Production of hydrogen from sewage biosolids in a continuously fed bioreactor: Effect of hydraulic retention time and sparging

Hydrogen was produced from primary sewage biosolids via mesophilic anaerobic fermentation in a continuously fed bioreactor. Prior to fermentation the sewage biosolids were heated to 70 °C for 1 h to inactivate methanogens and during fermentation a cellulose degrading enzyme was added to improve substrate availability. Hydraulic retention times (HRT) of 18, 24, 36 and 48 h were evaluated for the duration of hydrogen production. Without sparging a hydraulic retention time of 24 h resulted in the longest period of hydrogen production (3 days), during which a hydrogen yield of 21.9 L H₂ kg⁻¹ VS added to the bioreactor was achieved. Methods of preventing the decline of hydrogen production during continuous fermentation were evaluated. Of the techniques evaluated using nitrogen gas to sparge the bioreactor contents proved to be more effective than flushing just the headspace of the bioreactor. Sparging at 0.06 L L min⁻¹ successfully prevented a decline in hydrogen production and resulted in a yield of 27.0  L H₂ kg⁻¹ VS added, over a period of greater than 12 days or 12 HRT. The use of sparging also delayed the build up of acetic acid in the bioreactor, suggesting that it serves to inhibit homoacetogenesis and thus maintain hydrogen production.     

Use of coffee by‐products for the cultivation of Pleurotus citrinopileatus and Pleurotus salmoneo‐stramineus and its impact on biological properties of extracts thereof

Incorporating spent coffee grounds (SCGs), a by‐product from coffee brewing, in growth substrate of beneficial edible mushrooms is an approach that has to be further studied due to its potential positive outcomes: environmental impact mitigation, production costs reduction and beneficial impact on consumer health. Hence, cultivation of Pleurotus citrinopileatus and Pleurotus salmoneo‐stramineus was tested using SCG which enabled maximum production yield of P. citrinopileatus which was of 25.1% (w/w). Variable antidiabetic potential was observed between aqueous and enzymatic extracts (3.8%–29% inhibition) regardless species and substrates, whereas aqueous extract of P. citrinopileatus grown in substrate without SCG stood out presenting the highest antioxidant activity and inhibition activity of angiotensin I‐converting enzyme (IC₅₀ = 123 μg mL^?1). Ethanolic and aqueous extracts of both Pleurotus species grown in the presence or absence of SGC proved to be an interesting prebiotic source for growth of Bifidobacterium animalis Bo in comparison with fructooligosaccharides (FOS).     

Hydrothermal Carbonization of Sewage Sludge: Analysis of Process Severity and Solid Content

Hydrothermal carbonization of sewage sludge was carried out with the aim to evaluate the influence of process severity and initial solid content. Response surface methodology was applied to model yield and C yield responses. Enhanced dewaterability performance was recorded under mild processing conditions. The treatment promoted concentration and immobilization of Pb, Cd, Ni, Zn, and Cu. Variation of the solid content showed a stronger influence than severity on average yield and C yield. Higher heating values (HHVs) and energy retention efficiencies (EREs) of hydrochars obtained at the lowest solid content displayed the lowest values. Hence, the energy requirements of a first dewatering step should be compared with the related improvement in terms of HHV and ERE when sludge is used as feedstock.     

Engineering of Saccharomyces cerevisiae for the production of (+)-ambrein

The triterpenoid (+)-ambrein is the major component of ambergris, a coprolite of the sperm whale that can only be rarely found on shores. Upon oxidative degradation of (+)-ambrein, several fragrance molecules are formed, amongst them (−)-ambrox, one of the highest valued compounds in the perfume industry. In order to generate a Saccharomyces cerevisiae whole-cell biocatalyst for the production of (+)-ambrein, intracellular supply of the squalene was enhanced by overexpression of two central enzymes in the mevalonate and sterol biosynthesis pathway, namely the N-terminally truncated 3-hydroxy-3-methylglutaryl-CoA reductase 1 (tHMG) and the squalene synthase (ERG9). In addition, another key enzyme in sterol biosynthesis, squalene epoxidase (ERG1) was inhibited by an experimentally defined amount of the inhibitor terbinafine in order to reduce flux of squalene towards ergosterol biosynthesis while retaining sufficient activity to maintain cell viability and growth. Heterologous expression of a promiscuous variant of Bacillus megaterium tetraprenyl-β-curcumene cyclase (BmeTC-D373C), which has been shown to be able to catalyse the conversion of squalene to 3-deoxyachillol and then further to (+)-ambrein resulted in production of these triterpenoids in S. cerevisiae for the first time. Triterpenoid yields are comparable with the best microbial production chassis described in literature so far, the methylotrophic yeast Pichia pastoris. Consequently, we discuss similarities and differences of these two yeast species when applied for whole-cell (+)-ambrein production.     

Biological Characteristics and Osteogenic Differentiation of Ovine Bone Marrow Derived Mesenchymal Stem Cells Stimulated with FGF-2 and BMP-2

Cell-based therapies using mesenchymal stem cells (MSCs) are a promising tool in bone tissue engineering. Bone regeneration with MSCs involves a series of molecular processes leading to the activation of the osteoinductive cascade supported by bioactive factors, including fibroblast growth factor-2 (FGF-2) and bone morphogenetic protein-2 (BMP-2). In this study, we examined the biological characteristics and osteogenic differentiation potential of sheep bone marrow MSCs (BM-MSCs) treated with 20 ng/mL of FGF-2 and 100 ng/mL BMP-2 in vitro. The biological properties of osteogenic-induced BM-MSCs were investigated by assessing their morphology, proliferation, phenotype, and cytokine secretory profile. The osteogenic differentiation was characterized by Alizarin Red S staining, immunofluorescent staining of osteocalcin and collagen type I, and expression levels of genetic markers of osteogenesis. The results demonstrated that BM-MSCs treated with FGF-2 and BMP-2 maintained their primary MSC properties and improved their osteogenic differentiation capacity, as confirmed by increased expression of osteocalcin and collagen type I and upregulation of osteogenic-related gene markers BMP-2, Runx2, osterix, collagen type I, osteocalcin, and osteopontin. Furthermore, sheep BM-MSCs produced a variety of bioactive factors involved in osteogenesis, and supplementation of the culture medium with FGF-2 and BMP-2 affected the secretome profile of the cells. The results suggest that sheep osteogenic-induced BM-MSCs may be used as a cellular therapy to study bone repair in the preclinical large animal model.     

Intergenerational Metabolomic Analysis of Mothers with a History of Gestational Diabetes Mellitus and Their Offspring

Shared metabolomic patterns at delivery have been suggested to underlie the mother-to-child transmission of adverse metabolic health. This study aimed to investigate whether mothers with gestational diabetes mellitus (GDM) and their offspring show similar metabolomic patterns several years postpartum. Targeted metabolomics (including 137 metabolites) was performed in plasma samples obtained during an oral glucose tolerance test from 48 mothers with GDM and their offspring at a cross-sectional study visit 8 years after delivery. Partial Pearson’s correlations between the area under the curve (AUC) of maternal and offspring metabolites were calculated, yielding so-called Gaussian graphical models. Spearman’s correlations were applied to investigate correlations of body mass index (BMI), Matsuda insulin sensitivity index (ISI-M), dietary intake, and physical activity between generations, and correlations of metabolite AUCs with lifestyle variables. This study revealed that BMI, ISI-M, and the AUC of six metabolites (carnitine, taurine, proline, SM(-OH) C14:1, creatinine, and PC ae C34:3) were significantly correlated between mothers and offspring several years postpartum. Intergenerational metabolite correlations were independent of shared BMI, ISI-M, age, sex, and all other metabolites. Furthermore, creatinine was correlated with physical activity in mothers. This study suggests that there is long-term metabolic programming in the offspring of mothers with GDM and informs us about targets that could be addressed by future intervention studies.     

Solution Structure and Dynamics of the Small Protein HVO_2922 from Haloferax volcanii

Past sequencing campaigns overlooked small proteins as they seemed to be irrelevant due to their small size. However, their occurrence is widespread, and there is growing evidence that these small proteins are in fact functionally very important in organisms found in all kingdoms of life. Within a global proteome analysis for small proteins of the archaeal model organism Haloferax volcanii, the HVO_2922 protein has been identified. It is differentially expressed in response to changes in iron and salt concentrations, thus suggesting that its expression is stress-regulated. The protein is conserved among Haloarchaea and contains an uncharacterized domain of unknown function (DUF1508, UPF0339 family protein). We elucidated the NMR solution structure, which shows that the isolated protein forms a symmetrical dimer. The dimerization is found to be concentration-dependent and essential for protein stability and most likely for its functionality, as mutagenesis at the dimer interface leads to a decrease in stability and protein aggregation.     

A proactive and resilient seismic risk mitigation strategy for existing school buildings

Different methods and procedures have been developed to define prioritisation strategies of retrofit interventions aimed at reducing the seismic risk of school buildings on a large territorial scale. However, these approaches fail to demonstrate how risk analysis has been used successfully to quantitatively assess and select the optimal risk management decision. This article proposes innovative and useful metrics to measure the potential costs and benefits related to the prioritisation of retrofit intervention and the resilience of the analysed school system by directly integrating engineering, organisational, socio-economic and political aspects in the realm of seismic resilience assessment. Based on probabilistic risk assessments considering the new vulnerability of the prioritised school buildings, these measures could predict the expected economic and functional losses associated with a disastrous seismic event, as well as the possible post-disaster recovery of the system. In order to help decision-makers in selecting the optimal mitigation strategy with a multidisciplinary and multidimensional perspective, different political scenarios, the relative prioritisations of interventions and their intervention options are also defined. The proposed framework is demonstrated in a complex case study of 1,825 public schools in the Lima metropolitan area, Peru. Policymakers, planners and engineering professionals could benefit from results.     

Development and comparison of loop-mediated isothermal amplification and quantitative polymerase chain reaction assays for the detection of Mycoplasma bovis in milk

Bovine mastitis is an economic burden for dairies worldwide. Mycoplasma species, and especially Mycoplasma bovis, are among the most important causative agents, and rapid, precise, and low-cost methods for Mycoplasma detection are urgently needed. For this purpose, loop-mediated isothermal amplification (LAMP) and quantitative PCR (qPCR) assays were developed and compared. The LAMP assay was designed and primer concentrations optimized to M. bovis oppD, encoding oligopeptide permease D. For qPCR, a Taqman assay (Applied Biosystems, Carlsbad, CA) targeting M. bovis gltX, encoding glutamate transfer RNA ligase, was optimized for primer concentration, annealing temperature, and DNA polymerase. Both assays were similarly sensitive, with a detection limit of approximately 10⁴ to 10⁵ M. bovis cells/mL. Both assays were also successful in confirming M. bovis identity in laboratory culture suspensions and in bovine milk. The LAMP and qPCR assays combined with the MoBio DNA extraction kit (MoBio Laboratories Inc., Carlsbad, CA) resulted in the correct detection of 13 out of 13 M. bovis isolates and 14 out of 16 M. bovis-positive milk samples collected from commercial dairies in California. When combined with the PrepMan Ultra reagent (Applied Biosystems), the qPCR assay resulted in confirming 21 out of 21 M. bovis-positive milk samples. Comparison of the assays to milk containing either Mycoplasma arginini, Mycoplasma bovigenitalium, Mycoplasma californicum, M. alkalescens, or Acholeplasma laidlawii or milk lacking any detectable Mycoplasma species or relatives resulted in 3 out of 17 (LAMP with MoBio), 1 out of 17 (qPCR with MoBio), and 2 out of 36 (qPCR with PrepMan Ultra) false positives. Overall, the qPCR assay was more robust than LAMP and could be used on DNA recovered from milk prepared with the PrepMan Ultra reagent, a method that does not include a DNA purification step. The use of this qPCR method enables M. bovis detection in bovine milk in 40 to 55 min, and therefore provides new opportunities to accelerate and simplify M. bovis detection in unpasteurized milk to reduce the incidence of M. bovis mastitis outbreaks.