The project implementation is done according to the Consortium Agreement. The purpose of the consortium agreement is to specify the relationship among the Partners, in particular concerning the organization of the work between the Partners, the management of the Project and the rights and obligations of the Partners regarding the project and the results of the FUNCELL. The project works and the responsibilities for the tasks of each Partner are to be arranged as agreed in the Project Plan.
DFR Systems SRL, as the Coordinating Partner and will be responsible for the project management in accordance with the Manunet Guidelines.
DFR Systems has obtained results in using MBBR technology and holds a patent for a model of biofilm carrier (Patent no. RO 123174/28.01.2011).
In MBBR systems the biofilm grows protected within small plastic carriers, which are carefully designed with high internal surface area. These biofilm carriers are suspended and mixed throughout the water phase. The biological wastewater treatment process consists of adding biofilm carriers in aerated or anoxic basins to support biofilm growth. When the microorganisms in the attached biological film die, the film breaks up and peels off from the solid support being carried away by the liquid current. The destroyed cellular material is removed as sludge.
Cerioporus squamosus is a basidiomycete bracket fungus (also known as dryad’s saddle or pheasant’s back mushroom) (Spahr, 2009), and has special importance in natural ecosystems, being able to degrade a wide range of cellulosic substrates. The strain develops Dimitic hyphae, which are composed of skeletal hyphae (axial or inflated) (Zmitrovich, 2016).
Fresh culture strain was started in Potato-Dextrose Nutrient Broth (Scharlau), for 4 days at 28°C, at fermenter level (Biotec FE 007), in a final volume of 500 mL, environmental volume for 72 hours at 28°C under aerobic conditions (continuous oxygenation) and agitation set to 250 rpm.
After strain growing, for 4 days at 28oC, MBBR pieces (previously sterilized at 121oC for 15’) were put in the fermenter, and the process was further continued for 7 days at 28oC.
Microscopic analysis and morphological characterization was performed by scanning electron microscopy (SEM) using a Quanta 200, Fei (Netherlands) electron microscope (Figure 3). SEM analysis was also conducted on Cerioporus squamosus strain grown on Sabouraud-Agar plates, incubated at 28oC for 14 days, for hyphae development, and compared to liquid media growth, which was used for MBBRs functionalization.
Morphological analyses were performed on the GSED detector, ESEM mode, spot beam size of 4.0, 10kV filament voltage, with image acquisition at 27.2 seconds.
Optical microscopy analysis was carried out on an Olympus SZX7 stereomicroscope, with 7:1 zoom ratio, built-in electrostatic discharge protection, and advanced Galilean optical system for highly resolved images. Analyses were carried out at a magnification level of 0.8X, on both treated and untreated MBBRs.
Indigenous Ascomycota are the dominant fungal phylum in polluted environments, where they are able to transform or remove also significantly stable and toxic recalcitrants (Marco-Urrea et al., 2015). Polluted wastewaters represent a source of Ascomycota, putative degraders for which studies are surprisingly scarce. In this context, many unstudied fungal species need to be explored, to understand their specific interactions in engineered ecosystems, because they appear to play a primary role in actual polluted scenarios. Some Ascomycota undoubtedly possess great potential for bioremediation purposes (Mariner et al., 2008), and the key seems to be the intracellular enzymatic machinery coded in their genome, rather than the extracellular battery of oxidative enzymes already described for White Rot Fungi.
Optical microscopy analyses revealed Cerioporus squamosus biomass deposition inside the MBBR structure (b) when compared to untreated MBBR (a).
Analysis highlighted attachment of microbial biomass inside the MBBRs, but not on the external spaces of the polymeric structures. It can be highlighted that a longer process (more than 7 days) could translate into a higher quantity of biomass that will be internalized between the MBBRs empty spaces (inside the structure). Another aspect that was noted is that the biomass was not firmly attached by the polymer surface, and could be easily washed.
SEM analysis on Cerioporus squamosus standalone strain, grown on Sabouraud-Agar Petri plates, carried out at pressure levels between 200Pa and 208Pa highlighted a fibrillated, branched structure with a high degree of coverage on the carbon band and hyphae size ranging from 1.60μm to 2.72μm, for 4000x magnification analysis. The strain has a multi branches hyphae organization, these having dimensional width variations, and pyramidal structure in section (see below a SEM analysis of morphological characters for Cerioporus squamosum strain).