In a previous study, we identified Congo red-binding and -nonbinding phase variants of Escherichia coli serotype O157:H7 strain ATCC 43895. The Congo red-binding variant, strain 43895OR, produced a dry, aggregative colony that was similar to the red, dry, and rough (rdar) phenotype characteristic of certain strains of Salmonella. In contrast, variant 43895OW produced a smooth and white colony morphology. In this study, we show that, similar to rdar strains of Salmonella enterica serovar Typhimurium, strain 43895OR forms large aggregates in broth cultures, firm pellicles at the air-medium interface on glass, and dense biofilms on glass and polystyrene. However, unlike S. enterica serovar Typhimurium, strain 43895OR does not stain positive for cellulose production. When strain 43895OR was fixed on agar, scanning electron microscopy showed cells expressing extracellular matrix (ECM) containing curli fibers. Strain 43895OW was devoid of any ECM or curli fibers on agar but showed expression of curli fibers during attachment to glass. Strain 43895OR produced >4-fold-larger amounts of biofilm than strain 43895OW on polystyrene, glass, stainless steel, and Teflon; formation was >3-fold higher in rich medium than in nutrient-limited medium. Biofilm-associated cells of both strains showed statistically greater resistance (P < 0.05) to hydrogen peroxide and quaternary ammonium sanitizer than their respective planktonic cells. This study shows that the rdar phenotype of E. coli O157:H7 strain 43895OR is important in multicellular growth, biofilm formation, and resistance to sanitizers. However, the lack of cellulose production by strain 43895OR indicates important differences in the ECM composition compared to that of Salmonella.

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There are three classes of myofilaments in vertebrate smooth muscle fibers. The thin filaments correspond to actin and the thick filaments are identified with myosin. The third class of myofilaments (100 A diam) is distinguished from both the actin and the myosin on the basis of fine structure, solubility, and pattern of localization in the muscle fibers. Direct structural evidence is presented to show that the 100-A filaments constitute an integrated filamentous network with the dense bodies in the sarcoplasm, and that they are not connected to either the actin or myosin filaments. Examination of (a) isolated dense bodies, (b) series of consecutive sections through the dense bodies, and (c) redistributed dense bodies in stretched muscle fibers supports this conclusion. It follows that the 100-A filament complexes constitute a structurally distinct filamentous network. Analysis of polyacrylamide gels after electrophoresis of cell fractions that are enriched with respect to the 100-A filaments shows the presence of a new muscle protein with a molecular weight of 55,000. This protein can form filamentous segments that closely resemble in structure the native, isolated 100-,~ filaments. The results indicate that the filamentous network has a structure and composition that distinguish it from the actin and myosin in vertebrate smooth muscle.Despite the uniform optical density of the sarcoplasm in smooth muscle fibers, some classical histologists were able to demonstrate longitudinally oriented striations embedded in a homogeneous background material (36, 51). These striations were believed to reflect the presence of underlying fibrils in the sarcoplasm. This view was disputed (31), but later was proved in a study in which polarized light was used to show the fibrils in living muscle fibers (32). The structure of the fibrils was not further characterized until the advent of electron microscopy. Analysis then revealed that the sarcoplasm contained myofilaments described as ranging from 100 to 300 A in diameter (1,20,35,56). When methods of preparation affording high resolution were developed, the myofilaments were found to constitute a single class with a diameter of 60-80 A that corresponds to the contractile protein F-actin (24, 48). There was no evidence that myosin was present in filamentous form (13,40), although this protein could be isolated and induced to form filaments in vitro (25,29,48). Further work established that filaments resembling the myosin-containing filaments in striated muscles could be found in experimentally treated smooth muscle (5,30,39) and, more recently, when smooth muscles were fixed under "physiological" conditions, numerous thick myofilaments and

Three essential oils, oregano, red thyme, and cassia (100% pure oil), were encapsulated by phase separation into zein nanospheres. Topographical images indicated that the powders were made up of irregularly shaped particles ( approximately 50 mum) containing close-packed nanospheres. Approximately 31% of the oregano encapsulated particles had mean diameters greater than 100 nm compared to 19% for the zein alone particles. In vitro digestion of zein particles with pepsin at a concentration ratio of 10:1 was complete after 52 h in phosphate-citrate buffer, pH 3.5, at 37 degrees C by spectroscopic analysis. Nonenzymatic, aqueous in vitro release of essential oils from encapsulated zein particles was carried out in phosphate buffered saline at pH 7.4 and 37 degrees C. Release occurred at varying rates over 20 h probably from different locations within the closely packed nanospheres of different sizes. Gel electrophoresis SDS-PAGE of zein incubated with freeze-dried swine manure solids at 37 degrees C indicated that preformed microbial enzymes capable of digesting zein within minutes were present in the manure. Except for differences in size of nanospheres, no structural differences were resolved by several microscopic methods, suggesting that the oil and proteins phases were blended during phase separation.

The intermolecular and interfibrillar spacings of collagen in bovine corneal stroma have been measured as a function of tissue hydration. Data were recorded from low- and high-angle x-ray diffraction patterns obtained using a high intensity synchrotron source. The most frequently occurring interfibrillar spacing varied from 34 nm in dry corneas to 76 nm at H = 5 (the hydration, H, is defined as the ratio of the weight of water to the dry weight). The most frequently occurring intermolecular Bragg spacing increased from 1.15 nm (dry) to approximately 1.60 nm at normal hydration (H approximately 3.2) and continued to increase only slowly above normal hydration. Most of the increase in the intermolecular spacing occurred between H = O and H = 1. Over this hydration range the interfibrillar and intermolecular spacings moved in tandem, which suggests that the initial water goes equally within and between the fibrils. Above H = 1 water goes preferentially between the fibrils. The results suggest that, even at normal hydration, water does not fill the interfibrillar space uniformly, and a proportion is located in another space or compartment. In dried-then-rehydrated corneas, a larger proportion of the water goes into this other compartment. In both cases, it is possible to postulate a second set or population of fibrils that are more widely and irregularly separated and therefore do not contribute significantly to the diffraction pattern.

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A new medium designated Liber A has been designed and used to successfully cultivate all three 'Candidatus Liberibacter spp.,' the suspect causative agents of huanglongbing (HLB) in citrus. The medium containing citrus vein extract and a growth factor sustained growth of 'Ca. Liberibacter spp.' for four or five single-colony transfers before viability declined. Colonies, positive for 'Ca. L. asiaticus' by a 16s-based rDNA real-time polymerase chain reaction (RT-PCR) assay and sequencing, were irregular-shaped, convex, and 0.1 to 0.3 mm after 3 to 4 days. Suspect 'Ca. L. asiaticus' and 'Ca. L. americanus' cells were observed in infected tissue and on agar culture by scanning electron microscopy. The cells were ovoid to rod shaped, 0.3 to 0.4 by 0.5 to 2.0 microm, often with fimbriae-like appendages. Two strains of 'Ca. L. asiaticus' and one of 'Ca. L. americanus' grown on Liber A medium were pathogenic on citrus and could be isolated from noninoculated tissues of inoculated trees and seedlings 9 and 2 months later, respectively. The identity was confirmed by RT-PCR and 16s rDNA sequencing. This is the first report of the cultivation and pathogenicity of 'Ca. L. asiaticus' and 'Ca. L. americanus' associated with symptoms of HLB.