Glutathione (GSH) and GSH-dependent enzymes play a key role in cellular detoxification processes that enable organism to cope with various internal and environmental stressors. However, it is often not clear, which components of the complex GSH-metabolism are required for tolerance towards a certain stressor. To address this question, a small scale RNAi-screen was carried out in Caenorhabditis elegans where GSH-related genes were systematically knocked down and worms were subsequently analysed for their survival rate under sub-lethal concentrations of arsenite and the redox cycler juglone. While the knockdown of γ-glutamylcysteine synthetase led to a diminished survival rate under arsenite stress conditions, GSR-1 (glutathione reductase) was shown to be essential for survival under juglone stress conditions. gsr-1 is the sole GSR encoding gene found in C. elegans. Knockdown of GSR-1 hardly affected total glutathione levels nor reduced glutathione/glutathione disulphide (GSH/GSSG) ratio under normal laboratory conditions. Nevertheless, when GSSG recycling was impaired by gsr-1(RNAi), GSH synthesis was induced, but not vice versa. Moreover, the impact of GSSG recycling was potentiated under oxidative stress conditions, explaining the enormous effect gsr-1(RNAi) knockdown had on juglone tolerance. Accordingly, overexpression of GSR-1 was capable of increasing stress tolerance. Furthermore, expression levels of SKN-1-regulated GSR-1 also affected life span of C. elegans, emphasising the crucial role the GSH redox state plays in both processes.

Cryptosporidium parvum is one of the most common human parasitic protozoa and is responsible for many waterborne outbreaks in several industrialized countries. The oocyst, which is the infective form, is known to be highly resistant to wastewater treatment procedures and represents a potential hazard to human populations through contaminated raw or treated wastewater. In this investigation, the occurrence of Cryptosporidium in wastewater samples was monitored and removal efficiency was assessed. Treated (effluent) and untreated (influent) wastewater samples were collected seasonally over a period of 2 years. Oocysts were repeatedly detected in influent and effluent samples collected from the treatment plant during all sampling seasons, with a mean concentration of 782 oocysts/L. The seasonal distribution showed that oocysts are predominant during autumn and winter. Molecular analyses via the small (18S) subunit of rRNA amplification and subsequent sequencing with an objective of characterizing the oocysts revealed that Cryptosporidium parvum was the dominant Cryptosporidium parasite present in wastewater.

Pyrimidines are important metabolites in all cells. Levels of cellular pyrimidines are controlled by multiple mechanisms, with one of these comprising the reductive degradation pathway. In the model invertebrate Caenorhabditis elegans, two of the three enzymes of reductive pyrimidine degradation have previously been characterized. The enzyme catalysing the final step of pyrimidine breakdown, 3‐ureidopropionase (β‐alanine synthase), had only been identified based on homology. We therefore cloned and functionally expressed the 3‐ureidopropionase of C. elegans as hexahistidine fusion protein. The purified recombinant enzyme readily converted the two pyrimidine degradation products: 3‐ureidopropionate and 2‐methyl‐3‐ureidopropionate. The enzyme showed a broad pH optimum between pH 7.0 and 8.0. Activity was highest at approximately 40 °C, although the half‐life of activity was only 65 s at that temperature. The enzyme showed clear Michaelis–Menten kinetics, with a Km of 147 ± 26 μm and a Vmax of 1.1 ± 0.1 U·mg protein−1. The quaternary structure of the recombinant enzyme was shown to correspond to a dodecamer by ‘blue native’ gel electrophoresis and gel filtration. The organ specific and subcellular localization of the enzyme was determined using a translational fusion to green fluorescent protein and high expression was observed in striated muscle cells. With the characterization of the 3‐ureidopropionase, the reductive pyrimidine degradation pathway in C. elegans has been functionally characterized. Structured digital abstract  http://mint.bio.uniroma2.it/mint/search/interaction.do?interactionAc=MINT-7986015: 3‐ureidopropionase (uniprotkb:http://www.uniprot.org/uniprot/Q19437) and 3‐ureidopropionase (uniprotkb:http://www.uniprot.org/uniprot/Q19437) bind (http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0407) by blue native page (http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0276)