Comparison of alcian blue and ruthenium red effects on preservation of outer envelope ultrastructure in methanotrophic bacteria.

Alcian blue (AB) and ruthenium red (RR) effects on ultrastructural preservation of the bacterial cell envelope of methanotrophs are compared. A previous successful method with RR that enhanced preservation of outer envelope layers in two representative methanotroph species is applied to other genera and species of methanotropic bacteria. Alcian blue is substituted for RR in this en bloc protocol. The effect of AB on preservation of these layers is assessed at the ultrastructural level and compared to RR for all species examined. Further, comparison with freeze etch and a fixation in the absence of either RR or AB is made. Both RR and AB are found to aid preservation and help visualize additional components of the cell envelope which are lost or minimized in a standard fixation not employing these cationic reagents. For some species, images obtained are similar between RR and AB procedures and agree with images seen by freeze etch. For other species, AB preserves extended filamentous material that is partially condensed even with the use of RR. Thus, use of AB improves the preservation of outer envelope structure in these organisms equally or more effectively than use of RR.

Effect of fixation-resin combinations and ruthenium red on elucidating outer envelope structure and surface morphology of two methanotrophic bacteria.

We examined the ultrastructure of the cell envelope in Type I, Methylomonas albus (BG8), and Type II, Methylosinus trichosporium (OB3b), methane-oxidizing bacteria by using different fixatives, ruthenium red (RR) combinations and resins. We compared LR White and Spurr embedments with the following fixations: glutaraldehyde/OsO4, two glutaraldehyde-paraformaldehyde, and two different en bloc ruthenium red procedures, one utilizing OsO4 and the other with glutaraldehyde/OsO4 in sequential fixation. These fixations were also studied by scanning electron microscopy (SEM). Unfixed cells prepared by freeze etch were used for comparison. Transmission electron microscopy of BG8 embedded in LR White resin (with or without red0 preserved a layer of cup-like structures that were not seen in Spurr resin-embedded cells unless ruthenium red was used. For OB3b, the second RR method preserved beads and filaments where only "spike-like" structures were seen in all other fixations in both resins. By SEM, all fixations preserved a capsular slime layer of BG8 that was removed from some cells by both RR methods. In all SEM fixations, a bead layer was preserved in OB3b that was enhanced by RR. Filaments seen by freeze-etch and thin-section techniques were not seen in SEM. Presence or absence of particular envelope structures in these methanotrophs is dependent on the combination of fixatives and/or resins employed and is species-specific. The chemical preparation methods used resulted in enhanced understanding of the structure and composition of the cell envelope.

Ultrastructural localization of cellulase in Trichoderma reesei using immunocytochemistry and enzyme cytochemistry.

Two components of the cellulase complex (E.C. 3.2.1.4) of the fungus Trichoderma reesei were localized at the ultrastructural level. Immunocytochemistry and enzyme cytochemistry demonstrated that cellobiohydrolase and beta-1,4 glucanase were localized within cisternae of endoplasmic reticulum and within membrane complexes of cellulose-grown hyphae. Both enzymes were also present in the culture medium. Glucose-grown control hyphae lacked enzyme-specific staining, and no enzyme activity was detected in the growth medium.

Individual differences in adult foveal visual asymmetries.

The existence and nature of asymmetries in the recognition of elements of a visually presented array have been topics of dispute. In the present study, 32 adults responded to a single vertical or horizontal bar embedded in one circle of a 5 times 7 array of circles by touching a plate corresponding to the orientation of the bar. Two thirds of the subjects were left superior, while one third were right superior. Performance was in general top superior and decreased with increasing distance from the center. Possible explanations for these asymmetries are examined in terms of sequential processing, acuity dominance, hemispheric specialization, and selective attention.