Electron microscopic study of the effects of antimicrobial agents on the cellular architecture of Leptospira.

Morphological changes of Leptospira induced by ampicillin, streptomycin, or ciprofloxacin were studied by transmission electron microscopy after staining with uranyl acetate. Irregularities in coiling, loss of hooks, spherical swellings, membrane vesicle formation, disruption of outer membrane, exposure and detachment of flagella, penetration of dye, and lysis of cells were observed. These changes appeared in a time-dependent manner and appeared to be associated with the sites of action of antimicrobial agents.

Recombinant Porphyromonas gingivalis FimA preproprotein expressed in Escherichia coli is lipidated and the mature or processed recombinant FimA protein forms a short filament in vitro.

The gram-negative anaerobic bacterium Porphyromonas gingivalis is an etiologically important pathogen for chronic periodontal diseases in adults. Our previous study suggested that the major structural components of both Fim and Mfa fimbriae in this organism are secreted through their lipidated precursors. In this study, we constructed Escherichia coli strains expressing various fimA genes with or without the 5'-terminal DNA region encoding the signal peptide, and we determined whether lipidation of recombinant FimA proteins occurred in E.

Biogenesis of Salmonella enterica serovar typhimurium membrane vesicles provoked by induction of PagC.

Gram-negative bacteria ubiquitously release membrane vesicles (MVs) into the extracellular milieu. Although MVs are the product of growing bacteria, not of cell lysis or death, the regulatory mechanisms underlying MV formation remained unknown. We have found that MV biogenesis is provoked by the induction of PagC, a Salmonella-specific protein whose expression is activated by conditions that mimic acidified macrophage phagosomes.

Electron microscopic examination of uncultured soil-dwelling bacteria.

Bacteria living in soil collected from a rice paddy in Fukuoka, Japan, were examined by electron microscopy using a freeze-substitution fixation method. Most of the observed bacteria could be categorized, based on the structure of the cell envelope and overall morphology, into one of five groups: (i) bacterial spore; (ii) Gram-positive type; (iii) Gram-negative type; (iv) Mycobacterium like; and (v) Archaea like. However, a few of the bacteria could not be readily categorized into one of these groups because they had unique cell wall structures, basically resembling those of Gram-negative bacteria, but with the layer corresponding to the peptidoglycan layer in Gram-negative bacteria being extremely thick, like that of the cortex of a bacterial spore.

Photoactivated ethidium monoazide directly cleaves bacterial DNA and is applied to PCR for discrimination of live and dead bacteria.

Ethidium monoazide (EMA) is a DNA intercalating agent and a eukaryotic topoisomerase II poison. We found that EMA treatment and subsequent visible light irradiation (photoactivation or photolysis) shows a bactericidal effect, hence the mechanism was analyzed. When bacterial cells were treated with more than 10 microg/ml of EMA for 1 hr plus photoactivation for 20 min, cleavage of bacterial DNA was confirmed by agarose gel electrophoresis and electron microscopic studies.

Electron microscopic examination of the dormant spore and the sporulation of Paenibacillus motobuensis strain MC10.

We previously reported a new species Paenibacillus motobuensis. The type strain MC10 was stained gram-negative, but had a gram-positive cell wall structure and its spore had a characteristic star shape. The spore and sporulation process of P.

Curli fibers are required for development of biofilm architecture in Escherichia coli K-12 and enhance bacterial adherence to human uroepithelial cells.

Sessile bacteria show phenotypical, biochemical, and morphological differences from their planktonic counterparts. Curli, extracellular structures important for biofilm formation, are only produced at temperatures below 30 C in Escherichia coli K-12 strains. In this report, we show that E.

Paenibacillus motobuensis sp. nov., isolated from a composting machine utilizing soil from Motobu-town, Okinawa, Japan.

A novel bacterial strain, MC10(T), was isolated from a compost sample produced in a composting machine utilizing soil from Motobu-town, Okinawa, Japan. The isolate was Gram-negative, but produced endospores. These conflicting characters prompted a taxonomic study of the isolate.

Release of Shiga toxin by membrane vesicles in Shigella dysenteriae serotype 1 strains and in vitro effects of antimicrobials on toxin production and release.

Effects of various antimicrobials on in vitro Shiga toxin production and release by Shigella dysenteriae serotype 1 was investigated in this study with particular reference to the role of outer membrane vesicles in toxin release by the organism. Five antimicrobials, namely nalidixic acid, ciprofloxacin, norfloxacin, fosfomycin and mitomycin C, were chosen for the study and the toxin titre was measured by the reverse passive latex agglutination (RPLA) method using an available kit. Only mitomycin C was found to induce production of Shiga toxin in the bacteria and its release by outer membrane vesicles.

Legionella dumoffii DjlA, a member of the DnaJ family, is required for intracellular growth.

Legionella dumoffii is one of the common causes of Legionnaires' disease and is capable of replicating in macrophages. To understand the mechanism of survival within macrophages, transposon mutagenesis was employed to isolate the genes necessary for intracellular growth. We identified four defective mutants after screening 790 transposon insertion mutants.

Vesicle-mediated export and assembly of pore-forming oligomers of the enterobacterial ClyA cytotoxin.

The ClyA protein is a pore-forming cytotoxin expressed by Escherichia coli and some other enterobacteria. It confers cytotoxic activity toward mammalian cells, but it has remained unknown how ClyA is surface exposed and exported from bacterial cells. Outer-membrane vesicles (OMVs) released from the bacteria were shown to contain ClyA protein.

Degradation process of Mycobacterium leprae cells in infected tissue examined by the freeze-substitution method in electron microscopy.

Mycobacterium leprae cells (strain Thai-53) harvested from infected mouse foot pads were examined by electron microscopy using the freeze-substitution technique. The population of M. leprae cells from the infected tissue consisted of a large number of degraded cells and a few normal cells.

Comparative studies of the cell structures of Mycobacterium leprae and M. tuberculosis using the electron microscopy freeze-substitution technique.

The cell envelope and cytoplasmic architecture of the Mycobacterium leprae Thai-53 strain were examined using the freeze-substitution technique of electron microscopy and compared with those of the M. tuberculosis H37Rv strain. Both strains had similarly multilayered envelope architectures composed of an electron-translucent layer, a peptidoglycan layer and the plasma membrane, from outside to inside.

The iron-binding protein Dps confers hydrogen peroxide stress resistance to Campylobacter jejuni.

We identified and characterized the iron-binding protein Dps from Campylobacter jejuni. Electron microscopic analysis of this protein revealed a spherical structure of 8.5 nm in diameter, with an electron-dense core similar to those of other proteins of the Dps (DNA-binding protein from starved cells) family.

Morphological variety of intracellular microcolonies of Legionella species in Vero cells.

Intracellular microcolonies of six Legionella species growing in Vero cells showed distinctly varied morphologies. The varieties were observed by light microscopy of Gimenez-stained, Legionella-infected Vero cells and by electron microscopy (EM). Legionella pneumophila Philadelphia-1 formed needle-shaped crystal-like microcolonies.

Virus entry is a major determinant of cell tropism of Edmonston and wild-type strains of measles virus as revealed by vesicular stomatitis virus pseudotypes bearing their envelope proteins.

The Edmonston strain of measles virus (MV) that utilizes the human CD46 as the cellular receptor produced cytopathic effects (CPE) in all of the primate cell lines examined. In contrast, the wild-type MV strains isolated in a marmoset B-cell line B95a (the KA and Ichinose strains) replicated and produced CPE in some but not all of the primate lymphoid cell lines. To determine the mechanism underlying this difference in cell tropism, we used a recently developed recombinant vesicular stomatitis virus (VSV) containing as a reporter the green fluorescent protein gene in lieu of the VSV G protein gene (VSVDeltaG*).

A comparison of solid and liquid media for resuscitation of starvation- and low-temperature-induced nonculturable cells of Aeromonas hydrophila.

Like many other gram-negative bacteria, starved cells of Aeromonas hydrophila can be induced into a viable but nonculturable (VBNC) state by incubation at low temperature, as shown here by using various bacterial enumeration methods. Starved A. hydrophila strain HR7 cells at 4 degrees C reached the nonculturable stage in about 45 days.

Resuscitation of viable but nonculturable cells of Vibrio parahaemolyticus induced at low temperature under starvation.

Vibrio parahaemolyticus is known to exist in a viable but nonculturable state when incubated at low temperature under starvation. It has long been debated whether the culturable cells which appear after temperature upshift are the result of true resuscitation or regrowth of a few residual culturable cells. Starved V.

Restoration of culturability of starvation-stressed and low-temperature-stressed Escherichia coli O157 cells by using H2O2-degrading compounds.

Late-exponential-phase cells of Escherichia coli O157:H- strain E32511/HSC became nonculturable in sterilized distilled water microcosms at 4 degrees C. Plate counts declined from 3 x 10(6) to less than 0.1 CFU/ml in about 21 days.

Isolation and characterization of rugose form of Vibrio cholerae O139 strain MO10.

An extracellular exopolysaccharide (slime) is produced by Vibrio cholerae O139 MO10 in response to nutrient starvation. The presence of this slime layer on the cell surface and its subsequent release have been shown to be associated with biofilm formation and the change from a normal smooth colony morphology to a rugose one. An immunoelectron microscopic examination demonstrated that there is an epitope common to the exopolysaccharide antigen of V.

Vibrio cholerae O1 strain TSI-4 produces the exopolysaccharide materials that determine colony morphology, stress resistance, and biofilm formation.

Vibrio cholerae O1 strain TSI-4 (El Tor, Ogawa) can shift to a rugose colony morphology from its normal translucent colony morphology in response to nutrient starvation. We have investigated differences between the rugose and translucent forms of V. cholerae O1 strain TSI-4.

Morphological features of a filamentous phage of Vibrio cholerae O139 Bengal.

A filamentous phage was isolated from carrier strain AI-1841 of Vibrio cholerae 0139 Bengal and thus was termed fs phage. The phage was measured to be approximately 1 microm in length and 6 nm in width. One end of the phage was slightly tapered and had a fibrous appendage.

Overproduction of Campylobacter ferritin in Escherichia coli and induction of paracrystalline inclusion by ferrous compound.

The ferritin gene (cft) of Campylobacter jejuni was overexpressed in cells of Escherichia coli using a T7 RNA polymerase expression system. Many round particles which were the same size as the ferritin particles purified from C. jejuni were observed in the lysate of the cft-overexpressed E.

Electron microscopy of the major outer membrane protein of Campylobacter jejuni.

The surfaces of the disrupted-cell surfaces of the Campylobacter jejuni strains FUM158432 and M1 were examined using the negative-staining technique and electron microscopy. The surfaces of the whole cells and the outer membranes were covered with small dark dots which, in some areas, were arranged in hexagonal patterns. The hexagonal arrangement was more clearly seen in extracted outer membrane.

The hydrophobic surface protein layer of enteroaggregative Escherichia coli strains.

The surface of three strains of enteroaggregative Escherichia coli (EAggEC) and three strains of enteropathogenic E. coli (EPEC) were examined using the freeze-substitution technique of electron microscopy and as a result an electron dense surface layer was found only on EAggEC strains but not on EPEC strains. The analysis of the outer membrane proteins by polyacrylamide gel electrophoresis revealed the existence of a unique 38 kDa protein in EAggEC strains.