RESUMO
Lungworms of the genus Dictyocaulus are causative agents of parasitic bronchitis in domestic and wild ungulates. This study investigates the distribution, morphology and genetic diversity of D. cervi and a new lungworm species, Dictyocaulus skrjabini n. sp. infecting red deer Cervus elaphus, fallow deer Dama dama and moose Alces alces in Poland and Sweden. The study was conducted on 167 red deer from Poland and on the DNA of lungworms derived from 7 fallow deer, 4 red deer and 2 moose collected in Sweden. The prevalence of D. cervi and D. skrjabini n. sp. in dissected red deer in Poland was 31.1% and 7.2%, respectively. Moreover, D. skrjabini n. sp. was confirmed molecularly in 7 isolates of fallow deer lungworms and 1 isolate of red deer lungworms from Sweden. Dictyocaulus skrjabini n. sp. was established based on combination of their distinct molecular and morphological features; these included the length of cephalic vesicle, buccal capsule (BC), buccal capsule wall (BCW), distance from anterior extremity to the nerve ring, the width of head, oesophagus, cephalic vesicle, BC and BCW, as well as the dimensions of reproductive organs of male and female. Additionally, molecular analyses revealed 0.9% nucleotide sequence divergence for 1,605 bp SSU rDNA, and 16.517.3% nucleotide sequence divergence for 642 bp mitochondrial cytB between D. skrjabini n. sp. and D. cervi, respectively, and 18.719% between D. skrjabini n. sp. and D. eckerti, which translates into 18.218.7% amino acid sequence divergence between D. skrjabini n. sp. and both lungworms.
Assuntos
Cervos , Infecções por Dictyocaulus , Nematoides , Animais , Feminino , Masculino , Dictyocaulus/genética , Cervos/parasitologia , Infecções por Dictyocaulus/epidemiologia , Nematoides/genética , Sequência de BasesRESUMO
Prototheca species are unicellular, nonphotosynthetic, saprophytic, and occasionally pathogenic, microalgae, with an extensive environmental reservoir. This study explores, for the first time, the occurrence of Prototheca in aquatic ecosystems by using a molecular profiling approach. A total of 362 samples were collected from 80 natural and artificial waterbodies at 88 sampling sites in 26 localities across Poland during a 1.5-year period. The overall isolation rate of Prototheca from water environments was 14.1%. Prototheca were most prevalent in rivers of urbanized areas, indicating that the algae are primarily adapted to lotic ecosystems with a high input of organic matter. Interestingly, it is not the amount of organic matter per se but its quality that seems to shape the habitat potential of the protothecae. The two most frequently isolated species were P. wickerhamii and P. pringsheimii, representing a third and a fourth of the strains, respectively. Additionally, three novel species were described, namely, P. fontanea, P. lentecrescens, and P. vistulensis. The high species diversity of the genus Prototheca may reflect the complexity of water ecosystems along with ecological and functional adaptations of the algae to such environments. For further investigations, the study provides a revised scheme for identification of all 18 Prototheca species currently recognized. IMPORTANCE The study investigates the occurrence of very rare and poorly studied microalgae of the genus Prototheca, potentially pathogenic to humans and animals, in different water environments. Given the potential hazard to human and animal health from exposure to water-inhabiting protothecae, the prevalence of the algae in aquatic habitats deserves an insightful examination. The study is the first since the 1980s to explore the aquatic habitat of Prototheca spp. and the first ever performed to do this by molecular methods. Although the Prototheca isolation rate was low, a high species diversity was observed. The algae appear to represent allochthonous microflora, brought into waterbodies from various anthropogenic sources. Large rivers of urbanized areas were the most Prototheca-abundant. The study provides a description of three new Prototheca species, namely, P. fontanea, P. lentecrescens, and P. vistulensis. The study also delivers a new identification scheme for all Prototheca species currently recognized.
Assuntos
Microalgas , Prototheca , Animais , Humanos , Ecossistema , Água , PolôniaRESUMO
There is an ongoing search for alternative treatments for Clostridioides difficile infections. The aim of the study was to investigate the antibacterial and antibiotic activity of bee products against C. difficile strains with different polymerase chain reaction ribotypes (RTs). The minimum inhibitory concentration (MICs) of Manuka honey 550+, goldenrod honey, pine honey, and bee bread were determined by the broth dilution method. C. difficile adhesion to HT-29, HT-29 MTX, and CCD 841 CoN cell lines was assessed. Biofilm was cultured in titration plates and visualized by confocal microscopy. The MICs of Manuka honey for C. difficile 630 and ATCC 9689 strains and control strain, M 120, were 6.25%, 6.25%, and 1.56% (v/v), respectively; of goldenrod honey, 50%, 50%, and 12.5%, respectively; of pine honey, 25%, 25%, and 25%, respectively; and of bee bread, 100 mg/L, 50 mg/L, and 100 mg/L, respectively. Manuka honey (1%) increased adhesion of C. difficile RT176 strains, and one strain of RT023, to the CCD 841 cell line. Pine honey (1%) increased RT027 adhesion to the HT-29 cell line. Manuka honey, pine honey, and bee bread at subinhibitory concentrations increased the adhesion of C. difficile. Our research proved that bee products are active against the tested strains of C. difficile.
Assuntos
Clostridioides difficile , Mel , Própole , Abelhas , Animais , Ribotipagem , Clostridioides , Própole/farmacologia , Biofilmes , Testes de Sensibilidade Microbiana , Antibacterianos/farmacologia , Antibacterianos/uso terapêuticoRESUMO
Eukaryotic cells rely on the accuracy and efficiency of vesicular traffic. In plants, disturbances in vesicular trafficking are well studied in quickly dividing root meristem cells or polar growing root hairs and pollen tubes. The development of the female gametophyte, a unique haploid reproductive structure located in the ovule, has received far less attention in studies of vesicular transport. Key molecules providing the specificity of vesicle formation and its subsequent recognition and fusion with the acceptor membrane are Rab proteins. Rabs are anchored to membranes by covalently linked geranylgeranyl group(s) that are added by the Rab geranylgeranyl transferase (RGT) enzyme. Here we show that Arabidopsis plants carrying mutations in the gene encoding the ß-subunit of RGT (rgtb1) exhibit severely disrupted female gametogenesis and this effect is of sporophytic origin. Mutations in rgtb1 lead to internalization of the PIN1 and PIN3 proteins from the basal membranes to vesicles in provascular cells of the funiculus. Decreased transport of auxin out of the ovule is accompanied by auxin accumulation in tissue surrounding the growing gametophyte. In addition, female gametophyte development arrests at the uni- or binuclear stage in a significant portion of the rgtb1 ovules. These observations suggest that communication between the sporophyte and the developing female gametophyte relies on Rab-dependent vesicular traffic of the PIN1 and PIN3 transporters and auxin efflux out of the ovule.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Ácidos Indolacéticos , Óvulo Vegetal/genética , Tubo PolínicoRESUMO
Auxin is a key regulator of plant development affecting the formation and maturation of reproductive structures. The apoplastic route of auxin transport engages influx and efflux facilitators from the PIN, AUX and ABCB families. The polar localization of these proteins and constant recycling from the plasma membrane to endosomes is dependent on Rab-mediated vesicular traffic. Rab proteins are anchored to membranes via posttranslational addition of two geranylgeranyl moieties by the Rab Geranylgeranyl Transferase enzyme (RGT), which consists of RGTA, RGTB and REP subunits. Here, we present data showing that seed development in the rgtb1 mutant, with decreased vesicular transport capacity, is disturbed. Both pre- and post-fertilization events are affected, leading to a decrease in seed yield. Pollen tube recognition at the stigma and its guidance to the micropyle is compromised and the seed coat forms incorrectly. Excess auxin in the sporophytic tissues of the ovule in the rgtb1 plants leads to an increased tendency of autonomous endosperm formation in unfertilized ovules and influences embryo development in a maternal sporophytic manner. The results show the importance of vesicular traffic for sexual reproduction in flowering plants, and highlight RGTB1 as a key component of sporophytic-filial signaling.
Assuntos
Arabidopsis/enzimologia , Sementes/enzimologia , Transdução de Sinais , Alquil e Aril Transferases/metabolismo , Alquil e Aril Transferases/fisiologia , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Transporte Biológico , Regulação da Expressão Gênica de Plantas , Ácidos Indolacéticos/metabolismo , Mutação , Tubo Polínico/fisiologia , Sementes/crescimento & desenvolvimento , Sementes/metabolismoRESUMO
Nearly half of patients with advanced and metastatic melanomas harbor a BRAF mutation. Vemurafenib (VEM), a BRAF inhibitor, is used to treat such patients, however, responses to VEM are very short-lived due to intrinsic, adaptive and/or acquired resistance. In this context, we present the action of the B-Raf serine-threonine protein kinase inhibitor (vemurafenib) on the glycans structure and metallomics profiles in melanoma cells without (MeWo) and with (G-361) BRAF mutations. The studies were performed using α1-acid glycoprotein (AGP), a well-known acute-phase protein, and concanavalin A (Con A), which served as the model receptor. The detection of changes in the structure of glycans can be successfully carried out based on the frequency shifts and the charge transfer resistance after interaction of AGP with Con A in different VEM treatments using QCM-D and EIS measurements. These changes were also proved based on the cell ultrastructure examined by TEM and SEM. The LA-ICP-MS studies provided details on the metallomics profile in melanoma cells treated with and without VEM. The studies evidence that vemurafenib modifies the glycans structures and metallomics profile in melanoma cells harboring BRAF mutation that can be further implied in the resistance phenomenon. Therefore, our data opens a new avenue for further studies in the short-term addressing novel targets that hopefully can be used to improve the therapeutic regiment in advanced melanoma patients. The innovating potential of this study is fully credible and has a real impact on the global patient society suffering from advanced and metastatic melanomas.
Assuntos
Melanoma/metabolismo , Metais/metabolismo , Mutação , Polissacarídeos/química , Proteínas Proto-Oncogênicas B-raf/genética , Vemurafenib/farmacologia , Concanavalina A/química , Concanavalina A/metabolismo , Humanos , Melanoma/tratamento farmacológico , Melanoma/genética , Melanoma/patologia , Metais/análise , Orosomucoide/química , Orosomucoide/metabolismo , Inibidores de Proteínas Quinases/farmacologiaRESUMO
Our study aimed at examining the phylogenetic position of the newly-found Setaria nematodes obtained from the red deer (Cervus elaphus) based on sequences of the mitochondrial cytochrome c oxidase subunit 1 (COX-1). Alignment and phylogenetic analyses, as well as SEM microscopic analysis, revealed the presence of two Setaria species: S. cervi and S. tundra. Setaria tundra was noted in only one individual, a calf of the red deer, while S. cervi was observed in three stages, two hinds and one calf of the red deer. According to our knowledge, it is the first case of S. cervi in the red deer in Poland confirmed in molecular studies and also the first case of S. tundra infection in the red deer.
Assuntos
Cervos/parasitologia , Filogenia , Setaria (Nematoide)/classificação , Setaria (Nematoide)/isolamento & purificação , Animais , DNA Mitocondrial/química , DNA Mitocondrial/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , Microscopia Eletrônica de Varredura , Polônia , Análise de Sequência de DNA , Setaria (Nematoide)/anatomia & histologia , Setaria (Nematoide)/genéticaRESUMO
Dolichol is a required cofactor for protein glycosylation, the most common posttranslational modification modulating the stability and biological activity of proteins in all eukaryotic cells. We have identified and characterized two genes, PPRD1 and -2, which are orthologous to human SRD5A3 (steroid 5α reductase type 3) and encode polyprenol reductases responsible for conversion of polyprenol to dolichol in Arabidopsis thaliana. PPRD1 and -2 play dedicated roles in plant metabolism. PPRD2 is essential for plant viability; its deficiency results in aberrant development of the male gametophyte and sporophyte. Impaired protein glycosylation seems to be the major factor underlying these defects although disturbances in other cellular dolichol-dependent processes could also contribute. Shortage of dolichol in PPRD2-deficient cells is partially rescued by PPRD1 overexpression or by supplementation with dolichol. The latter has been discussed as a method to compensate for deficiency in protein glycosylation. Supplementation of the human diet with dolichol-enriched plant tissues could allow new therapeutic interventions in glycosylation disorders. This identification of PPRD1 and -2 elucidates the factors mediating the key step of the dolichol cycle in plant cells which makes manipulation of dolichol content in plant tissues feasible.
Assuntos
Arabidopsis/enzimologia , Dolicóis/metabolismo , Oxirredutases/metabolismo , Processamento de Proteína Pós-Traducional , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Glicosilação , Mutação , Oxirredutases/genética , Infertilidade das PlantasRESUMO
Rab proteins, key players in vesicular transport in all eukaryotic cells, are post-translationally modified by lipid moieties. Two geranylgeranyl groups are attached to the Rab protein by the heterodimeric enzyme Rab geranylgeranyl transferase (RGT) αß. Partial impairment in this enzyme activity in Arabidopsis, by disruption of the AtRGTB1 gene, is known to influence plant stature and disturb gravitropic and light responses. Here it is shown that mutations in each of the RGTB genes cause a tip growth defect, visible as root hair and pollen tube deformations. Moreover, FM 1-43 styryl dye endocytosis and recycling are affected in the mutant root hairs. Finally, it is demonstrated that the double mutant, with both AtRGTB genes disrupted, is non-viable due to absolute male sterility. Doubly mutated pollen is shrunken, has an abnormal exine structure, and shows strong disorganization of internal membranes, particularly of the endoplasmic reticulum system.
Assuntos
Arabidopsis/enzimologia , Arabidopsis/genética , Flores/genética , Mutação , Transferases/genética , Transferases/metabolismo , Alquil e Aril Transferases/química , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Sequência de Aminoácidos , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Sequência de Bases , Fertilidade/genética , Flores/anatomia & histologia , Flores/crescimento & desenvolvimento , Dados de Sequência Molecular , Pólen/metabolismo , Tubo Polínico/crescimento & desenvolvimento , Reprodução , Transferases/químicaRESUMO
Morphological anomalies are considered a rare phenomenon among natural tick populations. New cases of abnormalities in ticks are being described, such as body assymetries, nanism, gynandromorphism and limb malformations. The tick removed from a cat was morphologically identified to species and developmental stage. The time of feeding on the host was determined. The specimen was tested using PCR and Real-Time PCR methods for the presence of the common tick-borne pathogens: Anaplasma phagocytophilum, Babesia spp, Borrelia spp., Neoehrlichia mikurensis, Rickettsia spp. For visualisation of the anomalous structures, scanning electron microscopy (SEM) was performed. The tick was identified as a slightly engorged adult female of I. ricinus exhibiting ectromely of leg I on the left side of the idiosoma. The specimen was tested positive for two medically important pathogens: A. phagocytophilum and N. mikurensis. The case report describes a rare case of a morphological anomaly in an I. ricinus tick from Poland.
Assuntos
Ixodes , Animais , Polônia , Ixodes/microbiologia , Ixodes/crescimento & desenvolvimento , Feminino , Gatos , Doenças do Gato/parasitologia , Doenças do Gato/microbiologia , Anaplasma phagocytophilum/isolamento & purificação , Anaplasma phagocytophilum/genética , Infestações por Carrapato/veterinária , Infestações por Carrapato/parasitologiaRESUMO
Micro- (MPs) and nanoplastics (NPs) are currently ubiquitous in the ecosystems, and freshwater biota is still insufficiently studied to understand the global fate, transport paths, and consequences of their presence. Thus, in this study, we investigated the role of bivalves and a trophic transfer of MPs and NPs in an experimental food chain. The food chain consisted of terrestrial non-selective detritivore Dendrobaena (Eisenia) sp., freshwater benthic filter feeder Unio tumidus, and freshwater benthic detritivore-collectors Asellus aquaticus or Gammarus sp. Animals were exposed to different fluorescently labeled micro- and nanoplastics (PMMA 20 µm, nanoPS 15-18 nm, and 100 nm, PS 1 µm and 20 µm, PE from cosmetics) as well as to the faeces of animals exposed to plastics to assess their influence on the environmental transportation, availability to biota, and bioaccumulation of supplied particles. Damaged and intact fluorescent particles were observed in the faeces of terrestrial detritivores and in the droppings of aquatic filter feeders, respectively. They were also present in the guts of bivalves and of crustaceans which were fed with bivalve droppings. Bivalves (Unio tumidus, and additionally Unio pictorum, and Sphaerium corneum) produced droppings containing micro- and nanoparticles filtered from suspension and deposited them onto the tank bottom, making them available for broader feeding guilds of animals (e.g. collectors, like crustaceans). Finally, the natural ageing of PS and its morphological changes, leakage of the fluorescent labelling, and agglomeration of particles were demonstrated. That supports our hypothesis of the crucial role of the characterization of physical and chemical materials in adequately understanding the mechanisms of their interaction with biota. Microscopical methods (confocal, fluorescent, scanning electron) and Raman and FT-IR spectroscopy were used to track the particles' passage in a food web and monitor structural changes of the MPs' and NPs' surface.
Assuntos
Bivalves , Unio , Poluentes Químicos da Água , Animais , Microplásticos , Cadeia Alimentar , Espécies Sentinelas , Ecossistema , Espectroscopia de Infravermelho com Transformada de Fourier , Poluentes Químicos da Água/análise , Plásticos , Água Doce/químicaRESUMO
Cytotoxic and genotoxic effects of novel mPEG-silane coated iron(III) oxide nanoparticles doped with magnesium (Mg0.1-γ-Fe2O3(mPEG-silane)0.5) have been investigated on human adenocarcinomic alveolar basal epithelial (A549) and human normal bronchial epithelial (BEAS-2B) cells. In the studies several molecular and cellular targets addressing to cell membrane, cytoplasm organelles and nucleus components were served as toxicological endpoints. The as-synthesized nanoparticles were found to be stable in the cell culture media and were examined for different concentration and exposure times. No cytotoxicity of the tested nanoparticles was found although these nanoparticles slightly increased reactive oxygen species in both cell types studied. Mg0.1-γ-Fe2O3(mPEG-silane)0.5 nanoparticles did not produce any DNA strand breaks and oxidative DNA damages in A549 and BEAS-2B cells. Different concentration of Mg0.1-γ-Fe2O3(mPEG-silane)0.5 nanoparticles and different incubation time did not affect cell migration. The lung cancer cells' uptake of the nanoparticles was more effective than in normal lung cells. Altogether, the results evidence that mPEG-silane coated iron(III) oxide nanoparticles doped with magnesium do not elucidate any deleterious effects on human normal and cancerous lung cells despite cellular uptake of these nanoparticles. Therefore, it seems reasonable to conclude that these novel biocompatible nanoparticles are promising candidates for further development towards medical applications.
Assuntos
Dano ao DNA , Pulmão , Magnésio , Polietilenoglicóis , Silanos , Humanos , Silanos/toxicidade , Silanos/química , Polietilenoglicóis/toxicidade , Polietilenoglicóis/química , Magnésio/química , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/citologia , Neoplasias Pulmonares/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Linhagem Celular , Compostos Férricos/toxicidade , Compostos Férricos/química , Movimento Celular/efeitos dos fármacos , Nanopartículas Magnéticas de Óxido de Ferro/toxicidade , Nanopartículas Magnéticas de Óxido de Ferro/química , Linhagem Celular Tumoral , Células A549RESUMO
Brassinosteroids are important plant hormones influencing, among other processes, chloroplast development, the electron transport chain during light reactions of photosynthesis, and the Calvin-Benson cycle. Medium-chain-length polyprenols built of 9-11 isoprenoid units (C45-C55 carbons) are a class of isoprenoid compounds present in abundance in thylakoid membranes. They are synthetized in chloroplast by CPT7 gene from Calvin cycle derived precursors on MEP (methylerythritol 4-phosphate) isoprenoid biosynthesis pathway. C45-C55 polyprenols affect thylakoid membrane ultra-structure and hence influence photosynthetic apparatus performance in plants such as Arabidopsis and tomato. So far nothing is known about the hormonal or environmental regulation of CPT7 gene expression. The aim of our study was to find out if medium-chain-length polyprenol biosynthesis in plants may be regulated by hormonal cues.We found that the CPT7 gene in Arabidopsis has a BZR1 binding element (brassinosteroid dependent) in its promoter. Brassinosteroid signaling mutants in Arabidopsis accumulate a lower amount of medium-chain-length C45-C55 polyprenols than control plants. At the same time carotenoid and chlorophyll content is increased, and the amount of PsbD1A protein coming from photosystem II does not undergo a significant change. On contrary, treatment of WT plants with epi-brassinolide increases C45-C55 polyprenols content. We also report decreased transcription of MEP enzymes (besides C45-C55 polyprenols, precursors of numerous isoprenoids, e.g. phytol, carotenoids are derived from this pathway) and genes encoding biosynthesis of medium-chain-length polyprenol enzymes in brassinosteroid perception mutant bri1-116. Taken together, we document that brassinosteroids affect biosynthetic pathway of C45-C55 polyprenols.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Brassinosteroides/metabolismo , Poliprenois/metabolismo , Cloroplastos/metabolismo , Terpenos/metabolismo , Carotenoides/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de PlantasRESUMO
Magnetic fluid hyperthermia (MFH) is a promising therapeutic strategy that targets malignant tissues by heating to 40-43 °C using magnetic nanoparticles (MNPs) subjected to an alternating magnetic field (AMF). In this study, novel magnetic iron(III) oxide nanoparticles doped with magnesium (Mg0.1-γ-Fe2O3(mPEG-silane)0.5) were synthesized, and their structural, chemical, and magnetic properties were analyzed using the following techniques: Fourier-transform infrared spectroscopy, Raman spectroscopy, vibrating magnetometer analysis, powder X-ray diffraction, inductively coupled plasma mass spectrometry, scanning electron microscopy, high-resolution transmission electron microscopy, and energy-dispersive X-ray spectroscopy. The as-synthesized MNPs were used as water ferrofluids for MFH under an AMF in two calorimetric setups, namely phantom and lung cancer cell (A549) models. The as-synthesized MNPs were hexagonal or rhombohedral shaped, with an average size of 27 nm. They showed a typical soft ferromagnetic behavior based on the hysteresis profile, with a magnetic saturation of 70 emu g-1 and remnant magnetization of 1.6 emu g-1. In phantom studies, the ferrofluid (3.0 mg mL-1) exposed to an AMF (18.3 kA m-1, 110.1 kHz) heated up extremely quickly, reaching more than 90 °C in the first 10 min of magnetization. In cell studies, the ferrofluid (0.25 mg mL-1) under an AMF (16.7 kA m-1, 110.1 kHz) showed a slight increase in temperature within the first 12 min, reaching a peak of ca. 43-45 °C, which was stable up to the end of the AMF exposure (45 min). Under these conditions, a pronounced cytotoxic effect on the lung cancer cells was observed (viability ca. 15-20%). No such deleterious effects were observed when the cells were treated with MNPs only without an AMF. Specific absorption rate (SAR) measurements were performed using three mathematical approaches, namely the initial slope method, the corrected slope method, and the Box-Lucas method, which ranged from ca. 429 to 596 W g-1 for phantom and cell studies. Iron(III) oxide MNPs doped with magnesium were found to be candidates for MFH in lung cancer treatments.
Assuntos
Hipertermia Induzida , Neoplasias Pulmonares , Nanopartículas de Magnetita , Humanos , Magnésio , Nanopartículas de Magnetita/uso terapêutico , Nanopartículas de Magnetita/química , Hipertermia Induzida/métodos , Ferro , Óxidos , Neoplasias Pulmonares/terapia , Hipertermia , Campos MagnéticosRESUMO
The influence of triterpenoid saponins on subcellular morphological changes in the cells of parasitic nematodes remains poorly understood. Our study examines the effect of oleanolic acid glucuronides from marigold (Calendula officinalis) on the possible modification of immunogenic proteins from infective Heligmosomoides polygyrus bakeri larvae (L3). Our findings indicate that the triterpenoid saponins alter the subcellular morphology of the larvae and prevent recognition of nematode-specific proteins by rabbit immune-IgG. TEM ultrastructure and HPLC analysis showed that microtubule and cytoskeleton fibres were fragmented by saponin treatment. MASCOT bioinformatic analysis revealed that in larvae exposed to saponins, the immune epitopes of their proteins altered. Several mitochondrial and cytoskeleton proteins involved in signalling and cellular processes were downregulated or degraded. As possible candidates, the following set of recognised proteins may play a key role in the immunogenicity of larvae: beta-tubulin isotype, alpha-tubulin, myosin, paramyosin isoform-1, actin, disorganized muscle protein-1, ATP-synthase, beta subunit, carboxyl transferase domain protein, glutamate dehydrogenase, enolase (phosphopyruvate hydratase), fructose-bisphosphate aldolase 2, tropomyosin, arginine kinase or putative chaperone protein DnaK, and galactoside-binding lectin. Data are available via ProteomeXchange with identifier PXD024205.
RESUMO
Tetraspanin CD9 is the only protein of the oocyte membrane (oolemma) known to be required for the fusion of gametes during fertilization in the mouse. Using electron microscopy and immunostaining we examined the differences in localization of CD9 between ovulated oocytes, zygotes and parthenogenetically activated eggs (parthenogenotes). Changes in ultrastructure of oolemma, which take place in oocytes after fertilization or artificial activation, were also assessed. We demonstrated that after fertilization the level of CD9 present on microvilli of zygote was two times lower than its level on the oolemma of the oocyte. In addition, we showed that the distribution of microvilli is less uniform in the zygotes than in the unfertilized oocytes. We propose that the changes of microvilli distribution and their CD9 content are responsible for the development of the oocyte membrane block to sperm penetration.
Assuntos
Antígenos CD/análise , Glicoproteínas de Membrana/análise , Oócitos/fisiologia , Oócitos/ultraestrutura , Interações Espermatozoide-Óvulo/fisiologia , Animais , Antígenos CD/metabolismo , Membrana Celular/metabolismo , Masculino , Glicoproteínas de Membrana/metabolismo , Camundongos , Microvilosidades/metabolismo , Oócitos/química , Tetraspanina 29RESUMO
Although the Dictyocaulus lungworm, the agent of dictyocaulosis, is one of parasitological threats to European bison, its systematic position remains unclear. The aim of the present study was to evaluate the morphological features of the lungworm and the pathological lesions it induces, and to analyse mitochondrial (mt) genetic markers for systematic and molecular epidemiological studies. The morphological findings indicate that Dictyocaulus lungworms of European bison can be distinguished from those of cattle on the basis of differences in buccal capsule wall length, total body length, and spicules length in males, all of which were significantly longer in those of European bison. Nucleotide diversity calculated from pairwise sequence alignments of partial cytochrome c oxidase subunit 1 (cox1), cytochrome B (cytB) and NADH dehydrogenase subunit 5 (nad5) of specimens from cattle and European bison varied from 1.7% for nad5, 2.1% for cytB, to 3.7% for cox1 gene. Thus, among the lungworms of European bison and cattle, nad5 and cytB were the most conserved proteins, whereas cox1 was the most diverse. The mt cytB marker gene may be a suitable candidate for distinguishing between the two genotypes, as nad5 demonstrated the greatest within-genus sequence variation. The lung tissue of infected European bison manifests signs of verminous pneumonia characterized by interstitial pneumonia, bronchitis and bronchiolitis. Therefore, it appears that European bison and cattle are infected with slightly diverged, morphologically-different, genotypes of D. viviparus, indicating they belong to two separate worm populations. We propose, therefore, that the lungworm of European bison should be classified as D. viviparus subsp. bisontis.
RESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Uncaria tomentosa (Willd.) DC. is the most popular Peruvian plant, used in folk medicine for different purposes. It contains thousands of active compounds with great content of alkaloids. AIM OF STUDY: Two different fractions of Alkaloid-Rich and Alkaloid-Free were researched on chromosome morphology, mitotic activity and phases indexes. MATERIALS AND METHODS: Cells of Allium Test (meristematic cells of root tips) were incubated up to 24h in different concentrations of Alkaloid-Free and Alkaloid-Rich fraction obtained from Uncaria tomentosa bark followed by 48 h of postincubation in water. The chromosome morphology was analyzed and the content of mitotic and phase indexes were done. Individual compounds, oxindole alkaloids, phenolic compounds and sugars were determined. RESULTS: In Alkaloid-Rich and Alkaloid-Free fractions (different in chemical composition) we observed condensation and contraction of chromosomes (more in Alkaloid-Rich) with retardation and/or inhibition of mitoses and changed mitotic phases. Postincubation reversed results in the highest concentration which was lethal (in mostly Alkaloid-Rich fraction). CONCLUSIONS: Our studies indicate that different action can depend on different groups of active compounds in a preparation either containing alkaloids or not. Other fraction analysis may be useful in the future.
Assuntos
Alcaloides/farmacologia , Antimitóticos/farmacologia , Unha-de-Gato/química , Núcleo Celular/efeitos dos fármacos , Cromossomos de Plantas/efeitos dos fármacos , Cebolas/efeitos dos fármacos , Alcaloides/isolamento & purificação , Antimitóticos/isolamento & purificação , Núcleo Celular/genética , Núcleo Celular/metabolismo , Cromossomos de Plantas/ultraestrutura , Meristema/efeitos dos fármacos , Meristema/genética , Metáfase/efeitos dos fármacos , Índice Mitótico , Cebolas/genética , Casca de Planta , Extratos Vegetais/química , Extratos Vegetais/farmacologiaRESUMO
AIM: To investigate the in vitro activity of silver NPs (AgNPs) against pathogenic microalgae of the Prototheca genus. MATERIALS & METHODS: The antialgal potential of AgNPs against Prototheca species of both clinical and environmental origin was assessed from minimum inhibitory (algistatic) and algicidal concentrations. The in vitro cytotoxicity of AgNPs against bovine mammary epithelial cell line was evaluated by means of the standard MTT assay. RESULTS: AgNPs showed a strong killing activity toward Prototheca algae, as the minimal algicidal concentration (MAC) values matched perfectly the corresponding minimum inhibitory concentration (MIC) values for all species (MAC = MIC, 1-4 mg/l), except P. stagnora (MIC > 8 mg/l). The concentrations inhibitory to pathogenic Prototheca spp. (MIC, 1-4 mg/l) were below the concentrations at which any toxicity in epithelial cells could be observed (CC20 > 6 mg/l). CONCLUSION: The study emphasizes the potential of AgNPs as a new therapeutic tool for the management of Prototheca infections.
Assuntos
Nanopartículas Metálicas/química , Microalgas/efeitos dos fármacos , Prototheca/efeitos dos fármacos , Prata/química , Prata/farmacologia , Animais , Bovinos , Linhagem Celular , Testes de Sensibilidade MicrobianaRESUMO
Silver nanoparticles (AgNPs) are promising new antimicrobial agents against a wide range of skin and mucosal pathogens. However, their interaction with the immune system is currently not fully understood. Dendritic cells (DCs) are crucial during development of T cell-specific responses against bacterial and viral pathogens. We have previously shown that tannic acid-modified silver nanoparticles (TA-AgNPs) consist of a promising microbicide against HSV-2. The aim of this study was to compare the ability of TA-AgNPs or TA-AuNPs of similar sizes (TA-Ag/AuNPs) to induce DCs maturation and activation in the presence of HSV-2 antigens when used at non-toxic doses. First, we used JAWS II DC line to test toxicity, ultrastructure as well as activation markers (MHC I and II, CD40, CD80, CD86, PD-L1) and cytokine production in the presence of TA-Ag/AuNPs. Preparations of HSV-2 treated with nanoparticles (TA-Ag/AuNPs-HSV-2) were further used to investigate HSV-2 antigen uptake, activation markers, TLR9 expression, and cytokine production. Additionally, we accessed proliferation and activation of HSV-2-specific T cells by DCs treated with TA-AgNP/AuNPs-HSV-2. We found that both TA-AgNPs and TA-AuNPs were efficiently internalized by DCs and induced activated ultrastructure. Although TA-AgNPs were more toxic than TA-AuNPs in corresponding sizes, they were also more potent stimulators of DCs maturation and TLR9 expression. TA-Ag/AuNPs-HSV-2 helped to overcome inhibition of DCs maturation by live or inactivated virus through up-regulation of MHC II and CD86 and down-regulation of CD80 expression. Down-regulation of CD40 expression in HSV-2-infected DCs was reversed when HSV-2 was treated with TA-NPs sized >30 nm. On the other hand, small-sized TA-AgNPs helped to better internalize HSV-2 antigens. HSV-2 treated with both types of NPs stimulated activation of JAWS II and memory CD8+ T cells, while TA-AgNPs treatment induced IFN-γ producing CD4+ and CD8+ T cells. Our study shows that TA-AgNPs or TA-AuNPs are good activators of DCs, albeit their final effect upon maturation and activation may be metal and size dependent. We conclude that TA-Ag/AuNPs consist of a novel class of nano-adjuvants, which can help to overcome virus-induced suppression of DCs activation.