Mutual Relationship between Antibiotics and Resting Spores of <i>Bacillus subtilis</i>: Binding of Cyclic Polypeptide and Aminoglycoside Antibiotics to Spores and Their Inhibitory Effect on Outgrowth and Vegetative Growth

Tochikubo, Kunio; Hayakawa, Yoshiaki; Kojima, Kiyohide

doi:10.1111/j.1348-0421.1981.tb00015.x

Cited by 10 publications

(18 citation statements)

References 19 publications

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“…Release of bacilli from phagocytes during an in vitro anthrax infection of RAW 264.7 cells was observed directly using light microscopy. To ensure that the observed events were dependent only on internalized endospores, infected Mf were treated with gentamicin to kill extracellular bacteria and to inhibit vegetative growth of unphagocytosed spores (Tochikubo et al, 1981). All observed events appeared indistinguishable with or without addition of the drug (gentamicin-free images not depicted).…”

Section: Resultsmentioning

confidence: 99%

Early Bacillus anthracis-macrophage interactions: intracellular survival and escape

et al. 2000

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This study describes early intracellular events occurring during the establishment phase of Bacillus anthracis infections. Anthrax infections are initiated by dormant endospores gaining access to the mammalian host and becoming engulfed by regional macrophages (Mφ). During systemic anthrax, late stage events include vegetative growth in the blood to very high titres and the synthesis of the anthrax exotoxin complex, which causes disease symptoms and death. Experiments focus on the early events occurring during the first few hours of the B. anthracis infectious cycle, from endospore germination up to and including release of the vegetative cell from phagocytes. We found that newly vegetative bacilli escape from the phagocytic vesicles of cultured Mφ and replicate within the cytoplasm of these cells. Release from the Mφ occurs 4–6 h after endospore phagocytosis, timing that correlates with anthrax infection of test animals. Genetic analysis from this study indicates that the toxin plasmid pXO1 is required for release from the Mφ, whereas the capsule plasmid pXO2 is not. The transactivator atxA, located on pXO1, is also found to be essential for release, but the toxin genes themselves are not required. This suggests that Mφ release of anthrax bacilli is atxA regulated. The putative ‘escape’ genes may be located on the chromosome and/or on pXO1.

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Section: Resultsmentioning

confidence: 99%

Early Bacillus anthracis-macrophage interactions: intracellular survival and escape

et al. 2000

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“…Judging from the morphological changes and the macromolecular synthesis, the cells derived from kanamycintreated resting spores may be inhibited at a late stage of outgrowth, whereas those from gentamicin-treated resting spores are inhibited at a middle stage of outgrowth. This difference between the antibiotics seems to be due to their antibacterial activity but not to a combined effect because of the difference in their MIC (22) and because the resting spores cultured in the presence of kanamycin showed the thickest cell wall around the MIC and the degree of thickness decreased with an increase in concentration of this antibiotic.…”

Section: Discussionmentioning

confidence: 99%

“…Other samples were obtained as follows: about 2.3 X 10 8 resting spores (per ml) were inoculated into heart infusion broth containing kanamycin at final concentrations between 1 and 10 Itg/ml and shaken at 37 C for 15 hr. The minimum inhibitory concentration (MIG) for the same number of resting spores was 6.3 Itg/ml (22).…”

Section: Methodsmentioning

confidence: 99%

“…Resting spores of B. subtilis PCI 219 were used in this study. The method of cleaning spore preparations, the treatment of resting spores with various antibiotics, and the growth conditions of untreated and antibiotic-treated resting spores have already been described (22). The antibiotics examined were polymyxin B, gramicidin S, streptomycin, kanamycin, and gentamicin.…”

Section: Methodsmentioning

confidence: 99%

“…Recently, however, our laboratory has demonstrated that cyclic polypeptide antibiotics such as polymyxin B, colistin, and gramicidin S, and aminoglycoside antibiotics such as streptomycin, kanamycin, and gentamicin bind to the resting spores of B. subtilis and consequently act as sporocidal or sporostatic agents under certain limited conditions (21,22). As a natural course of events, it has been necessary to clarify the mechanism of inhibition by the above antibiotics of outgrowth and vegetative growth after germination.…”

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Mutual Relationship between Antibiotics and Resting Spores of Bacillus subtilis: Morphological Changes and Macromolecular Synthesis after Germination of Spores Treated with Cyclic Polypeptide and Aminoglycoside Antibiotics

Hayakawa

Tochikubo

Kozuka

1981

Microbiology and Immunology

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Morphological changes and synthesis of DNA, RNA, protein, and cell wall were investigated during germination of resting spores of Bacillus subtilis exposed transiently to the cyclic polypeptide antibiotics, polymyxin B and gramicidin S, and the aminoglycoside antibiotics, streptomycin, kanamycin, and gentamicin. Normal germinated spores showed breaks of the spore coat, a diminution in size and a fibrillar appearance of the cortex, a swelling core, a cell wall as thick as that of vegetative cells, some mesosomes and DNA fibrils. On the other hand, no breaks of the spore coat, a spore core with a slight swelling and irregular form, a thin cell wall, no demonstration of the nuclear material and no granularity in the cytoplasm were characteristic of the germinated spores derived from polymyxin B-and gramicidin S-treated resting spores. With gramicidin S-treated germinated spores a few vacuoles were formed in the cytoplasm. Both polymyxin B-and gramicidin S-treated germinated spores showed little or no synthesis of DNA, RNA, and protein. The vegetative cells derived from streptomycin-treated resting spores demonstrated several finely granular regions in the cytoplasm and a disorder of the fibrillar nucleoid, and their autolysis occurred early. Their DNA and RNA synthesis was normal, whereas protein synthesis was low. In spite of no occurrence of cell division and very low protein synthesis, the most striking characteristics of the outgrowing cells derived from kanamycintreated resting spores were a markedly thickened cell wall and a continuous incorporation of labeled D-alanine suggesting cell wall synthesis; RNA synthesis was slightly lower and DNA synthesis was almost normal. The outgrowing cells from gentamicin-treated resting spores also revealed relatively thick cell walls and a very slight incorporation of labeled n-alanine, Their DNA and RNA synthesis was fairly low and protein synthesis was almost completely inhibited. These results coincide with the growth curves of individual antibiotic-treated resting spores. 655

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Germination initiation and outgrowth of spores ofBacillus brevis strain Nagano and its gramicidin S-negative mutant

Piret

Demain

1982

Arch. Microbiol.

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Bacillus brevis strain Nagano and its gramicidin S-negative mutant, BI-7, were compared with respect to germination of their spores produced in several media. Germination initiation occurred in the presence of nutrient broth or L-alanine but not with inosine, glucose, glycerol or fructose; the process was activated by heat. Parental and mutant spores behaved similarly in these experiments. During outgrowth, parental spores remained in this phase of germination much longer than did mutant spores, but only when the parental spores had been harvested from a sporulation medium where significant gramicidin S synthesis had occurred. When parental spores were extracted or treated with an enzyme that hydrolyzes gramicidin S, rapid outgrowth occurred. Adding exogenous gramicidin S or the extract from parental spores to mutant spores lengthened the outgrowth in a dose-dependent manner. The uptake of labeled L-alanine by parental spores was delayed compared to mutant spores in the presence or absence of chloramphenicol. These data suggest a mechanism of action for gramicidin S whereby it interferes in membrane function, such as transport or energy metabolism, in outgrowing spores.

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Mutual Relationship between Antibiotics and Resting Spores of Bacillus subtilis: Binding of Cyclic Polypeptide and Aminoglycoside Antibiotics to Spores and Their Inhibitory Effect on Outgrowth and Vegetative Growth

Cited by 10 publications

References 19 publications

Early Bacillus anthracis-macrophage interactions: intracellular survival and escape

Early Bacillus anthracis-macrophage interactions: intracellular survival and escape

Mutual Relationship between Antibiotics and Resting Spores of Bacillus subtilis: Morphological Changes and Macromolecular Synthesis after Germination of Spores Treated with Cyclic Polypeptide and Aminoglycoside Antibiotics

Germination initiation and outgrowth of spores ofBacillus brevis strain Nagano and its gramicidin S-negative mutant

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