This study highlights the role of specific outer bacterial structures, such as the glycocalix, in calcium carbonate crystallization in vitro. We describe the formation of calcite crystals by extracellular polymeric materials, such as exopolysaccharides (EPS) and capsular polysaccharides (CPS) isolated from Bacillus firmus and Nocardia calcarea. Organic matrices were isolated from calcifying bacteria grown on synthetic medium—in the presence or absence of calcium ions—and their effect on calcite precipitation was assessed. Scanning electron microscopy observations and energy dispersive X-ray spectrometry analysis showed that CPS and EPS fractions were involved in calcium carbonate precipitation, not only serving as nucleation sites but also through a direct role in crystal formation. The utilization of different synthetic media, with and without addition of calcium ions, influenced the biofilm production and protein profile of extracellular polymeric materials. Proteins of CPS fractions with a molecular mass between 25 and 70 kDa were overexpressed when calcium ions were present in the medium. This higher level of protein synthesis could be related to the active process of bioprecipitation.

Fish renal morphology has been studied since the 1960’s, but is still incomplete for most species, even for some of the most studied. The functional and morphological diversity and specialization of the fish kidney, due to environmental adaptations, makes this a large subject that remains understudied. References to studies on fish renal structure are given in Table 1. Although authors as Hentschel and Elger contributed to the knowledge of fish renal morphology in a comparative perspective, the knowledge concerning this issue is still reduced. Here we give a short comparative summary of fish renal morphology, using as baseline model the brown trout.

We examine simulated electron microdiffraction patterns from models of thin polycrystalline silicon. The models are made by a Voronoi tessellation of random points in a box. The Voronoi domains are randomly selected to contain either a randomly-oriented cubic crystalline grain or a region of continuous random network material. The microdiffraction simulations from coherent probes of different widths are computed at the ideal kinematical limit, ignoring inelastic and multiple scattering. By examining the normalized intensity variance that is obtained in fluctuation electron microscopy experiments, we confirm that intensity fluctuations increase monotonically with the percentage of crystalline grains in the material. However, anomalously high variance is observed for models that have 100% crystalline grains with no imperfections. We confirm that the reduced normalized variance, V(k,R) − 1, that is associated with four-body correlations at scattering vector k, varies inversely with specimen thickness. Further, for probe sizes R larger than the mean grain size, we confirm that the reduced normalized variance obeys the predicted form given by Gibson et al. [Ultramicroscopy, 83, 169–178 (2000)] for the kinematical coherent scattering limit.

Extended abstract of a paper presented at Microscopy and Microanalysis 2012 in Phoenix, Arizona, USA, July 29 – August 2, 2012.

The centrosome is the principal microtubule organization center in cells, giving rise to microtubule-based organelles (e.g., cilia, flagella). The aim was to study the osteocyte centrosome morphology at an ultrastructural level in relation to its mechanosensitive function. Osteocyte centrosomes and cilia in tibial cortical bone were explored by acetylated alpha-tubulin (AαTub) immunostaining under confocal microscopy. For the first time, fine ultrastructure and spatial orientation of the osteocyte centrosome were explored by transmission electron microscopy on serial ultrathin sections. AαTub-positive staining was observed in 94% of the osteocytes examined (222/236). The mother centriole formed a short primary cilium and was longer than the daughter centriole due to an intermediate zone between centriole and cilium. The proximal end of the mother centriole was connected with the surface of daughter centriole by striated rootlets. The mother centriole exhibited distal appendages that interacted with the cell membrane and formed a particular structure called “cilium membrane prolongation.” The primary cilium was mainly oriented perpendicular to the long axis of bone. Mother and daughter centrioles change their original mutual orientation during the osteocyte differentiation process. The short primary cilium is hypothesized as a novel type of fluid-sensing organelle in osteocytes.

Extended abstract of a paper presented at Microscopy and Microanalysis 2012 in Phoenix, Arizona, USA, July 29 – August 2, 2012.

With growing interest in nanostructured thin films produced by glancing angle deposition (GLAD), it becomes increasingly important to understand their overall growth mechanics and nanocolumn structure. We present a new method of isolating the individual nanocolumns of GLAD films, facilitating automated measurement of their broadening profiles. Data collected for α = 81° TiO2 vertical nanocolumns deposited across a range of substrate rotation rates demonstrates that these rates influence growth scaling parameters. Further, individual posts were found in each case that violate predicted Kardar-Parisi-Zhang growth scaling limits. The technique's current iteration is comparable to existing techniques in speed: though data were studied from 10,756 individual objects, the majority could not be confidently used in subsequent analysis. Further refinement may allow high-throughput automated film characterization and permit close examination of subtle growth trends, potentially enhancing control over GLAD film broadening and morphology.

The use of nonallergic, nontoxic, and eco-friendly natural dyes has become a matter of significant importance due to increased environmental awareness on the need to avoid hazardous synthetic dyes. This study was to determine the staining properties of the dye extract of Lonchocarpus cyanescens on histomorphology of the testis. Freshly cut leaves of L. cyanescens obtained from Akpan Ifia Inan village in Ikono local government area of Akwa Ibom state (latitude 5° 10′ 12″ N; longitude 7° 48′ 0″ E) were put into a plastic jar and boiling water was poured to cover the leaves. It was covered and left for an hour. The liquid was strained and potassium hydroxide was added to the dye water mixture to reach a pH of 9. A whisk was used to mix air into the liquid, and the mixuture was then allowed to sit until the blue indigo had settled to the bottom of the container. The dye was diluted with 70% ethanol to a concentration of 0.1 g/mL and was used to stain sections of testes. Its potential for use as a counterstain was also investigated. The testes sections were stained in shades of blue. The dye overshadowed the colors of haematoxylin and eosin. Preliminary phytochemical screening of L. cyanescens revealed that it contains alkaloids, saponins, flavonoids, and tannins.

In the last decade, the applicability of focus ion beam–field emission scanning electron microscopy (FIB-FESEM) in the biological field has begun to get relevance. Among the possibilities offered by FIB-FESEM, high-resolution three-dimensional (3D) reconstruction of biological structures is one of the most interesting. Using this tool, the 3D porosity of four different types of mouse oocyte zona pellucida (ZP) was analyzed. A surface analysis of the mouse oocyte ZP was first performed by SEM. Next, one oocyte per ZP type was selected, and an area of its ZP was completely milled, using the cut and view mode, in the FIB-FESEM. Through a 3D reconstruction of the milled area, a map of the distribution of the pores across the ZP was established and the number and volume of pores were quantified, thus enabling for the first time the study of the inner porosity of the mouse ZP. Differences in ZP porosity observed among the four types analyzed allowed us to outline a model to explain the changes that the ZP undergoes through immature, mature, predegenerative, and degenerative stages.

The distortions present in an electron diffraction pattern can be classified into two categories: one is radially invariant and the other is angularly invariant. We report a method to compensate these displacements undergone by diffraction features promoted by any kind of artifacts generated in parallel beam electron diffraction conditions. This approach is not aimed at quantifying these distortions but only intends to aid in the measurement of lattice parameters of crystals with a significant increase of accuracy and precision as compared to previous approaches. It is based on statistical estimations of the relative positions between diffraction rings and/or spots after performing a transformation of the digitalized patterns to polar coordinates. The analytical method is based on fitting a Gaussian type profile to intensity distributions. This makes it possible to determine the lattice parameters of a polycrystal or single crystal with relative errors smaller than 0.1% for diffractograms acquired in photographic films and below 0.01% for those collected in imaging plates.

The species of earthworms included in the Lumbricidae family have a very complex esophageal organ called calciferous or Morren gland, being the representatives of the genus Lumbricus the individuals where that gland is better developed. Previous studies have shown the secretion produced in the glandular structure is not crystalline but amorphous in the form of amorphous calcium carbonate (ACC) spherulites. Afterwards the spherulites undergo a crystallization process to form rhombohedral calcite crystals that finally result in macroaggregates of these crystals. This crystallization process can go through two intermediate states, vaterite or aragonite. These changes happen on the surface of the amorphous phases.

High-resolution monochromated electron energy loss spectroscopy (EELS) at subnanometric spatial resolution and <200 meV energy resolution has been used to assess the valence band properties of a distributed Bragg reflector multilayer heterostructure composed of InAlN lattice matched to GaN. This work thoroughly presents the collection of methods and computational tools put together for this task. Among these are zero-loss-peak subtraction and nonlinear fitting tools, and theoretical modeling of the electron scattering distribution. EELS analysis allows retrieval of a great amount of information: indium concentration in the InAlN layers is monitored through the local plasmon energy position and calculated using a bowing parameter version of Vegard Law. Also a dielectric characterization of the InAlN and GaN layers has been performed through Kramers-Kronig analysis of the Valence-EELS data, allowing band gap energy to be measured and an insight on the polytypism of the GaN layers.

The physiological condition of Solea solea and Solea senegalensis, caught in the natural environment and in aquaculture fish farms respectively, was determined by analyzing the histological changes occurred in the gills of the animals. These two species were used because both occupy the same ecological niche but only Solea senegalensis is reared in Portuguese aquaculture farms. The analysis was performed using 19 specimens in total from both environments collected between January and March of 2011 in two different locations of the Portuguese coast (Leixões and Aveiro, Portugal). The lesions found in the gills samples corresponded to four types of fusions: basal fusion corresponding to fusions at the base of the secondary lamella; medium fusion corresponding to fusions up to half of the secondary lamella; total fusion corresponding to fusions in the total length of the lamella; apical fusion corresponding to the complete fusion of the entire lamella. Another form of lesion found were lamellar edema which consisted of two different type: apical edema corresponding to a swelling at the apex of the secondary lamella; lamellar edema corresponding to a swelling in the full length of the tissue of the secondary lamella. The last physiological modification found was the alteration of the quantity of mucous cells found and also of mucous released in the primary and secondary lamellae of the gills. The results revealed that the most commonly type of lesions found were: basal fusion (Fig. 1. C); medium fusion (Figure 1 B); apical fusion (Figure 1 A); total fusion (Figure 1 D); lamellar edema (Figure 1 E); apical edema (Figure 1 F) and mucous cells disorganization in the gills (Figure 1 G). These data are very similar to other previously reported results in histopathologic studies, and are most likely to result from: contamination with heavy metals; organic or pesticides contamination; contamination by hydrocarbons (oils, fuels; rapid changes in the temperature of the water; the time of recovery of the fishing nets in the case of Solea solea; contamination of the ice used in the storage process of Solea solea. The histological analyses are very useful to evaluate the physiological condition of the fish. The histological analyses are a very important tool which enables the rapid development of processes and supports sound ecological management decisions in order to protect and/or recover habitats and ecosystems in which these animals live, therefore, improving their physiological condition with obvious benefits as to the human consumer is concerned. The novelty in this study was the use of bottom duelling fish and the comparison established between wild specimens and aquaculture reared specimens. The final conclusion is that several types of contamination occurred both in the natural environment as in the aquaculture fish farms, resulting in the different types of lesions and physiological changes observed during our experiment.

Following is a list of microscopy-related meetings and courses. The editors would greatly appreciate input to this list via the electronic submission form found in the MSA World-Wide Web page at www.msa.microscopy.com. We will gladly add hypertext links to the notice on the web and insert a listing of the meeting in the next issue of the Journal. Send comments and questions to Nan Yao, nyao@Princeton.edu.

This study presents the morphological description and histochemical characterization of gill filaments of the Brazilian endemic bivalve Diplodon expansus, aiming to broaden the morphological knowledge of this species and establish the structure of the gills that will serve as control in histopathological studies applied to biomonitoring. The gill filaments are divided into three zones: frontal, intermediate, and abfrontal. In the center of the filament, haemocytes circulate through the haemolymph vessel, which is internally lined by endothelium. The frontal surface of the filament is covered with cilia, the lateral surface exhibits aquifer ducts, and the abfrontal surface presents ciliated and nonciliated cells. The epithelium of the filaments is composed of ciliated cells, nonciliated absorptive cells, and mucocytes. The support of the filaments is made by two specialized structures called skeletal rod and skeletal loop. Based on the obtained information, the gill filaments of the studied species present some peculiar characteristics that are not yet reported in detail in the literature such as the simultaneous presence of skeletal rod and skeletal loop. On the other hand, the general constitution of the filament is similar to that described for both marine and limnic bivalves and seems to be suitable for ecotoxicological studies.

Self-assembled nanostructures of α,ω-dihexylsexithiophene (DH6T) formed by spreading DH6T solutions onto freshly cleaved mica surface were studied by atomic force microscopy. The effects of solvent and concentration on the nanostructures of DH6T molecules were studied. Flat, well-ordered, and platelet-like domains were observed on mica surfaces after treatment with various polar solvent solutions of DH6T. These domains form a uniform film with height of 2.4 ± 0.2 nm, which is consistent with a 45° tilt in the molecular conformation of DH6T on mica surfaces. The formation mechanism of these multilayers is discussed in detail.

Extended abstract of a paper presented at Microscopy and Microanalysis 2012 in Phoenix, Arizona, USA, July 29 – August 2, 2012.