Evaluation of acidogenesis products’ effect on biogas production performed with metagenomics and isotopic approaches

Background: During the acetogenic step of anaerobic digestion, the products of acidogenesis are oxidized to substrates for methanogenesis: hydrogen, carbon dioxide and acetate. Acetogenesis and methanogenesis are highly interconnected processes due to the syntrophic associations between acetogenic bacteria and hydrogenotrophic methanogens, allowing the whole process to become thermodynamically favorable. The aim of this study is to determine the influence of the dominant acidic products on the metabolic pathways of methane formation and to find a core microbiome and substrate-specific species in a mixed biogas-producing system. Results: Four methane-producing microbial communities were fed with artificial media having one dominant component, respectively, lactate, butyrate, propionate and acetate, for 896 days in 3.5-L Up-flow Anaerobic Sludge Blanket (UASB) bioreactors. All the microbial communities showed moderately different methane production and utilization of the substrates. Analyses of stable carbon isotope composition of the fermentation gas and the substrates showed differences in average values of δ13C(CH4) and δ13C(CO2) revealing that acetate and lactate strongly favored the acetotrophic pathway, while butyrate and propionate favored the hydrogenotrophic pathway of methane formation. Genome-centric metagenomic analysis recovered 234 Metagenome Assembled Genomes (MAGs), including 31 archaeal and 203 bacterial species, mostly unknown and uncultivable. MAGs accounted for 54%–67% of the entire microbial community (depending on the bioreactor) and evidenced that the microbiome is extremely complex in terms of the number of species. The core microbiome was composed of Methanothrix soehngenii (the most abundant), Methanoculleus sp., unknown Bacteroidales and Spirochaetaceae. Relative abundance analysis of all the samples revealed microbes having substrate preferences. Substrate-specific species were mostly unknown and not predominant in the microbial communities. Conclusions: In this experimental system, the dominant fermentation products subjected to methanogenesis moderately modified the final effect of bioreactor performance. At the molecular level, a different contribution of acetotrophic and hydrogenotrophic pathways for methane production, a very high level of new species recovered, and a moderate variability in microbial composition depending on substrate availability were evidenced. Propionate was not a factor ceasing methane production. All these findings are relevant because lactate, acetate, propionate and butyrate are the universal products of acidogenesis, regardless of feedstock

Functional Characterization of TetR-like Transcriptional Regulator PA3973 from Pseudomonas aeruginosa

Pseudomonas aeruginosa, a human opportunistic pathogen, is a common cause of nosocomial infections. Its ability to survive under different conditions relies on a complex regulatory network engaging transcriptional regulators controlling metabolic pathways and capabilities to efficiently use the available resources. P. aeruginosa PA3973 encodes an uncharacterized TetR family transcriptional regulator. In this study, we applied a transcriptome profiling (RNA-seq), genome-wide identification of binding sites using ChIP-seq, as well as the phenotype analyses to unravel the biological role of PA3973. Transcriptional profiling of P. aeruginosa PAO1161 overexpressing PA3973 showed changes in the mRNA level of 648 genes. Concomitantly, ChIP-seq analysis identified more than 300 PA3973 binding sites in the P. aeruginosa genome. A 13 bp sequence motif was indicated as the binding site of PA3973. The PA3973 regulon encompasses the PA3972-PA3971 genes encoding a probable acyl-CoA dehydrogenase and a thioesterase. In vitro analysis showed PA3973 binding to PA3973p. Accordingly, the lack of PA3973 triggered increased expression of PA3972 and PA3971. The DPA3972-71 PAO1161 strain demonstrated impaired growth in the presence of stress-inducing agents hydroxylamine or hydroxyurea, thus suggesting the role of PA3972-71 in pathogen survival upon stress. Overall our results showed that TetR-type transcriptional regulator PA3973 has multiple binding sites in the P. aeruginosa genome and influences the expression of diverse genes, including PA3972-PA3971, encoding proteins with a proposed role in stress response

Wielopierścieniowe węglowodory aromatyczne w kurzu mieszkań warszawskich

The content of polycyclic aromatic hydrocarbons (PAHs) in the dust from Warsaw apartments were studied. Samples were collected from 48 flats, from a few places in each flat: living room (floor, shelves), bedroom (bed), kitchen, lavatory. At the same time a survey by questionnaire was carried out, in which outside and internat factors were described. PAHs concentration in samples were determined by reversed-phase high performance liquid chromatography (HPLC). The levels of PAHs concentration varied between 8 and 173 mg/kg. The content of PAHs in dust from most of Warsaw apartments was high (from 5 to 50 ppm), and in more than ten was extremely high (> 50 ppm). It could be one of the sources of health hazards in home environment. In the greater part of analyzed samples, very high levels of the following hydrocarbons have been found: benzo[b]fluoranthene, fluoranthene, pyrene, fhiorene and phenanthrene. Among microenvironments, selected for the study, the sum of PAHs in kitchen and cellars has been two-three times higher as compared with that in other places. The results show a clear trend to an increased concentration of PAHs in apartment of tobacco smokers.Badano poziom zanieczyszczenia wielopierścieniowymi węglowodorami aromatycznymi (WWA[PAHs]) kurzu z mieszkań warszawskich. Próbki pobrano z 48 mieszkań, z kilku miejsc w każdym mieszkaniu: pokój dzienny (podłoga, półki), sypialnia (łóżko), kuchnia, łazienka. Równocześnie przeprowadzano ankietę, w której w kilku stopniowej skali oceniano różne czynniki zewnętrzne i wewnętrzne. Stężenie WWA w próbkach oznaczono metodą chromatografii cieczowej (HPLC). Uzyskane zakresy stężeń dla sumy szesnastu WWA, wahają się od 8 do 173 mg/kg. Wartości stężeń sumy WWA w większości próbek były duże, w granicach od 5 do 50 ppm, co może stanowić duże zagrożenie dla zdrowia. W kilkunastu próbkach stężenia WWA osiągały wartości bardzo duże. W większości próbek stwierdzono wyjątkowo duże stężenia takich węglowodorów, jak: benzo[b]fluoranten, fluoranten, piren, fluoren i fenantren. W wytypowanych do badań mikrośrodowiskach stwierdzono dwu-trzykrotnie większe stężenie sumy szesnastu WWA w kuchniach i piwnicach niż w pozostałych miejscach. Porównanie zawartości węglowodorów w kurzu ze zwaloryzowanymi danymi z ankiety wskazuje na wyraźną tendencję większego stężenia WWA w mieszkaniach osób palących

First ancient DNA sequences of the Late Pleistocene red deer (Cervus elaphus) from the Crimea, Ukraine

The Emine-Bair-Khosar Cave (EBK), situated on the northern edge of the Lower Plateau of the Chatyrdag Massif (Crimean Mountains) is rich in palaeontological material accumulated over most of the Pleistocene. mtDNA was isolated from bones of three specimens of red deer (Cervus elaphus) dated to the late Pleistocene (MIS 3). These are the first ancient DNA sequences obtained for this species. The position of the three red deer individuals on the phylogeographic tree is based on mtDNA sequences of contemporary representatives of the Cervinae inhabiting the Northern Hemisphere. The results confirm the notion that the Crimean Peninsula was the north-easternmost refugium in Europe, and that during and after the Late Pleistocene it played a major role in recolonisation and dispersal of temperate species in the whole Eurasian continent

Steinernema kraussei (Steiner, 1923) (Rhabditida: Steinernematidae) – the first record from Poland

This study reports the first record of Steinernema kraussei from Poland. The nematode was isolated from coniferous woodlands in 4 localities in central Poland. Preliminary identification of the species was done based on morphometric measurements. To confirm nematode species of the genus Steinernema the result was supported by the description of the ITS regio

Effect of heparan sulfate and gold nanoparticles on muscle development during embryogenesis

Marlena Zielinska1,2, Ewa Sawosz1, Marta Grodzik1, Mateusz Wierzbicki1, Maria Gromadka1, Anna Hotowy3, Filip Sawosz3, Andrzej Lozicki1, Andrè Chwalibog31Division of Biotechnology and Biochemistry of Nutrition, Warsaw University of Life sciences, Warsaw, 2The Kielanowski Institute of Animal Physiology and Nutrition, Jablonna, Poland; 3Department of Basic Animal and Veterinary sciences, University of copenhagen, Frederiksberg, DenmarkPurpose: It was hypothesized that heparan sulfate (HS) as an essential compound for myogenesis and nanoparticles of gold (nano-Au) as highly reactive compounds can affect muscle development as a consequence of molecular regulation of muscle cell formation, and that these effects may be enhanced by a complex of HS conjugated with nano-Au. The objective of the present study was to determine the effect of administration of nano-Au, HS, and a nano-Au+HS complex on the morphological and molecular characteristics of breast muscle during embryogenesis.Methods: Chicken embryos were used as in vivo model. Fertilized chicken eggs (n = 350) were randomly divided into the control group and the groups treated with nano-Au, HS, and nano-Au+HS. The experimental solutions were given in ovo on the first day of incubation and the embryos were evaluated on day 20 of incubation. The methods included biochemical indi- ces in blood, immunohistochemistry, microscopy (transmission electron microscopy, scanning electron microscopy, confocal), and gene expression at the messenger ribonucleic acid and protein levels.Results: The treatments did not adversely affect mortality, organ weight, and homeostasis of the embryos. HS stimulated the development and maturation of breast muscle by increasing the number of nuclei, satellite cells, and muscle fibers and affected the expression of basic fibroblast growth factor-2 and paired-box transcription factor-7. Furthermore, the nano-Au+HS complex contributed to the increased number of myocytes and nuclei in chicken embryo muscles.Conclusion: The results indicate that the administration of HS and nano-Au affects muscle development and that this effect is enhanced by conjugating HS with nano-Au.Keywords: gold nanoparticles, heparan sulfate, muscle development, embryogenesis, chicke

Novel calcineurin A (PPP3CA) variant associated with epilepsy, constitutive enzyme activation and downregulation of protein expression

PPP3CA encodes calmodulin-binding catalytic subunit of calcineurin, a ubiquitously expressed calcium/calmodulin-regulated protein phosphatase. Recently de novo PPP3CA variants were reported as a cause of disease in 12 subjects presenting with epileptic encephalopathy and dysmorphic features. We describe a boy with similar phenotype and severe early onset epileptic encephalopathy in whom a novel de novo c.1324C>T (p.(Gln442Ter)) PPP3CA variant was found by whole exome sequencing. Western blot experiments in patient's cells (EBV transformed lymphocytes and neuronal cells derived through reprogramming) indicate that despite normal mRNA abundance the protein expression level is strongly reduced both for the mutated and wild-type protein. By in vitro studies with recombinant protein expressed in E. coli we show that c.1324C>T (p.(Gln442Ter)) results in constitutive activation of the enzyme. Our results confirm the role of PPP3CA defects in pathogenesis of a distinct neurodevelopmental disorder including severe epilepsy and dysmorphism and provide further functional clues regarding the pathogenic mechanism