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Abundance profiling via 16S rRNA targeted next generation sequencing (NGS) is a common procedure to characterize mixtures of prokaryotic populations inhabiting an environment. Depending on the variable region/s addressed, different maps can be obtained due to their different information content. In this work, we focussed on wastewater microbial communities and we compared several recently developed universal primers that addressed regions V1-V3, V3-V4 and V4. They all proved to have good performance over a wide range of microbial phyla, but the phylum Planctomycetes was not optimally covered, especially for members of the Brocadiales family. Such bacteria are at the basis of the novel nitrogen removal strategy based on anammox process. To overcome this limitation we performed an extensive bioinformatic analysis that allowed the design of a primer (Pro341FB) that shows increased sensitivity for this specific phylum with respect to the previously proposed Pro341F primer. Upon validation using a 16S NGS survey on microbial communities from different wastewater treatment plant (activated sludge systems, anaerobic digesters, aerobic and anaerobic granules) we demonstrated that Pro341FB is able to reveal up to 5 times more members of the Candidatus Brocadiales family (plus many other previously under-covered prokaryotes) than Pro341F, without affecting its excellent previous coverage.

Ralstonia solanacearum (Rs), the causal agent of tomato southern bacterial wilt and of potato brown rot, affects more than 250 plant species and it is included in the EPPO list of quarantine pathogens (QP). On tomato, the disease is characterized by a general wilting of the plant due to the colonization of its vascular system. Rs is a species complex with a high genetic variability and, based on the host plants, is currently divided into 5 races. In Emilia-Romagna Region (Northern Italy), during the last 15 years, Rs presence has been detected in potatoes, tomatoes and in superficial waters in nearby cultivated areas. 27 Rs strains (belonging to races 1 and 3) were characterised by Rep-PCR (BOX, ERIC, REP). The binary matrices from PCR profiles were used for UPGMA analysis in single or concatenated way. PD 2762 Rs and DISTAL 9081 Xanthomonas axonopodis pv. vitians strains were used as race 3 reference and as outgroup, respectively. From the concatenated profiles, the 27 Rs strains were divided in the two macro-groups of races 3 and 1 with a 15% similarity. The 6 strains race 3 were subdivided in 2 groups with a 50% similarity and 5 of them were more similar to each other than the PD 2762 strain. The 21 strains race 1 were subdivided into 4 groups with more than 60% similarity. The high degree of genetic variability particularly among the strains of race 1 (isolated in Parma and Ferrara provinces) suggests the presence of different haplotypes probably due to the movement of non-symptomatic infected material.

Sourdough fermentation influences several properties of leavened baked goods also because Lactic acid bacteria (LAB) and yeasts produce bioactive peptides with a positive effect on human health. In an early study, three Lactobacilli strains (L. farciminis H3 and A11 and L. sanfranciscensis I4) possessing different proteolytic activities were used to produce sourdoughs containing peptides equipped with anti-inflammatory and/or antioxidant properties. This work was aimed to assess whether these properties could be retained after cooking. The selected LABs were used to produce breads from which low molecular weight (LMW-) peptides were extracted. The results provide solid proofs of keeping both antioxidant and anti-inflammatory activities of peptides from cooked products. Sequences of LMW-peptides either from doughs and breads were determined by mass spectrometry: differences have been noticed in amino acidic composition and in sequences, however, all the strains produce peptides equipped with antioxidant and anti-inflammatory activities.

Honey is a food known for its medical properties. In this work, we have studied the impact of different types of honey on insulin signalling pathway. We found that honey extracts inhibit the enzyme PTP1B, one of the main negative regulators of insulin receptor signalling. HPLC-MS analysis allowed us to confirm the presence of several natural PTP1B inhibitors in the honey extracts analysed. Statistical analysis methods show a correlation between specific 1H-NMR resonance frequencies/HPLC peaks and the inhibitory power of the samples. This finding will allow the prediction of the biological properties of honey samples applying relative simple analytical methods. Finally, we demonstrated that the treatment of HepG2 cells with honey extracts enhances the expression of insulin receptor, and stimulates glucose uptake. For the first time, our results demonstrate that bioactive components of honey could improve glycaemic control by both inhibiting PTP1B and stimulating the expression of insulin receptor in liver cells.

In this work, a non-metallicolous and a metallicolous population of S. paradoxa were exposed to copper excess and fungal elicitation, and investigated for phytohormone production and cytological alterations. Under the stress applied separately and in combination, S. paradoxa plants varied phytohormone concentration in a population-specific way, suggesting a different signalling in response to biotic and abiotic stimuli according to the environment of origin. Generally, the stress responses consisted in increased levels of salicylic acid, auxin, and gibberellin in the non-metallicolous population, and of jasmonic and abscisic acid in the metallicolous one. Interestingly, the metallicolous population increased the level of such phytohormones following exposure to the fungal elicitor only in the presence of copper. This alternative hormonal signalling could derive from the incompatibility between the ordinary ROS-mediated response to pathogens and the acquired mechanisms that prevent oxidative stress in the population from the metal-rich soil. Furthermore, stress-induced autophagic phenomena were more evident in the non-metallicolous plants than in the metallicolous ones, suggesting that the adaptation to the metalliferous environment has also affected autophagy intensity and signalling in response to copper excess and fungal elicitation.

This work reports on simultaneous nitrification, denitrification and phosphorus removal treating real domestic wastewater with low carbon/nitrogen (C/N) ratio by aerobic granular sludge (AGS). Operations at high sludge retention time (SRT = 61 ± 24 days) resulted in low biomass yield per chemical oxygen demand removed (CODrem) (0.21 ± 0.01 gCODx/gCODrem), lower COD demand for denitrification as well as high effluent quality in terms of total suspended solids (TSS) (22 ± 7 mgTSS/L). The average ratio between the biodegradable soluble COD stored anaerobically as polyhydroxyalkanoates (PHAs) and the N removed was 3.1 ± 0.6 gCODsto/gNrem, suggesting that nitrification/denitrification occurred partly via the nitrite pathway. Results revealed that stable AGS process with high C/N/P removal efficiency of 84/71/96% can be obtained besides a low organic loading rate (0.43 ± 0.11 g COD/L/d) and influent C/N ratio (3.8 ± 1.6 g/g), resulting in a high effluent quality characterized by 25 ± 6 mg sCOD/L, 0.09 ± 0.07 mgPO4-P/L, 9 ± 2 mgTIN/L (10 ± 2 mgTN/L) and 22 ± 7 mgTSS/L.

Cerato-platanin (CP) is a non-catalytic, cysteine-rich protein, the first member of the cerato-platanin family. It is a single-domain protein with a double Ψ/β barrel domain resembling the D1 domain of plant and bacterial expansins. Similarly to expansins, CP shows a cell wall-loosening activity on cellulose and can be defined as an expanisin-like protein, in spite of the missing D2 domain, normally present in plant expansins. The weakening activity shown on cellulose may facilitate the CP-host interaction, corroborating the role of CP in eliciting plant defence response. Indeed, CP is an elicitor of primary defences acting as a Pathogen-Associated Molecular Patterns (PAMP). So far, structure-function relationship study has been mainly performed on the bacterial BsEXLX1 expansin, probably due to difficulties in expressing plant expansins in heterologous systems. Here, we report a subcloning and purification method of CP in the engineered E. coli SHuffle cells, which proved to be suitable to obtain the properly folded and biologically active protein. The method also enabled the production of the mutant D77A, rationally designed to be inactive. The wild-type and the mutated CP were characterized for cellulose weakening activity and for PAMP activity (i.e. induction of Reactive Oxygen Species synthesis and phytoalexins production). Our analysis reveals that the carboxyl group of D77 is crucial for expansin-like and PAMP activities, thus permitting to establish a correlation between the ability to weaken cellulose and the capacity to induce defence responses in plants. Our results enable the structural and functional characterization of a mono-domain eukaryotic expansin and identify the essential role of a specific aspartic residue in cellulose weakening.