Epifluorescent microscopy was employed to compare the bacterial live counts (BC) in the gut of two earthworm species Aporrectodea caliginosa and Lumbricus rubellus, representing different ecophysiological groups. The average number of BC was 10.9 x 10(9) g-1 dry weight in the gut of A. caliginosa, 5.9 x 10(9) in that of L. rubellus, 8.1 x 10(9) in earthworm casts and 6.0 x 10(9) in the soil. The number of BC showed a great seasonal variability in all the materials studied, exhibiting maxima in spring and autumn, and a minimum in summer. The BC increased in number during the passage of food material through the gut of both L. rubellus and A. caliginosa. The difference between BC in fore-gut and hind-gut were significantly higher in L. rubellus (4.2 x 10(9) vs. 8.8 x 10(9)) than that in A. caliginosa (10.3 x 10(9) vs. 13.4 x 10(9)). Interspecific differences in the number of BC may result from the different chemical and microbiological composition of the material consumed by earthworms as related to different feeding habits of both species.

• MeSH
• Bacteria izolace a purifikace   ultrastruktura   MeSH
• fluorescenční mikroskopie MeSH
• Oligochaeta klasifikace   mikrobiologie   MeSH
• půdní mikrobiologie MeSH
• roční období MeSH
• stravovací zvyklosti MeSH
• střeva mikrobiologie   MeSH
• trávení MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

We describe a modification of epifluorescence microscopes that allows quantitative widefield imaging of samples labeled by upconverting nanoparticles (UCNP). A top-hat illumination profile on the sample was achieved with a 980-nm laser diode by using tandem microlens arrays, a moving diffuser and a telescope, which adjusts the top-hat area to the field of view. Illumination homogeneity is a critical factor for imaging of UCNP since the intensity of their luminescence typically scales with the second power of the excitation intensity. Our illuminator is combined with the epifluorescence attachment of the microscope, allowing easy switching between observation of UCNP and traditional fluorescent dyes. Illumination profile homogeneity of about 98% was measured for objectives with magnification from 4× to 100×, and the top-hat profile was also obtained with phase contrast objectives. We demonstrate capability of the illuminator by evaluating in vitro uptake of UCNP encapsulated in oleyl-hyaluronan micelles into breast cancer cells. Micelles bearing the targeting peptide were about an order of magnitude more efficient than nontargeted micelles.

• MeSH
• fluorescenční barviva MeSH
• fluorescenční mikroskopie přístrojové vybavení   MeSH
• lasery * MeSH
• lidé MeSH
• luminiscence MeSH
• nádorové buněčné linie MeSH
• nanočástice metabolismus   ultrastruktura   MeSH
• osvětlení MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fluorescenční barviva MeSH

Spermiogenesis and the spermatozoon were studied in the digenean Mesocoelium monas Rudolphi, 1819 (from the toad Bufo sp. in Gabon). An ultrastructural study revealed that spermiogenesis follows the usual pattern found in digeneans, i.e. proximo-distal fusion of axonemes with a median cytoplasmic process followed by elongation. The spermatozoon has two fully incorporated axonemes with the 9 +"1" trepaxonematan pattern. Indirect immunofluorescence localization of tubulin and fluorescent labelling of the nucleus were used to obtain additional information on the structure of the spermatozoon. It was thus shown that one of the axonemes is slightly shorter than the other (190 versus 220 microns) and that the filiform nucleus (65 microns in length) is located at the distal extremity of the spermatozoon (220 microns in length). Various monoclonal and polyclonal antibodies, specific to alpha, beta, acetylated-alpha, or general tubulin, were used and produced similar labelling.

• MeSH
• buněčné jádro ultrastruktura   MeSH
• imunohistochemie MeSH
• ploštěnci anatomie a histologie   MeSH
• počítačové zpracování obrazu MeSH
• ropuchy parazitologie   MeSH
• spermatidy ultrastruktura   MeSH
• spermatogeneze * MeSH
• spermie ultrastruktura   MeSH
• tubulin izolace a purifikace   ultrastruktura   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• tubulin MeSH

Measuring microbial abundance in glacier ice and identifying its controls is essential for a better understanding and quantification of biogeochemical processes in glacial ecosystems. However, cell enumeration of glacier ice samples is challenging due to typically low cell numbers and the presence of interfering mineral particles. We quantified for the first time the abundance of microbial cells in surface ice from geographically distinct sites on the Greenland Ice Sheet (GrIS), using three enumeration methods: epifluorescence microscopy (EFM), flow cytometry (FCM), and quantitative polymerase chain reaction (qPCR). In addition, we reviewed published data on microbial abundance in glacier ice and tested the three methods on artificial ice samples of realistic cell (10(2)-10(7) cells ml(-1)) and mineral particle (0.1-100 mg ml(-1)) concentrations, simulating a range of glacial ice types, from clean subsurface ice to surface ice to sediment-laden basal ice. We then used multivariate statistical analysis to identify factors responsible for the variation in microbial abundance on the ice sheet. EFM gave the most accurate and reproducible results of the tested methodologies, and was therefore selected as the most suitable technique for cell enumeration of ice containing dust. Cell numbers in surface ice samples, determined by EFM, ranged from ~ 2 × 10(3) to ~ 2 × 10(6) cells ml(-1) while dust concentrations ranged from 0.01 to 2 mg ml(-1). The lowest abundances were found in ice sampled from the accumulation area of the ice sheet and in samples affected by fresh snow; these samples may be considered as a reference point of the cell abundance of precipitants that are deposited on the ice sheet surface. Dust content was the most significant variable to explain the variation in the abundance data, which suggests a direct association between deposited dust particles and cells and/or by their provision of limited nutrients to microbial communities on the GrIS.

• Klíčová slova
• Greenland Ice Sheet, epifluorescence microscopy, flow cytometry, glacier ice, microbial abundance, multivariate analysis, quantitative PCR,
• Publikační typ
• časopisecké články MeSH

Transdermal drug delivery is a passive diffusion process of an active compound through the skin which is affected by drug solubility in the multilamellar lipidic matrix of the stratum corneum (SC). Widely used non-ionic surfactants (NIS) can be added into transdermal formulations to enhance the penetration of drugs by influencing the packing of the stratum corneum lipidic matrix. Objective of our study was to analyse the interaction between selected NIS and a simple SC lipidic matrix model system using a variety of surface-sensitive techniques based on the application of Langmuir monolayers. In this work, the well-known surfactant Polysorbate 80 was compared with a modern surfactant Sucrose monolaurate. Infrared reflection-absorption spectroscopy (IRRAS) and epifluorescence microscopy provide information about the effects of those surfactants on the SC model system. Monolayer isotherms of the SC model mixture indicate a very stiff and well-packed layer, however, packing defects are evidenced in epifluorescence studies. The injection of the two NIS underneath the SC monolayers proved their potential to penetrate into the SC model at the air-water interface having a maximum insertion pressure (MIP) above the assumed lateral pressure of biological membranes. The NIS adsorbed preferentially into packing defects seen in epifluorescence microscopy studies with Sucrose monolaurate being more active than Polysorbate 80 in disordering the SC monolayer.

• Klíčová slova
• Epifluorescence microscopy, IRRAS, Langmuir monolayer, Non-ionic surfactants, Penetration enhancers, Polysorbate 80, Stratum Corneum, Sucrose monolaurate,
• MeSH
• aplikace kožní MeSH
• biologické modely MeSH
• kůže * MeSH
• lipidy MeSH
• povrchově aktivní látky * MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• lipidy MeSH
• povrchově aktivní látky * MeSH

The cortical microtubule and actin meshworks play a central role in the shaping of plant cells. Transgenic plants expressing fluorescent protein markers specifically tagging the two main cytoskeletal systems are available, allowing noninvasive in vivo studies. Advanced microscopy techniques, in particular confocal laser scanning microscopy (CLSM), spinning disk confocal microscopy (SDCM), and variable angle epifluorescence microscopy (VAEM), can be nowadays used for imaging the cortical cytoskeleton of living cells with unprecedented spatial and temporal resolution. With the aid of free computing tools based on the publicly available ImageJ software package, quantitative information can be extracted from microscopic images and video sequences, providing insight into both architecture and dynamics of the cortical cytoskeleton.

• Klíčová slova
• Actin, CLSM, Fluorescent proteins, Image analysis, ImageJ, Microtubules, SDCM, VAEM,
• MeSH
• Arabidopsis ultrastruktura   MeSH
• cytoskelet ultrastruktura   MeSH
• fluorescenční mikroskopie metody   MeSH
• konfokální mikroskopie metody   MeSH
• mikrotubuly ultrastruktura   MeSH
• počítačové zpracování obrazu metody   MeSH
• rostlinné buňky ultrastruktura   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Plant cell morphogenesis involves concerted rearrangements of microtubules and actin microfilaments. We previously reported that FH1, the main Arabidopsis thaliana housekeeping Class I membrane-anchored formin, contributes to actin dynamics and microtubule stability in rhizodermis cells. Here we examine the effects of mutations affecting FH1 (At3g25500) on cell morphogenesis and above-ground organ development in seedlings, as well as on cytoskeletal organization and dynamics, using a combination of confocal and variable angle epifluorescence microscopy with a pharmacological approach. Homozygous fh1 mutants exhibited cotyledon epinasty and had larger cotyledon pavement cells with more pronounced lobes than the wild type. The pavement cell shape alterations were enhanced by expression of the fluorescent microtubule marker GFP-microtubule-associated protein 4 (MAP4). Mutant cotyledon pavement cells exhibited reduced density and increased stability of microfilament bundles, as well as enhanced dynamics of microtubules. Analogous results were also obtained upon treatments with the formin inhibitor SMIFH2 (small molecule inhibitor of formin homology 2 domains). Pavement cell shape in wild-type (wt) and fh1 plants in some situations exhibited a differential response towards anti-cytoskeletal drugs, especially the microtubule disruptor oryzalin. Our observations indicate that FH1 participates in the control of microtubule dynamics, possibly via its effects on actin, subsequently influencing cell morphogenesis and macroscopic organ development.

• Klíčová slova
• Arabidopsis thaliana, Confocal microscopy, Cotyledon pavement cells, Cytoskeleton, Formin, Variable angle epifluorescence microscopy,
• MeSH
• aktiny metabolismus   MeSH
• Arabidopsis cytologie   účinky léků   metabolismus   MeSH
• biologické markery metabolismus   MeSH
• biologické modely MeSH
• cytoskelet účinky léků   metabolismus   MeSH
• fluorescence MeSH
• forminy MeSH
• klathrin metabolismus   MeSH
• kotyledon účinky léků   metabolismus   MeSH
• membránové proteiny metabolismus   MeSH
• mikrofilamenta účinky léků   metabolismus   MeSH
• mikrotubuly účinky léků   metabolismus   MeSH
• mutace genetika   MeSH
• proteiny huseníčku metabolismus   MeSH
• semenáček účinky léků   růst a vývoj   metabolismus   MeSH
• thioketony farmakologie   MeSH
• tvar buňky * účinky léků   MeSH
• uracil analogy a deriváty   farmakologie   MeSH
• zelené fluorescenční proteiny metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• AFH1 protein, Arabidopsis MeSH Prohlížeč
• aktiny MeSH
• biologické markery MeSH
• forminy MeSH
• klathrin MeSH
• membránové proteiny MeSH
• proteiny huseníčku MeSH
• SMIFH2 compound MeSH Prohlížeč
• thioketony MeSH
• uracil MeSH
• zelené fluorescenční proteiny MeSH

Ultrastructural analysis revealed that the spermatozoon of Discocotyle sagittata (Leuckart, 1842) is composed of two parallel axonemes, mitochondrion, nucleus and cortical microtubules. The nucleus, which occupies a central/distal position and has an unusual crescent-shaped profile, is slightly shorter than the mitochondrial rod. The two axonemes, which are of unequal length, and the cortical microtubules (up to 68 forming a continuous ring in the principal region) extend almost the entire length of the spermatozoon. A fold of the plasma membrane creates a unilateral flange or undulating membrane. Epifluorescence microscopy indicated that spermatogenesis gives rise to clusters of 64 spermatids connected to a common cytophore. Spermiogenesis and the structure of the filiform sperm of D. sagittata conform to the typical polyopisthocotylean pattern.

• MeSH
• buněčné jádro ultrastruktura   MeSH
• elektronová mikroskopie metody   MeSH
• endoplazmatické retikulum ultrastruktura   MeSH
• fluorescenční mikroskopie metody   MeSH
• Oncorhynchus mykiss parazitologie   MeSH
• ploštěnci fyziologie   ultrastruktura   MeSH
• spermatidy ultrastruktura   MeSH
• spermatocyty ultrastruktura   MeSH
• spermatogeneze * MeSH
• spermie ultrastruktura   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

This study was aimed at elucidating the fate of three important nuclear envelope components - lamins B and A/C and nucleoporin Nup160, during meiotic maturation of mouse oocytes. These proteins were localized by epifluorescence and confocal microscopy using specific antibodies in oocytes at different stages from prophase I (germinal vesicle) to metaphase II. In immature germinal vesicle oocytes, all three proteins were detected at the nuclear periphery. In metaphase I and metaphase II, lamin B co-localized with the meiotic spindle, lamin A/C was found in a diffuse halo surrounding the spindle and to a lesser degree throughout the cytoplasm, and Nup160 was concentrated to the spindle poles. To our knowledge, this is the first report on nucleoporin localization in mammalian oocytes and the first successful detection of lamins in mature oocytes. While the distribution patterns of both lamins closely paralleled the respective stages of mitosis, Nup160 localization in metaphase oocytes corresponded to that in mitotic prometaphase rather than metaphase. The peculiar distribution of this nucleoporin in oocytes may reflect its role in meiosis-specific mechanisms of spindle assembly and its regulation.

• MeSH
• buněčná diferenciace * MeSH
• jaderné proteiny metabolismus   MeSH
• konfokální mikroskopie MeSH
• laminy metabolismus   MeSH
• meióza * MeSH
• metafáze MeSH
• myši inbrední BALB C MeSH
• oocyty cytologie   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• jaderné proteiny MeSH
• laminy MeSH

Metagenome-assembled genomes (MAGs) of Asgardarchaeota have been recovered from a variety of habitats, broadening their environmental distribution and providing access to the genetic makeup of this archaeal lineage. The recent success in cultivating the first representative of Lokiarchaeia was a breakthrough in science at large and gave rise to new hypotheses about the evolution of eukaryotes. Despite their singular phylogenetic position at the base of the eukaryotic tree of life, the morphology of these bewildering organisms remains a mystery, except for the report of an unusual morphology with long, branching protrusions of the cultivated Lokiarchaeion strain "Candidatus Prometheoarchaeum syntrophicum" MK-D1. In order to visualize this elusive group, we applied a combination of fluorescence in situ hybridization and catalyzed reporter deposition (CARD-FISH) and epifluorescence microscopy on coastal hypersaline sediment samples, using specifically designed CARD-FISH probes for Heimdallarchaeia and Lokiarchaeia lineages, and provide the first visual evidence for Heimdallarchaeia and new images of a lineage of Lokiarchaeia that is different from the cultured representative. Here, we show that while Heimdallarchaeia are characterized by a uniform cellular morphology typified by a centralized DNA localization, Lokiarchaeia display a plethora of shapes and sizes that likely reflect their broad phylogenetic diversity and ecological distribution.IMPORTANCE Asgardarchaeota are considered to be the closest relatives to modern eukaryotes. These enigmatic microbes have been mainly studied using metagenome-assembled genomes (MAGs). Only very recently, a first member of Lokiarchaeia was isolated and characterized in detail; it featured a striking morphology with long, branching protrusions. In order to visualize additional members of the phylum Asgardarchaeota, we applied a fluorescence in situ hybridization technique and epifluorescence microscopy on coastal hypersaline sediment samples, using specifically designed probes for Heimdallarchaeia and Lokiarchaeia lineages. We provide the first visual evidence for Heimdallarchaeia that are characterized by a uniform cellular morphology typified by an apparently centralized DNA localization. Further, we provide new images of a lineage of Lokiarchaeia that is different from the cultured representative and with multiple morphologies, ranging from small ovoid cells to long filaments. This diversity in observed cell shapes is likely owing to the large phylogenetic diversity within Asgardarchaeota, the vast majority of which remain uncultured.

• Klíčová slova
• Asgardarchaeota, CARD-FISH, Heimdallarchaeia, Lokiarchaeia, morphology,
• MeSH
• Archaea klasifikace   genetika   MeSH
• fluorescenční mikroskopie MeSH
• fylogeneze MeSH
• geologické sedimenty mikrobiologie   MeSH
• hybridizace in situ fluorescenční metody   MeSH
• oligonukleotidové sondy genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• oligonukleotidové sondy MeSH