Nitrification–denitrification is an environmentally friendly and cost-effective way to treat landfill leachates. Special attention has been given to the nitrification step, usually the limiting one due to its special sensitivity to environmental factors. Here, the effect of the acclimatization of the nitrifying biomass to two different intermediate landfill leachates with different salt concentrations, COD and BOD₅ has been studied. Despite the complete nitrification being successfully performed, the specific nitritation rates were reduced after the biomass adaptation to both landfill leachates caused by the presence of heavy metals and the high salt concentration. NGS analysis of the biomass samples revealed that Proteobacteria (48.5%), Actinobacteriota (14.4%) and Chloroflexi (9.5%) were the dominant phyla in the non-adapted biomass. The leachate feeding led to a decrease in OTU diversity and favored the growth of the phyla Bacteroidetes (27.2%), Euryarchaeota (26.6%) and Proteobacteria (20.0%) accounting for more than 70% of relative abundance. Several OTUs capable of performing the nitritation belong to the Xanthobacteraceae and the Xanthomonadaceae families, the Saccharimonadales order, and the genus Nitrosomonas, Nitrosospira and Paracoccus. In the nitratation process, the Xanthobacteraceae family and Lautropia and Nitrolancea genera were found. Graphical Abstract

Earth’s climate is warming as a result of anthropogenic emissions of greenhouse gases from fossil fuel combustion. Bioenergy, which includes biodiesel, biohydrogen and bioethanol, has emerged as a sustainable alternative fuel source. For this reason, in recent years biodiesel production has become widespread but this industry currently generates a huge amount of glycerol as a by-product, which has become an environmental problem in its own right. A feasible possibility to solve this problem is the use of waste glycerol as a carbon source for microbial transformation into biofuels such as hydrogen and ethanol. For instance, Escherichia coli is a microorganism that can synthesize these compounds under anaerobic conditions. In this work an experimental procedure was established for screening E. coli single mutants to identify strains with enhanced ethanol and/or H2 productions compared to the wild type strain. In an initial screening of 150 single mutants, 12 novel strains (gnd, tdcE, rpiA nanE, tdcB, deoB, sucB, cpsG, frmA, glgC, fumA and gadB) were found to provide enhanced yields for at least one of the target products. The mutations, that improve most significantly the parameters evaluated (gnd and tdcE genes), were combined with other mutations in three engineered E. coli mutant strains in order to further redirect carbon flux towards the desired products. This methodology can be a useful tool to disclose the metabolic pathways that are more susceptible to manipulation in order to obtain higher molar yields of hydrogen and ethanol using glycerol as main carbon source in multiple E. coli mutants.

Benzohydroxamic acids, such as 4-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one (D-DIBOA), exhibit interesting herbicidal, fungicidal and bactericidal properties. Recently, the chemical synthesis of D-DIBOA has been simplified to only two steps. In a previous paper, we demonstrated that the second step could be replaced by a biotransformation using Escherichia coli to reduce the nitro group of the precursor, ethyl 2-(2′-nitrophenoxy)acetate and obtain D-DIBOA. The NfsA and NfsB nitroreductases and the NemA xenobiotic reductase of E. coli have the capacity to reduce one or two nitro groups from a wide variety of nitroaromatic compounds, which are similar to the precursor. By this reason, we hypothesised that these three enzymes could be involved in this biotransformation. We have analysed the biotransformation yield (BY) of mutant strains in which one, two or three of these genes were knocked out, showing that only in the double nfsA/nfsB and in the triple nfsA/nfsB/nemA mutants, the BY was 0%. These results suggested that NfsA and NfsB are responsible for the biotransformation in the tested conditions. To confirm this, the nfsA and nfsB open reading frames were cloned into the pBAD expression vector and transformed into the nfsA and nfsB single mutants, respectively. In both cases, the biotransformation capacity of the strains was recovered (6.09 ± 0.06% as in the wild-type strain) and incremented considerably when NfsA and NfsB were overexpressed (40.33% ± 9.42% and 59.68% ± 2.0% respectively).