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We evaluated the performance of four anti-fading agents during acquisition of multiple optical sections near the widest diameter of Drosophila accessory gland nuclei using indirect immunofluorescence and confocal laser scanning microscopy. Two commercially available agents, Vectashield and SlowFade showed anti-fading properties that alleviated fluorochrome fading associated with the acquisition of multiple fluorescent optical Z-series from a single specimen by a confocal laser scanning system. Using these reagents, we were able to colocalize polypeptides through immunostained whole Drosophila nuclei.

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Strain 831-C4T (T = type strain), isolated from the surface of lettuce plants (Lactuca sativa) obtained from a retail food market, was shown to be a sterol-requiring mollicute. Morphological examination of this organism by electron and dark-field microscopic techniques showed that it consists of small, nonhelical, nonmotile, pleomorphic coccoid cells, with individual cells surrounded by a single cytoplasmic membrane. No evidence of a cell wall was observed. The organism grew rapidly in all conventional culture medium formulations for mollicutes in either aerobic or anaerobic environments. The optimum temperature for growth was 30 degrees C, but multiplication occurred at 18 to 37 degrees C. Strain 831-C4T catabolized glucose, but hydrolysis of arginine or urea could not be demonstrated. The genome size of strain 831-C4T was determined to be about 569 megadaltons, while the base composition (guanine-plus-cytosine content) of the DNA was 30.0 mol%. Recent studies in which we compared the 16S rRNA sequences of strain 831-C4T with those of more than 40 other mollicutes indicated that this organism is phylogenetically related to the Spiroplasma-Mycoplasma mycoides clade. Strain 831-C4T was serologically unrelated to the type strains of previously described Mycoplasma species and to 18 other unclassified sterol-requiring isolates cultivated from various animal, plant, or insect sources. Strain 831-C4T (= ATCC 49193) is the type strain of Mycoplasma lactucae sp. nov.

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Strain PYAN-1T (T = type strain), which was isolated from a pupal gut of the firefly beetle Pyractonema angulata, and strains PIMN-1T and PIPN-2T, which were isolated from guts of adult Photinus marginalis and Photinus pyralis fireflies, respectively, were demonstrated to be sterol-requiring mollicutes. Cells of the three strains were shown by electron and dark-field microscopy to be small, pleomorphic, nonhelical, nonmotile bodies surrounded by single membranes. No evidence of a cell wall was observed, and the organisms were not susceptible to 500 U of penicillin per ml. The three strains grew rapidly in SP-4 broth medium. Strains PIMN-1T and PIPN-2T grew in medium supplemented with bovine serum fraction, but strain PYAN-1T did not. All three strains grew on solid media when the cultures were incubated aerobically, but only strains PYAN-1T and PIPN-2T formed colonies when anaerobic conditions were employed. The three strains catabolized glucose but hydrolyzed neither arginine nor urea. All of the strains grew at temperatures of 18 to 32 degrees C; strains PYAN-1T and PIMN-1T also grew at 10 degrees C. The optimal temperature for growth for strains PYAN-1T and PIPN-2T was 30 degrees C; strain PIMN-1T grew equally well at 30 or 32 degrees C. None of the three strains grew at 37 degrees C. The genome sizes of strains PYAN-1T, PIMN-1T, and PIPN-2T were about 527 (478 to 589), 570 (480 to 630), and 762 (635 to 871) megadaltons, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

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Although nearly two decades have passed since de Villafranca (1961) described A-band shortening, controversy persists. Here we will review the data which has been amassed since de Villafranca 's description. We will conclude that A-bands and thick filaments shorten during sarcomere shortening in Limulus striated muscle. Further we will suggest that two machines operate in this muscle: a tension generating sliding filament system and a tension generating thick filament shortening system. Also we will suggest a mechanism of force generation of the filament shortening system and provide evidence for a cycling bridge mechanism for this muscle.

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Photoelectron count autocorrelation function of light scattered by Limulus thick myofilament suspensions was measured as a function of scattering angle in the relaxed, activated and rerelaxed states. By using the cumulants method of data analysis, the average linewidth over large ranges of KL (up to 120), has been calculated with K and L being, respectively, the magnitude of the momentum transfer vector and the length of the myofilament. We have observed a dramatic increase in the average linewidth denoting the presence of additional high frequency components for the myofilament suspension on the activated state. By confirming our results on the size of the myofilaments from electron micrographs, we are able to attribute the high frequency (kHz) components to the "correlated" cross-bridge motions, representing, to our knowledge, the first direct experimental evidence of such movements in isolated thick myofilament suspensions.

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Here we describe the change in thick filament length in striated muscle of Limulus, the horseshoe crab. Long thick filaments (4.0 microns) are isolated from living, unstimulated Limulus striated muscle while those isolated from either electrically or K+-stimulated fibers are significantly shorter (3.1 microns) (P less than 0.001). Filaments isolated from muscle glycerinated at long sarcomere lengths are long (4.4 microns) while those isolated from muscle glycerinated at short sarcomere lengths are short (2.9 microns) and the difference is significant (P less than 0.001). Thin filaments are 2.4 microns in length. The shortening of thick filaments is related to the wide range of sarcomere lengths exhibited by Limulus telson striated muscle.

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The musculature of the telson of Limulus polyphemus L. consists of three dorsal muscles: the medial and lateral telson levators and the telson abductor, and one large ventral muscle; the telson depressor, which has three major divisions: the dorsal, medioventral, and lateroventral heads. The telson muscles are composed of one type of striated muscle fiber, which has irregularly shaped myofibrils. The sarcomeres are long, with discrete A and I and discontinuous Z bands. M lines are not present. H zones can be identified easily, only in thick (1.0 microm) longitudinal sections or thin cross sections. In lengthened fibers, the Z bands are irregular and the A bands appear very long due to misalignment of constituent thick filaments. As the sarcomeres shorten, the Z lines straighten somewhat and the thick filaments become more aligned within the A band, leading to apparent decrease in A band length. Further A band shortening, seen at sarcomere lengths below 7.4 microm may be a function of conformational changes of the thick filaments, possibly brought about by alterations in the ordering of their paramyosin cores.

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