prevalence between the Lambourn control and sewage-treated samples (ANOVA (interaction between treatment and river), F = 6.42, p = 0.028). Mechanisms that lead to such differences could include macronutrient/biofilm or phage/bacteria interactions. Our findings highlight the role that low-level exposure to complex polluting mixtures such as sewage effluent can play in the spread of antibiotic resistance genes. The results also highlight that certain conditions, such as macronutrient load, might accelerate spread of antibiotic resistance genes.

Effects of larval diet and metamorphosis cue on survival and growth of sea urchin post-larvae (Paracentrotus lividus; Lamarck, 1816)

In this study, we present the results of two experiments; in the first one we evaluated the effects of four larval dietary treatments on the survival and growth of the sea urchin Paracentrotus lividus, larvae and post-larvae. In the second experiment we have measured the effects of two different settlement substrates, combined with the presence of conspecifics, on metamorphosis, survival and growth of post-larvae. The microalgae dietary treatments consisted in: Dunaliella tertiolecta (Duna); 50% mixture of Isochrysis galbana and D. tertiolecta (ID); 50% mixture of Chaetoceros gracilis and D. tertiolecta (CD); 33% mixture of I. galbana, C. gracilis and D. tertiolecta (ICD). Although all dietary treatments resulted in a good survival at competence, significant difference in post-larval survival was observed between treatments, and indeed, only larvae fed Duna and CD survived to 180days post settlement (DPS).In the second experiment, the settlement substrates consisted in a film of cultured Ulvella lens or a naturally developing biofilm of diatoms, and the employed rearing water was either natural seawater or seawater previously exposed to P. lividus adults. At 10DPS, larger (p<0.05) post-larvae were observed in the natural biofilm treatment, whilst the presence of conspecifics significantly increased larval settlement in both substrates (p<0.01).These results indicate that it is important to consider the survival of post-larvae and juveniles to establish the efficiency of the dietary treatment on the hatchery production of P. lividus. Furthermore, it suggests that improved settlement protocols, such as the use of conspecifics, could contribute to increase hatchery outputs. Finally, it confirms the suitability of U. lens as settlement cue but also highlights that further research is required to establish its effectiveness for post-larvae first feeding. Statement of relevanceOur work contributes to improving hatchery rearing methods of larvae and post-larvae of the sea urchin Paracentrotus lividus.