Search

Background: Biomarkers with strong predictive capacity towards transplantation outcome for livers undergoing normothermic machine perfusion (NMP) are needed. We investigated lactate clearing capacity as a basic function of liver viability during the first 6 h of NMP. Methods: A trial conducted in 6 high-volume transplant centres in Europe. All centres applied a back-to-base NMP approach with the OrganOx metra system. Perfusate lactate levels at start, 1, 2, 4 and 6 h of NMP were assessed individually and as area under the curve (AUC) and correlated with EAD (early allograft dysfunction), MEAF (model for early allograft function) and modified L-GrAFT (liver graft assessment following transplantation) scores. Results: A total of 509 livers underwent ≥6 h of NMP before transplantation in 6 centres in the UK, Germany and Austria. The donor age was 53 (40–63) years (median, i.q.r.). The total NMP time was 10.8 (7.9–15.7) h. EAD occurred in 26%, MEAF was 4.72 (3.54–6.05) and L-GrAFT10 −0.96 (−1.52–−0.32). Lactate at 1, 2 and 6 h correlated with increasing robustness with MEAF. Rather than a binary assessment with a cut-off value at 2 h, the actual 2 h lactate level correlated with the MEAF (P = 0.0306 versus P = 0.0002, Pearson r = 0.01087 versus r = 0.1734). The absolute lactate concentration at 6 h, the AUC of 0–6 h and 1–6 h (P < 0.0001, r = 0.3176) were the strongest predictors of MEAF. Conclusion: Lactate measured 1–6 h and lactate levels at 6 h correlate strongly with risk of liver allograft dysfunction upon transplantation. The robustness of predicting MEAF by lactate increases with perfusion duration. Monitoring lactate levels should be extended to at least 6 h of NMP routinely to improve clinical outcome. Biomarkers with strong predictive capacity towards transplantation outcome for livers undergoing normothermic machine perfusion are needed. We investigated lactate clearing capacity as a basic function of liver viability during the first 6 hours of NMP in a multicentre trial in 509 livers and found the absolute lactate concentration at 6h, the AUC of 0-6h and 1-6h were the strongest predictors of clinical outcome. Monitoring lactate levels should be extended to at least 6h of NMP routinely to improve clinical outcome. [ABSTRACT FROM AUTHOR]

During the last decades, hypervirulent strains of Clostridioides difficile with frequent disease recurrence and increased mortality appeared. Clostridioides difficile DSM 101085 was isolated from a patient who suffered from several recurrent infections and colonizations, likely contributing to a fatal outcome. Analysis of the toxin repertoire revealed the presence of a complete binary toxin locus and an atypical pathogenicity locus consisting of only a tcdA pseudogene and a disrupted tcdC gene sequence. The pathogenicity locus shows upstream a transposon and has been subject to homologous recombination or lateral gene transfer events. Matching the results of the genome analysis, neither TcdA nor TcdB production but the expression of cdtA and cdtB was detected. This highlights a potential role of the binary toxin C. difficile toxin in this recurrent colonization and possibly further in a host-dependent virulence. Compared with the C. difficile metabolic model strains DSM 28645 (630 Δerm) and DSM 27147 (R20291), strain DSM 101085 showed a specific metabolic profile, featuring changes in the threonine degradation pathways and alterations in the central carbon metabolism. Moreover, products originating from Stickland pathways processing leucine, aromatic amino acids, and methionine were more abundant in strain DSM 101085, indicating a more efficient use of these substrates. The particular characteristics of strain C. difficile DSM 101085 may represent an adaptation to a low-protein diet in a patient with recurrent infections. [ABSTRACT FROM AUTHOR]

Plant-parasitic cyst nematodes infect plants and form highly sophisticated feeding sites in roots. It is not known which plant cell signalling mechanisms trigger plant defence during the early stages of nematode parasitism. Mitogen-activated protein kinases (MAPKs) are central components of protein phosphorylation cascades transducing extracellular signals to plant defence responses. MAPK phosphatases control kinase activities and the signalling outcome. The involvement and the role of MPK3 and MPK6, as well as the MAPK phosphatase AP2C1, is demonstrated during parasitism of the beet cyst nematode Heterodera schachtii in Arabidopsis. Our data reveal notable activation patterns of plant MAPKs and the induction of AP2C1 suggesting the attenuation of defence signalling in plant cells during early nematode infection. It is demonstrated that the ap2c1 mutant that is lacking AP2C1 is more attractive but less susceptible to nematodes compared with the AP2C1-overexpressing line. This implies that the function of AP2C1 is a negative regulator of nematode-induced defence. By contrast, the enhanced susceptibility of mpk3 and mpk6 plants indicates a positive role of stress-activated MAPKs in plant immunity against nematodes. Evidence is provided that phosphatase AP2C1, as well as AP2C1-targeted MPK3 and MPK6, are important regulators of plant-nematode interaction, where the co-ordinated action of these signalling components ensures the timely activation of plant defence. [ABSTRACT FROM AUTHOR]

Above- and belowground plant parts are simultaneously attacked by different pests and pathogens. The host mediates these interactions and physiologically reacts, e.g. with local and systemic alterations of endogenous hormone levels coupled with coordinated transcriptional changes. This in turn affects attractiveness and susceptibility of the plant to subsequent attackers. Here, the model plant Arabidopsis thaliana is used to study stress hormone-based systemic responses triggered by simultaneous root parasitism by the cyst nematode Heterodera schachtii and shoot herbivory by the thrips Frankliniella occidentalis and the spider mite Tetranychus urticae. First, HPLC/MS and quantitative reverse transcriptase PCR are used to show that nematode parasitism strongly affects stress hormone levels and expression of hormone marker genes in shoots. Previous nematode infection is then demonstrated to affect the behavioural and life history performance of both arthropods. While thrips explicitly avoid nematode-infected plants, spider mites prefer them. In addition, the life history performance of T. urticae is significantly enhanced by nematode infection. Finally, systemic changes triggered by shoot-feeding F. occidentalis but not T. urticae are shown to make the roots more attractive for H. schachtii. This work emphasises the importance of above- and belowground signalling and contributes to a better understanding of plant systemic defence mechanisms against plant-parasitic nematodes. [ABSTRACT FROM AUTHOR]

Changes in the gene expression of sucrose synthases and invertases affected the local and systemic plant metabolism and communication, source–sink relationships, thus nutrition and development of sedentary endo-parasitic nematodesSedentary endoparasitic nematodes of plants induce highly specific feeding cells in the root central cylinder. From these, the obligate parasites withdraw all required nutrients. The feeding cells were described as sink tissues in the plant’s circulation system that are supplied with phloem-derived solutes such as sugars. Currently, there are several publications describing mechanisms of sugar import into the feeding cells. However, sugar processing has not been studied so far. Thus, in the present work, the roles of the sucrose-cleaving enzymes sucrose synthases (SUS) and invertases (INV) in the development of Heterodera schachtii were studied. Gene expression analyses indicate that both enzymes are regulated transcriptionally. Nematode development was enhanced on multiple INV and SUS mutants. Syncytia of these mutants were characterized by altered enzyme activity and changing sugar pool sizes. Further, the analyses revealed systemically affected sugar levels and enzyme activities in the shoots of the tested mutants, suggesting changes in the source–sink relationship. Finally, the development of the root-knot nematode Meloidogyne javanica was studied in different INV and SUS mutants and wild-type Arabidopsis plants. Similar effects on the development of both sedentary endoparasitic nematode species (root-knot and cyst nematode) were observed, suggesting a more general role of sucrose-degrading enzymes during plant–nematode interactions. [ABSTRACT FROM PUBLISHER]

The plant-parasitic nematode Heterodera schachtii stimulates plant root cells to form syncytial feeding structures which synthesize all nutrients required for successful nematode development. Cellular re-arrangements and modified metabolism of the syncytia are accompanied by massive intra- and intercellular solute allocations. In this study the expression of all genes annotated as sugar transporters in the Arabidopsis Membrane Protein Library was investigated by Affymetrix gene chip analysis in young and fully developed syncytia compared with non-infected Arabidopsis thaliana roots. The expression of three highly up-regulated (STP12, MEX1, and GTP2) and three highly down-regulated genes (SFP1, STP7, and STP4) was analysed by quantitative RT-PCR (qRT-PCR). The most up-regulated gene (STP12) was chosen for further in-depth studies using in situ RT-PCR and a nematode development assay with a T-DNA insertion line revealing a significant reduction of male nematode development. The specific role of STP12 expression in syncytia of male juveniles compared with those of female juveniles was further shown by qRT-PCR. In order to provide evidence for sugar transporter activity across the plasma membrane of syncytia, fluorescence-labelled glucose was used and membrane potential recordings following the application of several sugars were performed. Analyses of soluble sugar pools revealed a highly specific composition in syncytia. The presented work demonstrates that sugar transporters are specifically expressed and active in syncytia, indicating a profound role in inter- and intracelluar transport processes. [ABSTRACT FROM PUBLISHER]

The plant parasitic nematode Heterodera schachtii induces specific syncytial feeding sites in the roots of Arabidopsis thaliana from where it withdraws all required nutrients. Therefore, syncytia have to be well supplied with assimilates and generate strong sinks in the host plant's transport system. Import mechanisms and consequent accumulation of sucrose in syncytia were described recently. In this work, we studied the starch metabolism of syncytia. Using high-performance liquid chromatography and microscopic analyses, we demonstrated that syncytia store carbohydrates by starch accumulation. Further, we monitored the expression of genes involved in the starch metabolic pathway by gene chip analysis and quantitative reverse transcription-PCR. Finally, we provide functional proof of the importance of starch synthesis for nematode development using T-DNA insertion lines. We conclude that syncytia accumulate starch as a carbohydrate buffer to compensate for changing solute uptake by the nematode and as long-term storage during juvenile development. [ABSTRACT FROM AUTHOR]

The plant parasitic nematode Heterodera schachtii induces syncytial feeding structures in the roots of host plants. Nematode-induced syncytia become strong sink tissues in the plant solute circulation system as the parasites start withdrawing nutrients. In the present work, the expression pattern of the phloem-specific sucrose transporter AtSUC4 (also described as AtSUT4) is analysed in syncytia induced by H. schachtii and it is compared with that of AtSUC2, another phloem-specific sucrose transporter, which is expressed in syncytia. The temporal expression pattern was monitored by GUS-tests and real-time RT-PCR, while the localization within the syncytia was performed using in situ RT-PCR. In this context, the concentration of sucrose in infection sites was also analysed and, in fact, an increase in response to syncytium development was found. Silencing of the AtSUC4 gene finally resulted in a significant reduction of female nematode development, thus demonstrating a function for this gene for the first time. It is therefore concluded that AtSUC4 plays a significant role in the early phase of syncytium differentiation when functional plasmodesmata to the phloem are not yet established. It is further concluded that, during syncytium establishment, transporters are responsible for sucrose supply and, at a later stage, when a connection to the phloem is established via plasmodesmata, transporters are required for sucrose retrieval. [ABSTRACT FROM PUBLISHER]