1321. Toxicity of the Diatom Genus Pseudo-nitzschia (Bacillariophyceae) : insights from toxicity tests and genetic screening in the Northern Adriatic SeaTimotej Turk Dermastia, Sonia Dall’Ara, Jožica Dolenc, Patricija Mozetič, 2022, original scientific article Abstract: Diatoms of the genus Pseudo-nitzschia H.Peragallo are known to produce domoic acid (DA), a toxin involved in amnesic shellfish poisoning (ASP). Strains of the same species are often classified as both toxic and nontoxic, and it is largely unknown whether this difference is also genetic. In the Northern Adriatic Sea, there are virtually no cases of ASP, but DA occasionally occurs in shellfish samples. So far, three species—P. delicatissima (Cleve) Heiden, P. multistriata (H. Takano) H. Takano, and P. calliantha Lundholm, Moestrup, & Hasle—have been identified as producers of DA in the Adriatic Sea. By means of enzme-linked immunosorbent assay (ELISA), high-performance liquid chromatography with UV and visible spectrum detection (HPLC-UV/VIS), and liquid chromatography with tandem mass spectrometry (LC-MS/MS), we reconfirmed the presence of DA in P. multistriata and P. delicatissima and detect for the first time in the Adriatic Sea DA in P. galaxiae Lundholm, & Moestrup. Furthermore, we attempted to answer the question of the distribution of DA production among Pseudo-nitzschia species and strains by sequencing the internal transcribed spacer (ITS) phylogenetic marker and the dabA DA biosynthesis gene and coupling this with toxicity data. Results show that all subclades of the Pseudo-nitzschia genus contain toxic species and that toxicity appears to be strain dependent, often with geographic partitioning. Amplification of dabA was successful only in toxic strains of P. multistriata and the presence of the genetic architecture for DA production in non-toxic strains was thus not confirmed.
Keywords: Adriatic, dabA, domoic acid, Pseudo-nitzschia galaxiae
Published in DiRROS: 16.07.2024; Views: 276; Downloads: 177
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1322. Phytoplankton pigments reveal size structure and interannual variability of the coastal phytoplankton community (Adriatic Sea)Vesna Flander-Putrle, Janja Francé, Patricija Mozetič, 2022, original scientific article Abstract: In coastal seas, a variety of environmental variables characterise the average annual pattern of the physico-chemical environment and influence the temporal and spatial variations of phytoplankton communities. The aim of this study was to track the annual and interannual variability of phytoplankton biomass in different size classes in the Gulf of Trieste (Adriatic Sea) using phytoplankton pigments. The seasonal pattern of phytoplankton size classes showed a co-dominance of the nano and micro fractions during the spring peak and a predominance of the latter during the autumn peak. The highest picoplankton values occurred during the periods with the lowest total phytoplankton biomass, with chlorophytes dominating during the colder months and cyanobacteria during the summer. The highest number of significant correlations was found between phytoplankton taxa and size classes and temperature, nitrate and nitrite. The most obvious trend observed over the time series was an increase in picoplankton in all water layers, with the most significant trend in the bottom layer. Nano- and microplankton showed greater variation in biomass, with a decrease in nanoplankton biomass in 2011 and 2012 and negative trend in microplankton biomass in the bottom layer. These results suggest that changes in trophic relationships in the pelagic food web may also have implications for biogeochemical processes in the coastal sea.
Keywords: phytoplankton, biomass, phytoplankton size classes, phytoplankton pigments, HPLC, interannual variability, trends, Adriatic Sea
Published in DiRROS: 16.07.2024; Views: 314; Downloads: 222
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1323. The physiological impact of GFLV virus infection on grapevine water status : first observationsAnastazija Jež Krebelj, Maja Cigoj, Marija Stele, Marko Chersicola, Maruša Pompe Novak, Paolo Sivilotti, 2022, original scientific article Abstract: In a vineyard, grapevines are simultaneously exposed to combinations of several abiotic (drought, extreme temperatures, salinity) and biotic stresses (phytoplasmas, viruses, bacteria). With climate change, the incidences of drought in vine growing regions are increased and the host range of pathogens with increased chances of virulent strain development has expanded. Therefore, we studied the impact of the combination of abiotic (drought) and biotic (Grapevine fanleaf virus (GFLV) infection) stress on physiological and molecular responses on the grapevine of cv. Schioppettino by studying the influence of drought and GFLV infection on plant water status of grapevines, on grapevine xylem vessel occlusion, and on expression patterns of 9-cis-epoxycarotenoid dioxygenase 1 (NCED1), 9-cis-epoxycarotenoid dioxygenase 2 (NCED2), WRKY encoding transcription factor (WRKY54) and RD22-like protein (RD22) genes in grapevines. A complex response of grapevine to the combination of drought and GFLV infection was shown, including priming in the case of grapevine water status, net effect in the case of area of occluded vessels in xylem, and different types of interaction of both stresses in the case of expression of four abscisic acid-related genes. Our results showed that mild (but not severe) water stress can be better sustained by GFLV infection rather than by healthy vines. GFLV proved to improve the resilience of the plants to water stress, which is an important outcome to cope with the challenges of global warming.
Keywords: grapevine, water status, virus infection, GFLV, xylem vessel occlusion, gene expression
Published in DiRROS: 16.07.2024; Views: 271; Downloads: 257
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1324. Lethal and sub-lethal effects and modulation of gene expression induced by T kinase inhibitors in zebrafish (Danio Rerio) embryosTina Eleršek, Matjaž Novak, Mateja Mlinar, Igor Virant, Nika Bahor, Karin Leben, Bojana Žegura, Metka Filipič, 2022, original scientific article Abstract: Tyrosine kinase inhibitors (TKIs) are designed for targeted cancer therapy. The consumption of these drugs during the last 20 years has been constantly rising. In the zebrafish (Danio rerio) embryo toxicity test, we assessed the toxicity of six TKIs: imatinib mesylate, erlotinib, nilotinib, dasatinib, sorafenib and regorafenib. Imatinib mesylate and dasatinib induced lethal effects, while regorafenib, sorfenib and dasatinib caused a significant increase of sub-lethal effects, predominantly oedema, no blood circulation and formation of blood aggregates. The analyses of the changes in the expression of selected genes associated with the hormone system after the exposure to imatinib mesylate, dasatinib and regorafenib demonstrated that all three tested TKIs deregulated the expression of oestrogen receptor esr1, cytochrome P450 aromatase (cypa19b) and hydroxysteroid-dehydrogenase (hsd3b), regorafenib, and also thyroglobulin (tg). The expression of genes involved in the DNA damage response (gadd45 and mcm6) and apoptosis (bcl2) was deregulated only by exposure to regorafenib. The data indicate that common mechanisms, namely antiangiogenic activity and interference with steroidogenesis are involved in the TKI induced sub-lethal effects and potential hormone disrupting activity, respectively. The residues of TKIs may represent an environmental hazard; therefore, further ecotoxicological studies focusing also on the effects of their mixtures are warranted.
Keywords: aquatic toxicity, tyrosine kinase inhibitors, zebrafish embryo toxicity test, gene expression, environmental hazard
Published in DiRROS: 16.07.2024; Views: 306; Downloads: 196
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1325. Hydrodynamic cavitation efficiently inactivates potato virus Y in waterArijana Filipić, Tadeja Lukežič, Katarina Bačnik, Maja Ravnikar, Meta Ješelnik, Tamara Košir, Martin Petkovšek, Mojca Zupanc, Matevž Dular, Ion Gutiérrez-Aguirre, 2022, original scientific article Abstract: Waterborne plant viruses can destroy entire crops, leading not only to high financial losses but also to food shortages. Potato virus Y (PVY) is the most important potato viral pathogen that can also affect other valuable crops. Recently, it has been confirmed that this virus is capable of infecting host plants via water, emphasizing the relevance of using proper strategies to treat recycled water in order to prevent the spread of the infectious agents. Emerging environmentally friendly methods such as hydrodynamic cavitation (HC) provide a great alternative for treating recycled water used for irrigation. In the experiments conducted in this study, laboratory HC based on Venturi constriction with a sample volume of 1 L was used to treat water samples spiked with purified PVY virions. The ability of the virus to infect plants was abolished after 500 HC passes, corresponding to 50 min of treatment under pressure difference of 7 bar. In some cases, shorter treatments of 125 or 250 passes were also sufficient for virus inactivation. The HC treatment disrupted the integrity of viral particles, which also led to a minor damage of viral RNA. Reactive species, including singlet oxygen, hydroxyl radicals, and hydrogen peroxide, were not primarily responsible for PVY inactivation during HC treatment, suggesting that mechanical effects are likely the driving force of virus inactivation. This pioneering study, the first to investigate eukaryotic virus inactivation by HC, will inspire additional research in this field enabling further improvement of HC as a water decontamination technology.
Keywords: hydrodynamic cavitation, potato virus Y, virus inactivation, water decontamination
Published in DiRROS: 16.07.2024; Views: 314; Downloads: 212
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1326. Differential responses of coexisting owls to annual small mammal population fluctuations in temperate mixed forestUrška Ratajc, Martin Breskvar, Sašo Džeroski, Al Vrezec, 2022, original scientific article Abstract: Montane temperate forests in central and southern Europe host diverse small mammal assemblages, but the fluctuations in these assemblages in correlation with owl predators are still poorly explored. The key questions of our study were how coexisting owls responded to different prey fluctuations and whether any particular small mammal species governed predator–prey co-dynamics. We conducted a long-term study (2004–2020) in low-elevation (300–1100 m above sea level) mixed Beech and Silver Fir forest in the northern Dinaric Alps (central Slovenia). Monitoring data on the main small mammal groups – mice Muridae, voles Cricetidae, dormice Gliridae and shrews Soricidae – and three owl species – the Ural Owl Strix uralensis, Tawny Owl Strix aluco and Boreal Owl Aegolius funereus – were collected annually. To find relationships between prey and predator populations, we used two types of supervised machine learning approaches and addressed three predictive modelling tasks of multi-target regression. The dominant species in the small mammal assemblage, the Yellow-necked Mouse Apodemus flavicollis, had a key role in determining predator populations and their breeding performance. We noted higher sensitivity to small mammal fluctuations in boreal zone owl species (Boreal Owl and Ural Owl), which reach their southern distribution limit in the Dinaric Alps, whereas the temperate zone species (Tawny Owl) seemed to be less affected. In years of prey shortage, the Boreal Owl was found to presumably abandon its territories, the Ural Owl suppressed breeding and the Tawny Owl sustained breeding activity by shifting prey selection. Low-elevation forests appeared to be suboptimal habitat for the competitive subordinate Boreal Owl, which may exploit occasional outbreaks of small mammal populations in these habitats even in the presence of larger competitors. Whether low-elevation forests can play a role in maintaining threatened and cold-adapted Boreal Owl populations in central and southern Europe in the face of recent ecosystem changes due to climate and environmental changes remains an open scientific question.
Keywords: sove, mali sesalci, populacijska dinamika, strojno učenje
Published in DiRROS: 16.07.2024; Views: 274; Downloads: 121
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1327. HepG2 spheroids as a biosensor-like cell-based system for (geno)toxicity assessmentMartina Štampar, Sonja Žabkar, Metka Filipič, Bojana Žegura, 2022, original scientific article Abstract: 3D spheroids developed from HepG2 cells were used as a biosensor-like system for the detection of (geno)toxic effects induced by chemicals. Benzo(a)pyrene (B(a)P) and amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) with well-known mechanisms of action were used for system validation. HepG2 spheroids grown for 3 days were exposed to BaP and PhIP for 24 and 72 h. The growth and viability of spheroids were monitored by planimetry and Live/Dead staining of cells. Multi-parametric flow cytometric analysis was applied for simultaneous detection of specific end-effects including cell cycle analysis (Hoechst staining), cell proliferation (KI67 marker), and DNA double-strand breaks (ℽH2AX) induced by genotoxic compounds. Depending on the exposure concentration/time, BaP reduced spheroid growth, affected cell proliferation by arresting cells in S and G2 phase and induced DNA double-strand breaks (DSB). Simultaneous staining of ℽH2AX formation and cell cycle analysis revealed that after BaP (10 μM; 24 h) exposure 60% of cells in G0/G1 phase had DNA DSB, while after 72 h only 20% of cells contained DSB indicating efficient repair of DNA lesions. PhIP did not influence the spheroid size whereas accumulation of cells in the G2 phase occurred after both treatment times. The evaluation of DNA damage revealed that at 200 μM PhIP 50% of cells in G0/G1 phase had DNA DSB, which after 72-h exposure dropped to 40%, showing lower repair capacity of PhIP-induced DSB compared to BaP-induced. The developed approach using simultaneous detection of several parameters provides mechanistic data and thus contributes to more reliable genotoxicity assessment of chemicals as a high-content screening tool.
Keywords: in vitro 3D cell model, HepG2, flow cytometry, cell cycle, proliferation, DNA strand, breaks
Published in DiRROS: 16.07.2024; Views: 312; Downloads: 180
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1328. An assessment of the reproducibility of reverse transcription digital PCR quantification of HIV-1Samreen Falak, Rainer Macdonald, Eloise J. Busby, Denise M. O'Sullivan, Mojca Milavec, Annabell Plauth, Martin Kammel, Heinz Zeichhardt, Hans-Peter Grunert, Andreas Kummrow, Jim F. Huggett, 2022, original scientific article Abstract: Viral load monitoring in human immunodeficiency virus type 1 (HIV-1) infection is often performed using reverse transcription quantitative PCR (RT-qPCR) to observe response to treatment and identify the development of resistance. Traceability is achieved using a calibration hierarchy traceable to the International Unit (IU). IU values are determined using consensus agreement derived from estimations by different laboratories. Such a consensus approach is necessary due to the fact that there are currently no reference measurement procedures available that can independently assign a reference value to viral reference materials for molecular in vitro diagnostic tests. Digital PCR (dPCR) is a technique that has the potential to be used for this purpose. In this paper, we investigate the ability of reverse transcriptase dPCR (RT-dPCR) to quantify HIV-1 genomic RNA without calibration. Criteria investigated included the performance of HIV-1 RNA extraction steps, choice of reverse transcription approach and selection of target gene with assays performed in both single and duplex format. We developed a protocol which was subsequently applied by two independent laboratories as part of an external quality assurance (EQA) scheme for HIV-1 genome detection. Our findings suggest that RT-dPCR could be used as reference measurement procedure to aid the value assignment of HIV-1 reference materials to support routine calibration of HIV-1 viral load testing by RT-qPCR.
Published in DiRROS: 16.07.2024; Views: 266; Downloads: 145
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1329. The performance of human cytomegalovirus digital PCR reference measurement procedure in seven external quality assessment schemes over four yearsMojca Milavec, Jernej Pavšič, Alexandra Bogožalec Košir, Gerwyn M. Jones, Denise M. O'Sullivan, Alison S. Devonshire, Fran Van Heuverswyn, Maria Karczmarczyk, Jannika Neeb, Annabell Plauth, Philippe Corbisier, Heinz Schimmel, Andreas Kummrow, Jörg Neukammer, Carole A. Foy, Martin Kammel, Hans-Peter Grunert, Heinz Zeichhardt, Jim F. Huggett, 2022, original scientific article Published in DiRROS: 15.07.2024; Views: 371; Downloads: 148
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