Dibothriocephalus ditremus and Dibothriocephalus latus are diphyllobothriidean tapeworms autochthonous to Europe. Their larval stages (plerocercoids) may seriously alter health of their intermediate fish hosts (D. ditremus) or cause intestinal diphyllobothriosis of the final human host (D. latus). Despite numerous data on the internal structure of broad tapeworms, many aspects of the morphology and physiology related to host–parasite co-existence remain unclear for these 2 species. The main objective of this work was to elucidate functional morphology of the frontal part (scolex) of plerocercoids, which is crucial for their establishment in fish tissues and for an early attachment in final hosts. The whole-mount specimens were labelled with different antibodies and examined by confocal microscope to capture their complex 3-dimensional microanatomy. Both species exhibited similar general pattern of immunofluorescent signal, although some differences were observed. In the nervous system, FMRF amide-like immunoreactivity (IR) occurred in the bi-lobed brain, 2 main nerve cords and surrounding nerve plexuses. Differences between the species were found in the structure of the brain commissures and the size of the sensilla. Synapsin IR examined in D. ditremus occurred mainly around FMRF amide-like IR brain lobes and main cords. The unexpected finding was an occurrence of FMRF amide-like IR in terminal reservoirs of secretory gland ducts and excretory canals, which has not been observed previously in any tapeworm species. This may indicate that secretory/excretory products, which play a key role in host–parasite relationships, are likely to contain FMRF amide-related peptide/s.

Alga in the genus Chroothece have been reported mostly from aquatic or subaerial continental environments, where they grow in extreme conditions. The strain Chroothece mobilis MAESE 20.29 was exposed to different light intensities, red and green monochromatic light, ultraviolet (UV) radiation, high nitrogen concentrations, and high salinity to assess the effect of those environmental parameters on its growth. Confocal laser scanning microscopy (CLSM) was used as an “in vivo” noninvasive single-cell method for the study. The strain seemed to prefer fairly high light intensities and showed a significant increase in allophycocyanin (APC) and chlorophyll a [photosystem I (PSI) and photosystem II (PSII)] fluorescence with 330 and 789 μM/cm2/s intensities. Green monochromatic light promoted a significant increase in the fluorescence of APC and chlorophyll a (PSI and PSII). UV-A significantly decreased phycocyanin and increased APC, while UV-A + B showed a greater decreasing effect on c-Phycocyanin but did not significantly change concentrations of APC. The increase in nitrogen concentration in the culture medium significantly and negatively affected all pigments, and no effect was observed with an increase in salinity. Our data show that CLSM represents a very powerful tool for ecological research of microalgae in small volumes and may contribute to the knowledge of phycobiliproteins in vivo behavior and the parameters for the large-scale production of these pigments.

Imaging of cellular layers in a gut-on-a-chip system has been confined to two-dimensional (2D)-imaging through conventional light microscopy and confocal laser scanning microscopy (CLSM) yielding three-dimensional- and 2D-cross-sectional reconstructions. However, CLSM requires staining and is unsuitable for longitudinal visualization. Here, we compare merits of optical coherence tomography (OCT) with those of CLSM and light microscopy for visualization of intestinal epithelial layers during protection by a probiotic Bifidobacterium breve strain and a simultaneous pathogen challenge by an Escherichia coli strain. OCT cross-sectional images yielded film thicknesses that coincided with end-point thicknesses derived from cross-sectional CLSM images. Light microscopy on histological sections of epithelial layers at the end-point yielded smaller layer thicknesses than OCT and CLSM. Protective effects of B. breve adhering to an epithelial layer against an E. coli challenge included the preservation of layer thickness and membrane surface coverage by epithelial cells. OCT does not require staining or sectioning, making OCT suitable for longitudinal visualization of biological films, but as a drawback, OCT does not allow an epithelial layer to be distinguished from bacterial biofilms adhering to it. Thus, OCT is ideal to longitudinally evaluate epithelial layers under probiotic protection and pathogen challenges, but proper image interpretation requires the application of a second method at the end-point to distinguish bacterial and epithelial films.

Statoliths are the only hard structures in the gelatinous bell of most scyphozoan medusae and investigations on these structures could promote investigations of the understudied population dynamics and phylogeny of jellyfish. We examined the statoliths of Aurelia aurita jellyfish of different ages by light microscopic and microtomographic measurements supplemented by scanning electron microscopy. The morphometric analyses confirmed that statolith numbers and sizes increase during jellyfish development and revealed that newly-formed statoliths had similar shapes that may change during statolith growth. Nevertheless, most statoliths had a typical compact rod shape with an aspect ratio of 1–2.5 at all ages and we suggest that the composition of statolith shapes may be taxa specific. We developed a new approach allowing exact measurements of statolith growth for the first time. The application of calcein as a fluorescent marker resulted in clear fluorescent lines within the statoliths, allowing calculations of the statolith side face growth increments (0.1 µm/day; n=252). A single-crystal analysis revealed that the calcein incubation did not affect the statolith crystal structure. In conclusion, calcein labeling is an excellent method to follow the growth of bassanite statoliths.

Labeling of organelles for microscopy is achieved generally by specific dyes that either accumulate in a cellular compartment such as cyanine dyes in mitochondria or are only fluorescent under specific conditions such as the low pH in the lysosome. Here we demonstrate that Cy5—a fluorescent molecule that does not enter cells by itself—can be loaded into cells by attaching a short oligonucleotide. This very inexpensive labeling procedure can be done in the presence of serum. Therefore, very sensitive cell types should also be amenable to this procedure, and longer observations can be achieved compared to other commercially available dyes as the labeling reagent does not need to be washed out. This also points to the pitfall of using fluorescently labeled oligonucleotides for live cell imaging where the oligonucleotide is supposed to detect a specific target sequence in its subcellular distribution.

We describe the architecture of nitrifying bacteria biofilms immobilized on a three-dimensional (3D) polyurethane foam that permits efficient water flow through a bioreactor. The 3D spatial organization of immobilized bacterial colonies is characterized on three resolution levels with X-ray tomography, light confocal microscopy, and scanning electron microscopy (SEM). Using these techniques we demonstrate biofilm distribution in the foam and the existence of several modes of binding of bacteria to the foam. Computed X-ray tomography permits observation of the distribution of the biofilm in the whole open cellular polyurethane material volume and estimation of biofilm volume. SEM and confocal laser scanning microscopy techniques permit 3D visualization of biofilm structure. Three distinct immobilization patterns could be observed in the open cellular polyurethane material: (1) large irregular aggregates of bacterial biofilm that exist as irregular biofilm fragments, rope-like structures, or biofilm layers on the foam surface; (2) spherical (pom-pom) aggregates of bacteria localized on the external surface of biofilm; and (3) biofilm threads adherent to the surface of polyurethane foam. Finally, we demonstrate that immobilized bacteria exhibit metabolic activity and growth.