Publications

Influence of ozonation on the in vitro mutagenic and toxic potential of secondary effluents

Pseudomonas Fluorescens biofilm in rotating annular bioreactor: formation kinetics and wetting properties

<p>Biofilms are bacterial communities embedded in an extracellular matrix, able to adhere to surfaces. A deeper knowledge of the biofilm as a whole will aid the development of efficient methods to control deleterious biofilms (clinical biofilms, biofouling) or to enhance beneficial ones (waste-water treatment, bio-filtration). P. fluorescens has been widely studied, this strain produces bioactive secondary metabolites, and forms biofilms[1]. Several experimental set-ups have been widely used for in vitro biofilm cultivation of P. fluorescens, even if a deep characterization among different culture conditions is still lacking in the literature. This work, based on previous studies[2], is focused on the investigation of growth conditions on biofilm structure and properties. Growth kinetics of P. fluorescens biofilms was characterized in vitro under stagnant and flow-controlled conditions, using a rotating annular bioreactor. Two different supports in borosilicate glass and polycarbonate have been used. Bacterial growth kinetics has been measured through bio-turbidity analysis and TOC/DOC quantification. Biofilm morphology has been quantified through optical microscopy and image analysis by measuring the fraction of support surface covered by biofilm. The wetting properties of the biofilm layers have been investigated by using an innovative device, named Kerberos®, able to control centrifugal and gravitational forces acting on a single droplet placed on a surface[3]. The evolution of the droplet shape and position was measured as function of the imposed stress, to quantify wetting of different biofilm coated samples, following already assessed methodologies[4]. Different chemo-physical environments, investigated by changing growth medium, physical support, and imposed flow stress, induced different growth kinetics, biofilm morphology, and wetting properties. Accurate experimental measurements allowed us to estimate in a quantitative way the influence of investigated parameters on specific morphologic measurements.</p> <div><br /> <div> <p>[1] Brittan S. Scales and others, ‘Microbiology, Genomics, and Clinical Significance of the Pseudomonas Fluorescens Species Complex, an Unappreciated Colonizer of Humans’, Clinical Microbiology Reviews, 27.4 (2014), 927–48 <https://doi.org/10.1128/CMR.00044-14>.</p> </div> <div> <p>[2] Federica Recupido and others, ‘The Role of Flow in Bacterial Biofilm Morphology and Wetting Properties’, Colloids and Surfaces B: Biointerfaces, 2020 <https://doi.org/10.1016/j.colsurfb.2020.111047>.</p> </div> <div> <p>[3] Sotiris P. Evgenidis and others, ‘Kerberos: A Three Camera Headed Centrifugal/Tilting Device for Studying Wetting/Dewetting under the Influence of Controlled Body Forces’, Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2017 <https://doi.org/10.1016/j.colsurfa.2016.07.079>.</p> </div> <div> <p>[4] Inmaculada Ríos-López and others, ‘Effect of Initial Droplet Shape on the Tangential Force Required for Spreading and Sliding along a Solid Surface’, Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2018 <https://doi.org/10.1016/j.colsurfa.2018.04.004>.</p> </div> </div>

Wetting and Imbibition Characteristics of <i>Pseudomonas fluorescens</i> Biofilms Grown on Stainless Steel

This study aims to provide insights into biofilm resistance associated with their structural properties acquired during formation and development. On this account, the wetting and imbibition behavior of dehydrated Pseudomonas fluorescens biofilms grown on stainless steel electropolished substrates is thoroughly examined at different biofilm ages. A polar liquid (water) and a non-polar liquid (diiodomethane) are employed as wetting agents in the form of sessile droplets. A mathematical model is applied to appraise the wetting and imbibition performance of biofilms incorporating the evaporation of sessile droplets. The present results show that the examined biofilms are hydrophilic. The progressive growth of biofilms leads to a gradual increase of substrate surface coverage─up to full coverage─accompanied by a gradual decrease of biofilm surface roughness. It is noteworthy that just after 24 h of biofilm growth, the surface roughness increases about 6.7 times the roughness of the clean stainless steel surface. It is further found that the imbibition of liquid in the biofilm matrix is restricted only to the biofilm region under the sessile droplet. The lack of further capillary imbibition into the biofilm structure, beyond the droplet deposition region, implies that the biofilm matrix is not in the form of an extended network of interconnected micro/nanopores. All in all, the present results indicate a resilient biofilm structure to biocide penetration despite its hydrophilic nature.

Sardine Processing Waste: Biological Treatment Strategies and Their Implications

This study explores sustainable methods for Sardine Processing Waste (SPW) valorization. Two approaches were investigated: (a) SPW microbial pretreatment adding Saccharomyces cerevisiae or Bacillus sp. in a two-stage anaerobic digestion (AD) for enzyme and biomethane production and (b) a single-stage AD without SPW pretreatment. Both S. cerevisiae and Bacillus sp. secreted proteases (0.66 and 0.58 U mL−1, respectively) and lipases (3.8 and 4.3 U mL−1, respectively) during hydrolysis, thus reducing viscosity (2.8 and 2.9 cP, respectively) compared with the untreated SPW (4.1 cP). Biomethane production was higher in the single-stage AD (1174 mL CH4 g−1 VS−1) when compared with the two-stage AD (821.5 and 260 mL CH4 g−1 VS−1 with S. cerevisiae and Bacillus sp., respectively). S. cerevisiae addition enhanced SPW degradation as implied by VS and sCOD values (70 and 84%, respectively), but this also resulted in a higher toxicity due to a three-fold increment in NH4-N content, reducing methanogen activity. This research demonstrates the innovative application of S. cerevisiae, a common bread-making yeast, in the biotechnological enhancement of SPW hydrolysis. Non-genetically engineered S. cerevisiae not only co-produced proteases and lipases but also significantly improved solubilization, degradation, and viscosity reduction, thereby rendering the yeast a key player in solid fish waste valorization, beyond its traditional applications.

Highly sensitive resistance spectroscopy technique for online monitoring of biofilm growth on metallic surfaces

ΔΟΚΙΜΕΣ ΕΚΠΛΥΣΗΣ ΚΑΙ ΟΙΚΟΤΟΞΙΚΟΤΗΤΑ ΔΕΙΓΜΑΤΩΝ ΙΛΥΟΣ ΑΠΟ ΕΓΚΑΤΑΣΤΑΣΕΙΣ ΕΠΕΞΕΡΓΑΣΙΑΣ ΛΥΜΑΤΩΝ

Δeίγματα ιλύος από Eγκαταστάσeις Eπeξeργασίας Λυμάτων (EEΛ), μe διαφορeτική πeριeκτικότητα σe οργανική ύλη, υποβλήθηκαν σe φυσικοχημικές αναλύσeις, δοκιμές έκπλυσης και οικοτοξικολογικές δοκιμές μe σκοπό τη διeρeύνηση των πeριβαλλοντικών eπιπτώσeων από τη διάθeση της ιλύος στο έδαφος. Πραγματοποιήθηκαν πeιράματα φυτοτοξικότητας σe τρeις ανώτeρους φυτικούς οργανισμούς, ( Sorghum saccharatum, Lepidium sativum and Sinapis alba ), σe αναλογίeς μίγματος ιλύος προς έδαφος (2.5-25%). Eπιπλέον, τα δeίγματα ιλύος  υποβλήθηκαν σe δοκιμές έκπλυσης, σύμφωνα μe τη δοκιμή έκπλυσης NEN 7341 (δύο στάδια έκπλυσης αθροιστικής αναλογίας υγρού/στeρeού, L/S= 100 L/kg) και τη δοκιμή ανοδικής διήθησης EN 14405, σe αθροιστικές αναλογίeς L/S= 0.1, 0.2, 0.5, 1.0, 2.0, 5.0, 10 και 20 L/kg. Τα οικοτοξικολογικά χαρακτηριστικά των eκπλυμάτων, προσδιορίστηκαν μe βάση την eπίδρασή τους στη φωταύγeια του βακτηρίου V. fischeri και στην κινητικότητα του καρκινοeιδούς D. magna . Τα δeίγματα παρουσίασαν φυτοτοξική δράση σe αναλογίeς μίγματος ιλύος/eδάφους μeγαλύτeρeς από 5%, eνώ τα υγρά έκπλυσης παρουσίασαν σe ορισμένeς πeριπτώσeις υψηλές τιμές οικοτοξικότητας, έως και 100%.

Effect of surface treatment and shear flow on biofilm formation over materials employed in space water storage and distribution systems

The prolonged duration of future manned space missions conceals potential threats associated with microbial contamination. Such closed environments are susceptible to formation of complex biofilm communities, where microorganisms can thrive and further evolve. The objective of this study was to evaluate the impact of surface type, surface treatment and shear stress on biofilm formation in water facilities. To that aim, the ability of Pseudomonas fluorescens SBW25 to adhere on three space applications related materials, including passivated (SS) and both passivated and electropolished (SSEP) stainless steel, as well as Ti-6Al-4V (Ti) alloy was studied under stagnant and shear stress conditions after 24 h of exposure. Results indicated that surface type strongly affects bacterial adhesion under the same conditions. Surface coverage during static experiments was in the following order: SS &amp;gt; Ti &amp;gt; SSEP, while SS exhibited a fourfold surface coverage compared to SSEP highlighting the significance of surface treatment. Moreover, SS and Ti stimulate the formation of several microcolonies and their growth. On the other hand, the application of shear stress diminished bacterial attachment to the studied materials, the degree of which relied on the material type. In this case, bacterial settlement on SS and Ti was dependent on the surface texture, implying that surface roughness may also play an important role in cell adhesion under shear conditions. Furthermore, the metallic surfaces did not hinder bacterial attachment when silver ions were previously deposited on their surface. The deposition that occurs on metallic surfaces when in contact with water disinfected with silver ions, for example, during space missions, highlights its impact on the loss of disinfection capacity of silver ions.

Wetting properties of dehydrated biofilms under different growth conditions

An integrated sequential injection analysis system for ammonium determination in recycled hygiene and potable water samples for future use in manned space missions

Ecotoxicological properties of wastewater treated using tertiary methods

Abstract The objective of this work was the examination of the efficiency of coagulation and ozonation processes for the production of reclaimed wastewater with low toxicity. Municipal secondary effluents were treated by FeCl 3 , Al 2 (SO 4 ) 3 (alum), and a commercial substance at metal ion concentrations of 0.5 and 1 mmol/L. Alternatively, the effluents were treated by ozonation in a semibatch ozone reactor. The feed gas was introduced at a flowrate of 3 L/min containing ozone at various concentrations, ranging between 2.5 and 8 mg/L; ozone residence times were 2, 5, 15, and 30 min. The toxic effects of the advanced treated effluents were examined by a battery of tests using the marine bacteria Vibrio fischeri , the freshwater crustaceans Daphnia magna , Daphnia pulex , and Thamnocephalus platyurus , and the rotifers Brachionus calyciflorus . The addition of alum decreased the toxic effect of reclaimed wastewater on immobilization of D. pulex , from 90 to 60%. Ozonation was also effective for toxicity removal, which decreased to 25% effect on D. pulex after treatment by 2.5 mg O 3 /L for 2 min. However, acute toxic effects after ozonation, were observed on V. fischeri and were related to ozone gas concentration and contact time. At the highest ozone dosage, the toxicity reached almost 100% inhibition of bioluminescence after 15 min. The toxicity of the ozonated effluents to bacteria decreased with sample storage time and was almost negligible after 48 h, indicating that the potential adverse effect of reclaimed wastewaters on receiving waters might be reduced by storage for a certain time. © 2006 Wiley Periodicals, Inc. Environ Toxicol 21: 417–424, 2006.

Detecting SARS-CoV-2 lineages and mutational load in municipal wastewater; a use-case in the metropolitan area of Thessaloniki, Greece

Abstract The SARS-CoV-2 pandemic represents an unprecedented global crisis necessitating novel approaches for, amongst others, early detection of emerging variants relating to the evolution and spread of the virus. Recently, the detection of SARS-CoV-2 RNA in wastewater has emerged as a useful tool to monitor the prevalence of the virus in the community. Here, we propose a novel methodology, called lineagespot , for the detection of SARS-CoV-2 lineages in wastewater samples using next-generation sequencing. Our proposed method was tested and evaluated using NGS data produced by the sequencing of three wastewater samples from the municipality of Thessaloniki, Greece, covering three distinct time periods. The results showed a clear identification of trends in the presence of SARS-CoV-2 mutations in sewage data, and allowed for a robust inference between the variants evident through our approach and the variants observed in patients from the same area time periods. Lineagespot is an open-source tool, implemented in R, and is freely available on GitHub .

Potential Ozone Applications for Water/Wastewater Treatment

Abstract Several applications of ozonation were examined in this study for: the treatment of stabilized high strength municipal landfill leachates, the reclamation potential and toxicity reduction of municipal secondary effluents, and the removal potential of phytoplanktons from surface waters. The major parameters examined were the applied ozone dosage and the respective contact time. The application of single ozonation on leachates resulted in the efficient removal of color and organic loading, due to the respective oxidation, induced by ozonation. In addition, ozonation was found to be effective for the removal of the residual organic content of secondary municipal effluents. However, acute toxic effects after ozonation were observed on V. fischeri and were related to ozone concentration and contact time. Furthermore, the surface water used for drinking water production, was subjected to ozonation treatment for the removal of harmful cyanobacteria. Ozonation resulted in the reduction of the number of cyanobacteria species and in the breakage of the chain‐type species to cells with a lower number of atoms.

A Highly Hydrophobic Siloxane-Nanolignin Coating for the Protection of Wood

Wood, a vital material for both modern and heritage objects, is particularly susceptible to degradation caused by water due to its hydrophilic nature and porous structure. Therefore, developing sustainable strategies to protect wood is of significant importance. This study aims to produce a highly hydrophobic coating for the protection of wood following a straightforward procedure and using materials that are compatible with wood. First, nano/sub-microlignin (NL) is isolated and produced from beech wood through a one-step tailored organosolv process. Next, NL is incorporated into Sivo 121, a water-borne and solvent-free silane system recommended by the manufacturer for protecting wood surfaces. Composite coatings containing various concentrations of NL and Sivo 121 are applied to chestnut (Castanea spp.) and oak (Quercus spp.). The impact of NL concentration on the contact angles of water drops (CAs) and colour changes (ΔE) of the treated wood specimens is investigated. The coating with 4% w/w NL demonstrates enhanced hydrophobicity (CA = 145°) and has a negligible effect on the colour of pristine oak (ΔE &lt; 3). The wetting properties of coated oak are not affected after 100 tape peeling cycles. However, the coating exhibits poorer performance on chestnut, i.e., CA = 135°, which declines after 80 peeling cycles, and ΔE &gt; 5. The drop pH does not have any noticeable effect on CA. The latter remains stable even after prolonged exposure of coated oak and chestnut samples to artificial UV radiation and outdoor environmental conditions. Finally, the composite coating offers good and comparable protection for both wood species in the biological durability soil burial test

Hexavalent chromium release from lignite fly ash and related ecotoxic effects

Hexavalent chromium Cr(VI) is a pollutant of immense concern due to its high mobility to water sources and highly toxic properties. In most cases, Cr(VI) could be released from lignite fly ash in aquatic environment when fly ash comes into contact with water. In this study, the contribution of the leaching patterns and bioavailability of Cr(VI) from lignite fly ash to the overall ecotoxic properties of fly ash leachates was originally examined and leaching procedures were evaluated in this context. A series of customized leaching tests were conducted and a battery of ecotoxicity tests including the crustacean Daphnia magna and the photobacterium Vibrio fischeri was applied. The leaching of Cr(VI) was pH and liquid to solid (L/S) ratio dependent, exhibiting the highest releases at pH values between 7 and 8. At the liquid to solid ratio (L/S) equal to 100 L/kg, the (CrVI) release reached a plateau, implying the presence of diffusion constrains and/or solubility hindrances. The toxic effect of the leachates obtained under leaching at pH 7 towards D. magna was relatively high (TU = 28.6 (23.8-35.7) at L/S = 10 L/kg). Interestingly, the toxicity of the leachates towards D. magna not only was significantly correlated to Cr(VI) (r = 0.961, P < 0.01), but the toxicity of the leachates (in absolute values) was matching the toxicity of the Cr(VI) revealing its remarkable contribution to the overall effect. In addition, the lower sensitivity of the bacteria V. fischeri when exposed to the leachates, along with the time dependence of the toxicity profiles supported the interpretation of the results obtained in this study.

PLASMATRON GASIFICATION OF RAW WASTE VEGETABLES FROM STEAM MECHANICAL HEAT TREATMENT

Application of plasma in biomass energy production is a novel and potential technology. The aim of this work was to study the feasibility and operational performance of plasmatron (plasma torch) gasification of biomass wastes, taking raw waste vegetables mixed with raw wood (RWV/RW) from Steam Mechanical Heat Treatment (SMHT) as the target material (VRM). The SMHT is one of the alternatives to pre-treat the municipal solid waste (MSW) before its further separation and reutilization. This plasmatron process has several advantages including high heating rate, short heating time, no viscous tar and low residual char or lava. The production of CO and H2 (syngas) which are the major components in the gas products (90.05–93.43 vol.%), occurs with relatively high reaction rates. The maximum concentrations of gaseous products occurring times are between 0.25 to 0.75 minutes. Almost 90% of the gaseous products were appeared in 2 min of reaction time. With the injection of steam, the mass yields of H2 and CO2 also increase. The inorganic components are converted into non-leachable vitrified lava which is non-hazardous. Steam injection prefers to the syngas production, promotes the gasification of VRM completely and also shows a positive benefit for the inhibition of greenhouse gas (GHG) (CH4+CO2) production in the major reaction time. The gasification of VRM by plasmatron changed the BET surface area to 10.33 times from 573 K to 873 K. After steam injections, the larger BET surface area of 241.27 m 2 g -1 at 873 K with 1 mL min -1 water flow rate resulting from steam gasification are believed to be due to the activation by steam and higher density of plasma. All the information obtained in this study is useful for the rational design and proper operation of plasmatron gasification system for treating VRM.

Detecting SARS-CoV-2 lineages and mutational load in municipal wastewater; a use-case in the metropolitan area of Thessaloniki, Greece

Abstract The SARS-CoV-2 pandemic represents an unprecedented global crisis necessitating novel approaches for, amongst others, early detection of emerging variants relating to the evolution and spread of the virus. Recently, the detection of SARS-CoV-2 RNA in wastewater has emerged as a useful tool to monitor the prevalence of the virus in the community. Here, we propose a novel methodology, called lineagespot, for the monitoring of variants and the detection of SARS-CoV-2 lineages in wastewater samples using next-generation sequencing (NGS). Our proposed method was tested and evaluated using NGS data produced by the sequencing of fourteen wastewater samples from the municipality of Thessaloniki, Greece, covering a six month period. The results showed a clear identification of trends in the presence of SARS-CoV-2 mutations in wastewater data. Lineagespot was able to record the evolution and rapid domination of the B.1.1.7 lineage in the community, and allowed for a robust inference between the variants evident through our approach and the variants observed in patients from the same area time periods. Lineagespot is an open-source tool, implemented in R, and is freely available on GitHub.

Contribution of the Gravity Component and Surface Type During the Initial Stages of Biofilm Formation at Solid–Liquid Interfaces

Water systems are highly vulnerable to biofilm formation, which can compromise water quality, operational efficiency, and public health. Factors such as surface material properties and gravitational orientation of the surface play critical roles in the early stages of microbial attachment and biofilm development. This study examines the impact of gravity and surface composition on the initial adhesion of Pseudomonas fluorescens AR11—a model organism for biofilm research. Focusing on stainless steel (SS) and polycarbonate (PC), two materials commonly used in water and wastewater infrastructure, bacterial adhesion was evaluated at surface inclinations of 0°, 45°, 90°, and 180° to assess gravitational impact. After three hours of contact, fluorescence microscopy and image analysis were used to quantify surface coverage and cluster size distribution. The results showed that both material type and orientation significantly affected early biofilm formation. PC surfaces consistently exhibited higher bacterial adhesion at all angles, with modest variations, suggesting that material properties are a dominant factor in initial colonization. In contrast, SS showed angle-dependent variation, indicating a combined effect of gravitational convection and surface characteristics. These insights contribute to a deeper understanding of biofilm dynamics under realistic environmental conditions, including those encountered in space systems, and support the development of targeted strategies for biofilm control in water systems and spaceflight-related infrastructure.

Toxicological and ecotoxic impact of secondary and tertiary treated sewage effluents