Tannery sludge valorization through zeolite-assisted anaerobic process for short-chain fatty acids (SCFAs) production.

Tannery sludge, a challenging waste, was utilized as a substrate for the production of Short-Chain Fatty Acids (SCFAs) through a series of six thermophilic Continuous Stirred-Tank Reactor runs. The sludge was subjected to a mild thermal pre-treatment and incorporated zeolites (chabazite in run II, and clinoptilolite in run III) in the acidification process. Results highlighted zeolites' impact on chromium concentration and the SCFAs/CODSOL ratio. Ammonia release remained consistent at around 47 % and 51 % for run I and II, respectively, but surpassed 60% in run III, suggesting limited zeolite effectiveness in NH4 absorption. Chromium release in the liquid fraction, due to thermal pretreatment, reached 335 mg/L. While in tests without zeolite, complete removal proved challenging, in zeolite-amended runs, complete removal was achieved, showcasing the materials' heavy metal absorption capacity. SCFA concentrations reached 20260 mgCOD/L, with acidification efficiency varying; runs I and III had ratios around 0.70 COD/COD, while run II showed substantial improvement (0.92) with chabazite. Anaerobic fermentation-digestion mass balance indicated a 41% reduction in landfill sludge mass, reducing its environmental footprint while yielding valuable byproducts like biogas and SCFAs. These findings underscore zeolites' potential in heavy metal absorption and acidification process enhancement, paving the way for applications with tannery sludge.

Oxygen limitation in aerobic polyhydroxyalkanoates production from sewage sludge anaerobic fermentation liquids under low and medium organic loading rate.

The present study describes the microbial production of polyhydroxyalkanoates (PHA) from thermally pre-treated sewage sludge at pilot scale level, investigating for the first time the effect of the organic loading rate (OLR) under oxygen limitation on biomass storage properties and kinetics. Polymer characteristics have been also evaluated. The selection/enrichment of PHA-storing biomass was successfully achieved in a Sequencing Batch Reactor (SBR) under short hydraulic retention time (HRT; 2 days). Low OLR (2.05 g COD/L d) was ideal for the selection of an efficient PHA-producing consortium cultivated under limited oxygen availability. In the fed-batch accumulation conducted under high DO regime, such biomass was characterized by 51% of PHA content on cell dry weight, with a related storage yield (YP/Sbatch) of 0.61 CODPHA/CODS. On the contrary, medium OLR (4.56 g COD/L d) was not technically feasible to sustain the required consortium's selection under low DO regime. The PHA produced by biomass cultivated under low DO regime was characterized higher thermal stability and crystalline domain compared to PHA traditionally produced under high DO regime. The mass balance assessment highlighted a global yield of 51 g PHA/kg VS (volatile solids of thickened sludge), which was 9% lower than yield obtained under high DO regime, in the face of a realistic reduction of the energy cost of the process.

Evidence for intraflagellar transport in butterfly spermatocyte cilia.

In the model organism insect Drosophila melanogaster short cilia assemble on spermatocytes that elaborate into 1.8 mm long flagella during spermatid differentiation. A unique feature of these cilia/flagella is their lack of dependence on intraflagellar transport (IFT) for their assembly. Here, we show that in the common butterfly Pieris brassicae, the spermatocyte cilia are exceptionally long: about 40 µm compared to less than 1 µm in Drosophila. By transmission electron microscopy, we show that P. brassicae spermatocytes display several features not found in melanogaster, including compelling evidence of IFT structures and features of motile cilia.

Producing volatile fatty acids and polyhydroxyalkanoates from foods by-products and waste: A review.

Dairy products, extra virgin olive oil, red and white wines are excellent food products, appreciated all around the world. Their productions generate large amounts of by-products which urge for recycling and valorization. Moreover, another abundant waste stream produced in urban context is the Organic Fraction of Municipal Solid Wastes (OFMSW), whose global annual capita production is estimated at 85 kg. The recent environmental policies encourage their exploitation in a biorefinery loop to produce Volatile Fatty Acids (VFAs) and polyhydroxyalkanoates (PHAs). Typically, VFAs yields are high from cheese whey and OFMSW (0.55-0.90 gCOD_VFAs/gCOD), lower for Olive Mill and Winery Wastewaters. The VFAs conversion into PHAs can achieve values in the range 0.4-0.5 gPHA/gVSS for cheese whey and OFMSW, 0.6-0.7 gPHA/gVSS for winery wastewater, and 0.2-0.3 gPHA/gVSS for olive mill wastewaters. These conversion yields allowed to estimate a huge potential annual PHAs production of about 260 M tons.

Cilium induction triggers differentiation of glioma stem cells.

Glioblastoma multiforme (GBM) possesses glioma stem cells (GSCs) that promote self-renewal, tumor propagation, and relapse. Understanding the mechanisms of GSCs self-renewal can offer targeted therapeutic interventions. However, insufficient knowledge of GSCs' fundamental biology is a significant bottleneck hindering these efforts. Here, we show that patient-derived GSCs recruit elevated levels of proteins that ensure the temporal cilium disassembly, leading to suppressed ciliogenesis. Depleting the cilia disassembly complex components is sufficient to induce ciliogenesis in a subset of GSCs via relocating platelet-derived growth factor receptor-alpha (PDGFR-α) to a newly induced cilium. Importantly, restoring ciliogenesis enabled GSCs to switch from self-renewal to differentiation. Finally, using an organoid-based glioma invasion assay and brain xenografts in mice, we establish that ciliogenesis-induced differentiation can prevent the infiltration of GSCs into the brain. Our findings illustrate a role for cilium as a molecular switch in determining GSCs' fate and suggest cilium induction as an attractive strategy to intervene in GSCs proliferation.

Human brain organoids assemble functionally integrated bilateral optic vesicles.

During embryogenesis, optic vesicles develop from the diencephalon via a multistep process of organogenesis. Using induced pluripotent stem cell (iPSC)-derived human brain organoids, we attempted to simplify the complexities and demonstrate formation of forebrain-associated bilateral optic vesicles, cellular diversity, and functionality. Around day 30, brain organoids attempt to assemble optic vesicles, which develop progressively as visible structures within 60 days. These optic vesicle-containing brain organoids (OVB-organoids) constitute a developing optic vesicle's cellular components, including primitive corneal epithelial and lens-like cells, retinal pigment epithelia, retinal progenitor cells, axon-like projections, and electrically active neuronal networks. OVB-organoids also display synapsin-1, CTIP-positive myelinated cortical neurons, and microglia. Interestingly, various light intensities could trigger photosensitive activity of OVB-organoids, and light sensitivities could be reset after transient photobleaching. Thus, brain organoids have the intrinsic ability to self-organize forebrain-associated primitive sensory structures in a topographically restricted manner and can allow interorgan interaction studies within a single organoid.

Effect of the temperature in a mixed culture pilot scale aerobic process for food waste and sewage sludge conversion into polyhydroxyalkanoates.

The utilisation of urban organic waste as feedstock for polyhydroxyalkanoates (PHA) production is growing since it allows to solve the main concerns about their disposal and simultaneously to recover added-value products. A pilot scale platform has been designed for this purpose. The VFA-rich fermentation liquid coming from the anaerobic treatment of both source-sorted organic fraction of municipal solid waste (OFMSW) and waste activated sludge (WAS) has been used as substrate for the aerobic process steps: a first sequencing batch reactor (SBR, 100 L) for the selection of a PHA-producing biomass, and a second fed-batch reactor (70 L) for PHA accumulation inside the cells. The SBR was operated at 2.0-4.4 kg COD/(m3 d) as OLR, under dynamic feeding regime (feast-famine) and short hydraulic retention time (HRT; 1 day). The selected biomass was able to accumulate up to 48% g PHA/g VSS. Both steps were performed without temperature (T) control, avoiding additional consumption of energy. In this regard, the applied OLR was tuned based on environmental T and, as a consequence, on biomass kinetic, in order to have a constant selective pressure. The latter was mainly quantified by the PHA storage yield (YP/Sfeast 0.34-0.45 CODP/CODS), which has been recognized as the main parameters affecting the global PHA productivity [1.02-1.82 g PHA/(L d)] of the process.

Dzip1 and Fam92 form a ciliary transition zone complex with cell type specific roles in Drosophila.

Cilia and flagella are conserved eukaryotic organelles essential for cellular signaling and motility. Cilia dysfunctions cause life-threatening ciliopathies, many of which are due to defects in the transition zone (TZ), a complex structure of the ciliary base. Therefore, understanding TZ assembly, which relies on ordered interactions of multiprotein modules, is of critical importance. Here, we show that Drosophila Dzip1 and Fam92 form a functional module which constrains the conserved core TZ protein, Cep290, to the ciliary base. We identify cell type specific roles of this functional module in two different tissues. While it is required for TZ assembly in all Drosophila ciliated cells, it also regulates basal-body growth and docking to the plasma membrane during spermatogenesis. We therefore demonstrate a novel regulatory role for Dzip1 and Fam92 in mediating membrane/basal-body interactions and show that these interactions exhibit cell type specific functions in basal-body maturation and TZ organization.

Drosophila Doublefault protein coordinates multiple events during male meiosis by controlling mRNA translation.

During the extended prophase of Drosophila gametogenesis, spermatocytes undergo robust gene transcription and store many transcripts in the cytoplasm in a repressed state, until translational activation of select mRNAs in later steps of spermatogenesis. Here, we characterize the Drosophila Doublefault (Dbf) protein as a C2H2 zinc-finger protein, primarily expressed in testes, that is required for normal meiotic division and spermiogenesis. Loss of Dbf causes premature centriole disengagement and affects spindle structure, chromosome segregation and cytokinesis. We show that Dbf interacts with the RNA-binding protein Syncrip/hnRNPQ, a key regulator of localized translation in Drosophila We propose that the pleiotropic effects of dbf loss-of-function mutants are associated with the requirement of dbf function for translation of specific transcripts in spermatocytes. In agreement with this hypothesis, Dbf protein binds cyclin B mRNA and is essential for translation of cyclin B in mature spermatocytes.

Plk1/Polo Phosphorylates Sas-4 at the Onset of Mitosis for an Efficient Recruitment of Pericentriolar Material to Centrosomes.

Centrosomes are the major microtubule-organizing centers, consisting of centrioles surrounded by a pericentriolar material (PCM). Centrosomal PCM is spatiotemporally regulated to be minimal during interphase and expands as cells enter mitosis. It is unclear how PCM expansion is initiated at the onset of mitosis. Here, we identify that, in Drosophila, Plk1/Polo kinase phosphorylates the conserved centrosomal protein Sas-4 in vitro. This phosphorylation appears to occur at the onset of mitosis, enabling Sas-4's localization to expand outward from meiotic and mitotic centrosomes. The Plk1/Polo kinase site of Sas-4 is then required for an efficient recruitment of Cnn and γ-tubulin, bona fide PCM proteins that are essential for PCM expansion and centrosome maturation. Point mutations at Plk1/Polo sites of Sas-4 affect neither centrosome structure nor centriole duplication but specifically reduce the affinity to bind Cnn and γ-tubulin. These observations identify Plk1/Polo kinase regulation of Sas-4 as essential for efficient PCM expansion.

The "transition zone" of the cilium-like regions in the Drosophila spermatocytes and the role of the C-tubule in axoneme assembly.

The fruit-fly Drosophila melanogaster harbours different types of ciliary structures: ciliary projections associated with neurons of type I and cilium-like regions (CLRs) found during male gametogenesis. The latter deserve particular attention since they are morphologically similar to vertebrate primary cilia and transform into the sperm axonemes during spermiogenesis. Although, all the centrioles are able to organize the CLRs, we found that the mother centriole docks first to the plasma membrane suggesting a new intrinsic functional asymmetry between the parent centrioles. We also show that the CLRs lack the Y-links that connect the axoneme doublets with the plasma membrane in conventional primary cilia. Moreover, the C-tubules, that are lacking in the axoneme of the primary cilia, persisted along the CLRs albeit modified into longitudinal blades. Remarkably, mutant flies in which the CLRs are devoid of the C-tubules or their number is reduced lack sperm axonemes or have incomplete axonemes. Therefore, the C-tubules are dispensable for the assembly of the CLRs but are essential for sperm axoneme elongation and maintenance in Drosophila.

Gorab is a Golgi protein required for structure and duplication of Drosophila centrioles.

We demonstrate that a Drosophila Golgi protein, Gorab, is present not only in the trans-Golgi but also in the centriole cartwheel where, complexed to Sas6, it is required for centriole duplication. In addition to centriole defects, flies lacking Gorab are uncoordinated due to defects in sensory cilia, which lose their nine-fold symmetry. We demonstrate the separation of centriole and Golgi functions of Drosophila Gorab in two ways: first, we have created Gorab variants that are unable to localize to trans-Golgi but can still rescue the centriole and cilia defects of gorab null flies; second, we show that expression of C-terminally tagged Gorab disrupts Golgi functions in cytokinesis of male meiosis, a dominant phenotype overcome by mutations preventing Golgi targeting. Our findings suggest that during animal evolution, a Golgi protein has arisen with a second, apparently independent, role in centriole duplication.

Centrobin is essential for C-tubule assembly and flagellum development in Drosophila melanogaster spermatogenesis.

Centrobin homologues identified in different species localize on daughter centrioles. In Drosophila melanogaster sensory neurons, Centrobin (referred to as CNB in Drosophila) inhibits basal body function. These data open the question of CNB's role in spermatocytes, where daughter and mother centrioles become basal bodies. In this study, we report that in these cells, CNB localizes equally to mother and daughter centrioles and is essential for C-tubules to attain the right position and remain attached to B-tubules as well as for centrioles to grow in length. CNB appears to be dispensable for meiosis, but flagellum development is severely compromised in Cnb mutant males. Remarkably, three N-terminal POLO phosphorylation sites that are critical for CNB function in neuroblasts are dispensable for spermatogenesis. Our results underpin the multifunctional nature of CNB that plays different roles in different cell types in Drosophila, and they identify CNB as an essential component for C-tubule assembly and flagellum development in Drosophila spermatogenesis.

Recent developments in biohythane production from household food wastes: A review.

Biohythane is a hydrogen-methane blend with hydrogen concentration between 10 and 30% v/v. It can be produced from different organic substrates by two sequential anaerobic stages: a dark fermentation step followed by a second an anaerobic digestion step, for hydrogen and methane production, respectively. The advantages of this blend compared to either hydrogen or methane, as separate biofuels, are first presented in this work. The two-stage anaerobic process and the main operative parameters are then discussed. Attention is focused on the production of biohythane from household food wastes, one of the most abundant organic substrate available for anaerobic digestion: the main milestones and the future trends are exposed. In particular, the possibility to co-digest food wastes and sewage sludge to improve the process yield is discussed. Finally, the paper illustrates the developments of biohythane application in the automotive sector as well as its reduced environmental burden.

The developing Drosophila eye - a new model to study centriole reduction.

In the developing Drosophila eye, the centrioles of the differentiating retinal cells are not surrounded by the microtubule-nucleating γ-tubulin, suggesting that they are unable to organize functional microtubule-organizing centers. Consistent with this idea, Cnn and Spd-2, which are involved in γ-tubulin recruitment, and the scaffold protein Plp, which plays a role in the organization of the pericentriolar material, are lost in the third-instar larval stage. However, the centrioles maintain their structural integrity, and both the parent centrioles accumulate Asl and Ana1. Although the loading of Asl points to the acquisition of the motherhood condition, the daughter centrioles fail to recruit Plk4 and do not duplicate. However, it is surprising that the mother centrioles that accumulate Plk4 also never duplicate. This suggests that the loading of Plk4 is not sufficient, in this system, to allow centriole duplication. By halfway through pupal life, the centriole number decreases and structural defects, ranging from being incomplete or lacking B-tubules, are detected. Asl, Ana1 and Sas-4 are still present, suggesting that the centriole integrity does not depend on these proteins.

Parthenogenesis in Insects: The Centriole Renaissance.

Building a new organism usually requires the contribution of two differently shaped haploid cells, the male and female gametes, each providing its genetic material to restore diploidy of the new born zygote. The successful execution of this process requires defined sequential steps that must be completed in space and time. Otherwise, development fails. Relevant among the earlier steps are pronuclear migration and formation of the first mitotic spindle that promote the mixing of parental chromosomes and the formation of the zygotic nucleus. A complex microtubule network ensures the proper execution of these processes. Instrumental to microtubule organization and bipolar spindle assembly is a distinct non-membranous organelle, the centrosome. Centrosome inheritance during fertilization is biparental, since both gametes provide essential components to build a functional centrosome. This model does not explain, however, centrosome formation during parthenogenetic development, a special mode of sexual reproduction in which the unfertilized egg develops without the contribution of the male gamete. Moreover, whereas fertilization is a relevant example in which the cells actively check the presence of only one centrosome, to avoid multipolar spindle formation, the development of parthenogenetic eggs is ensured, at least in insects, by the de novo assembly of multiple centrosomes.Here, we will focus our attention on the assembly of functional centrosomes following fertilization and during parthenogenetic development in insects. Parthenogenetic development in which unfertilized eggs are naturally depleted of centrosomes would provide a useful experimental system to investigate centriole assembly and duplication together with centrosome formation and maturation.

Feasibility of thermophilic anaerobic processes for treating waste activated sludge under low HRT and intermittent mixing.

Thermophilic anaerobic digestion (AD) arises as an optimized solution for the waste activated sludge (WAS) management. However, there are few feasibility studies using low solids content typically found in the WAS, and that consider uncommon operational conditions such as intermittent mixing and low hydraulic retention time (HRT). In this investigation, a single-stage pilot reactor was used to treat WAS at low HRT (13, 9, 6 and 5 days) and intermittent mixing (withholding mixing 2 h prior feeding). Thermophilic anaerobic digestion (55 °C) was initiated from a mesophilic digester (35 °C) by the one-step startup strategy. Although instabilities on partial alkalinity (1245-3000 mgCaCO3/L), volatile fatty acids (1774-6421 mg/L acetic acid) and biogas production (0.21-0.09 m3/m3reactor.d) were observed, methanogenesis started to recover in 18 days. The thermophilic treatment of WAS at 13 and 9 days HRT efficiently converted VS into biogas (22 and 21%, respectively) and achieved high biogas yield (0.24 and 0.22 m3/kgVSfed, respectively). Intermittent mixing improved the retention of methanogens inside the reactor and reduced the washout effect even at low HRT (<9 days). The negative thermal balance found was influenced by the low solids content in the WAS (2.1% TS) and by the heat losses from the digester walls. The energy balance and economic analyses demonstrated the feasibility of thermophilic AD of WAS in a hypothetical full-scale system, when the heat energy could be recovered from methane in a scenario of higher solids concentration in the substrate (>5% TS).

Rab1 interacts with GOLPH3 and controls Golgi structure and contractile ring constriction during cytokinesis in Drosophila melanogaster.

Cytokinesis requires a tight coordination between actomyosin ring constriction and new membrane addition along the ingressing cleavage furrow. However, the molecular mechanisms underlying vesicle trafficking to the equatorial site and how this process is coupled with the dynamics of the contractile apparatus are poorly defined. Here we provide evidence for the requirement of Rab1 during cleavage furrow ingression in cytokinesis. We demonstrate that the gene omelette (omt) encodes the Drosophila orthologue of human Rab1 and is required for successful cytokinesis in both mitotic and meiotic dividing cells of Drosophila melanogaster We show that Rab1 protein colocalizes with the conserved oligomeric Golgi (COG) complex Cog7 subunit and the phosphatidylinositol 4-phosphate effector GOLPH3 at the Golgi stacks. Analysis by transmission electron microscopy and 3D-SIM super-resolution microscopy reveals loss of normal Golgi architecture in omt mutant spermatocytes indicating a role for Rab1 in Golgi formation. In dividing cells, Rab1 enables stabilization and contraction of actomyosin rings. We further demonstrate that GTP-bound Rab1 directly interacts with GOLPH3 and controls its localization at the Golgi and at the cleavage site. We propose that Rab1, by associating with GOLPH3, controls membrane trafficking and contractile ring constriction during cytokinesis.

Recent Zika Virus Isolates Induce Premature Differentiation of Neural Progenitors in Human Brain Organoids.

The recent Zika virus (ZIKV) epidemic is associated with microcephaly in newborns. Although the connection between ZIKV and neurodevelopmental defects is widely recognized, the underlying mechanisms are poorly understood. Here we show that two recently isolated strains of ZIKV, an American strain from an infected fetal brain (FB-GWUH-2016) and a closely-related Asian strain (H/PF/2013), productively infect human iPSC-derived brain organoids. Both of these strains readily target to and replicate in proliferating ventricular zone (VZ) apical progenitors. The main phenotypic effect was premature differentiation of neural progenitors associated with centrosome perturbation, even during early stages of infection, leading to progenitor depletion, disruption of the VZ, impaired neurogenesis, and cortical thinning. The infection pattern and cellular outcome differ from those seen with the extensively passaged ZIKV strain MR766. The structural changes we see after infection with these more recently isolated viral strains closely resemble those seen in ZIKV-associated microcephaly.