Extracellular non-coding RNA signatures of the metacestode stage of Echinococcus multilocularis

Extracellular RNAs (ex-RNAs) are secreted by cells through different means that may involve association with proteins, lipoproteins or extracellular vesicles (EV). In the context of parasitism, ex-RNAs represent new and exciting communication intermediaries with promising potential as novel biomarkers. In the last years, it was shown that helminth parasites secrete ex-RNAs, however, most work mainly focused on RNA secretion mediated by EV. Ex-RNA study is of special interest in those helminth infections that still lack biomarkers for early and/or follow-up diagnosis, such as echinococcosis, a neglected zoonotic disease caused by cestodes of the genus Echinococcus. In this work, we have characterised the ex-RNA profile secreted by in vitro grown metacestodes of Echinococcus multilocularis, the casuative agent of alveolar echinococcosis.We have used high throughput RNA-sequencing together with RT-qPCR to characterise the ex-RNA profile secreted towards the extra- and intra-parasite milieus in EV-enriched and EV-depleted fractions. We show that a polarized secretion of small RNAs takes place, with microRNAs mainly secreted to the extra-parasite milieu and rRNA- and tRNA-derived sequences mostly secreted to the intra-parasite milieu. In addition, we show by nanoparticle tracking analyses that viable metacestodes secrete EV mainly into the metacestode inner vesicular fluid (MVF); however, the number of nanoparticles in culture medium and MVF increases > 10-fold when metacestodes show signs of tegument impairment. Interestingly, we confirm the presence of host miRNAs in the intra-parasite milieu, implying their internalization and transport through the tegument towards the MVF. Finally, our assessment of the detection of Echinococcus miRNAs in patient samples by RT-qPCR yielded negative results suggesting the tested miRNAs may not be good biomarkers for this disease.A comprehensive study of the secretion mechanisms throughout the life cycle of these parasites will help to understand parasite interaction with the host and also, improve current diagnostic tools.     

Effect of coexistence of Penicillium frequentans (westling) with Globodera rostochiensis (Woll.) behrens on potato

The aim of two‐year‐long pot experiments was to investigate the influence of P. frequentans on the growth of over‐ground parts, the intensity of assimilation and transpiration of plants, as well as ths influence on the yield of potato bulbs. It was determined how the above fungus influenced the populaüon of G rostochiensis, the influence being characterized by the density change of cysts, eggs, and larvae in the soil. Experiments in vitro allowed to determine the wholesomeness of cysts, eggs, and larvae (J2) that were in direct contact with the mycelium ofP. frequentans.

It was stated that presence of P. frequentans intensified the development of plants and relevantly stimulated their growth, especially in the initial vegetation phase, as compared with the remaining combinations (Fig. 1). Positive effect of P. frequentans on the growth of potato yield was proved, in comparison with the control, and by co‐occurrence of the fungus with G. rostochiensis, also in comparison to the combination with nematodes only (Fig. 2).

Pot experiments confirmed the antagonistic effect of P. frequentans on the population of G. rostochiensis. Presence of P. frequentans in the ground caused namely a relevant diminution of density of cysts, eggs and larvae (Tab. II), as well as morphological changes by females, which was connected with size reduction of the cysts (Tab. III). Experiments in vitro confirmed the destructive effect of P. frequentans on the cysts of G. rostochiensis (Tab. IV).     

Co-localization of zinc transporter 3 (ZnT3) with sensory neuromediators and/or neuromodulators in the enteric nervous system of the porcine esophagus

Zinc transporter 3 (ZnT3) is one of the zinc transporters family. It is closely connected to the nervous system, where enables the transport of zinc ions from the cytoplasm to synaptic vesicles. This substance has been described within the central and peripheral nervous system, especially in the enteric nervous system (ENS). The aim of the present study was to describe the co-localization of ZnT3 with selected neuromediators and/or neuromodulators participating in sensory stimuli conduction in neurons of the ENS within the porcine esophagus. Co-localization of ZnT3 with substance P (SP), leucine enkephalin (LENK) and calcitonin gene-related peptide (CGRP) was studied using standard double-immunofluorescence technique. The obtained results show that ZnT3, SP and/or LENK may occur in the same enteric neurons, and the degree of co-localization of these substances clearly depends on the fragment of esophagus studied and the type of enteric ganglia. In contrast, the co-localization of ZnT3 with CGRP was not observed during the present investigation. The obtained results suggest that ZnT3 in the ENS may be involved in the conduction of sensory and/or pain stimuli.     

Melatonin, an underestimated natural substance with great potential for agricultural application

Melatonin (MEL) was thought to be only a neurotransmitter found in vertebrates until its detection in other organisms including plants. Although the number of publications on MEL function in plants is expanding, the knowledge of this subject is still insufficient. Among many functions which MEL performs in plants, its role as an antioxidant and a growth promoter is most supported by experimental evidence. This compound is an independent plant growth regulator and it may mediate the activities of other plant growth regulators. Due to its antioxidant properties MEL may also stabilize cell redox status and protect tissues against reactive oxygen and nitrogen species which accumulated under stressful environment. Some researchers propose that MEL could be used to improve the phytoremediation efficiency of plants against different pollutants. In this paper we show that exogenous MEL applied into the seeds could be a good biostimulator improving not only seed germination, seedling/plant growth but also crop production especially under stress conditions. We also believe that this compound can increase food quality (the aspect of functional food) and may improve human health. Since MEL is inexpensive and safe for animals and humans its application as a biostimulator could be a good, feasible and cost-effective method useful in agriculture.     

Effect of Differentiation and Cell Density on Glycosphingolipid Class and Molecular Species Composition of Mouse Neuroblastoma NB2a Cells

The effects of cell density and retinoic acid-induced differentiation on the class and molecular species composition of mouse neuroblastoma NB2a cell glycosphingolipids were examined under conditions where the period of culture was controlled. The total amount of neutral glycosphingolipids per cell decreased both with differentiation and as the cells became confluent. The relative amount of the neutral glycosphingolipid classes was not affected by differentiation, whereas there were small but significant changes in the relative amount of the neutral glycosphingolipid classes as the cells became confluent. The total amount of the gangliosides was unaffected by either differentiation or cell density, but there were significant changes in the ganglioside class composition as a result of both cell density and differentiation, and the effects were additive. The molecular species of all the major neutral glycosphingolipid and ganglioside classes were essentially identical, and were altered only slightly by either differentiation or cell density.     

Elimination of by-products in 11β-hydroxylation of Substance S using Curvularia lunata clones regenerated from NTG-treated protoplasts

Summary Stable mutants showing improved 11-hydroxylation of Substance S were isolated, following treatment with N-methyl-N-nitro-N-nitrosuguanidine (NTG) and regeneration of uninucleate protoplasts of the appropriate fungal strains. This procedure was especially suitable for obtaining more directed 11-hydroxylation of Substance S with Curvularia lunata IM 2901. Apart from producing cortisol (11-hydroxy-S), the parent strain formed several by-products that significantly lowered the yield of the desired 11-hydroxyderivative. Isolated mutants of this microoraganism carried out directed 11-hydroxylation with only a small amount of one of the by-products, which resulted in a much higher yield of cortisol. Correspondence to: L. Sedlaczek     

Effects of zebra mussels on cladoceran communities under eutrophic conditions

The purpose of this study was to determine how zebra mussels affected cladoceran community structure under eutrophic conditions. We conducted a mesocosm study where we manipulated the presence of zebra mussels and the presence of large-bodied Daphnia (Daphnia magna and Daphnia pulicaria). We also conducted a complimentary life-table experiment to determine how water from the zebra mussel treatment affected the life history characteristics of the cladoceran species. We anticipated that small- and large-bodied cladoceran species would respond differently to changes in algal quality and quantity under the effects of zebra mussels. Large-bodied Daphnia successfully established in the zebra mussel treatment but failed to grow in the control. We did not observe positive relationships between food concentrations and cladoceran abundances. However, the phosphorus content in the seston indicated that food quality was below the threshold level for large-bodied cladocerans at the beginning of the experiment. We believe that zebra mussels quickly enhanced the phosphorus content in the seston due to the excretion of inorganic phosphorus, thus facilitating the development of large-bodied Daphnia. In conclusion, our results suggest that zebra mussels can alter the phosphorus content of seston in lakes and this can affect the dynamics of crustacean zooplankton.     

Molecular basis of diseases caused by the mtDNA mutation m.8969G>A in the subunit a of ATP synthase

The ATP synthase which provides aerobic eukaryotes with ATP, organizes into a membrane-extrinsic catalytic domain, where ATP is generated, and a membrane-embedded F_O domain that shuttles protons across the membrane. We previously identified a mutation in the mitochondrial MT-ATP6 gene (m.8969G>A) in a 14-year-old Chinese female who developed an isolated nephropathy followed by brain and muscle problems. This mutation replaces a highly conserved serine residue into asparagine at amino acid position 148 of the membrane-embedded subunit a of ATP synthase. We showed that an equivalent of this mutation in yeast (aS₁₇₅N) prevents F_O-mediated proton translocation. Herein we identified four first-site intragenic suppressors (aN₁₇₅D, aN₁₇₅K, aN₁₇₅I, and aN₁₇₅T), which, in light of a recently published atomic structure of yeast F_O indicates that the detrimental consequences of the original mutation result from the establishment of hydrogen bonds between aN₁₇₅ and a nearby glutamate residue (aE₁₇₂) that was proposed to be critical for the exit of protons from the ATP synthase towards the mitochondrial matrix. Interestingly also, we found that the aS₁₇₅N mutation can be suppressed by second-site suppressors (aP₁₂S, aI₁₇₁F, aI₁₇₁N, aI₂₃₉F, and aI₂₀₀M), of which some are very distantly located (by 20–30?Å) from the original mutation. The possibility to compensate through long-range effects the aS₁₇₅N mutation is an interesting observation that holds promise for the development of therapeutic molecules.