STUDIES OF ASPOROGENIC MUTANTS OF BACILLUS CEREUS: PRELIMINARY INVESTIGATIONS WITH CALCIUM AND ZINC

1962 ◽  
Vol 8 (6) ◽  
pp. 823-833 ◽  
Author(s):  
J. J. Cooney ◽  
D. G. Lundgren

The physiology of spore formation was studied in Bacillus cereus and a temperature-sensitive asporogenic mutant. The parent organism sporulates when cultured in a minimal medium at either 28 °C or 37 °C while the mutant sporulates only at 28 °C. The blocking of sporulation at 37 °C has been referred to as "abortive" sporulation. Uptake of calcium and zinc was followed during growth and sporulation or "abortive" sporulation. Calcium and dipicolinic acid (DPA) levels in sporogenic cultures increased as the medium calcium was increased. The asporogenic mutant took up less calcium and synthesized little DPA. Heat resistance of spores increased as the calcium and DPA increased. Over 99% of Ca45or Zn65were released from labelled spores when autoclaved to release DPA. Chemical fractionations were made of cells labelled with Zn65and Ca45and harvested at different times during the culture cycle. Smaller percentages of calcium than of zinc were located in the cold trichloroacetic acid soluble fraction. The alcohol-soluble, ether-insoluble fraction of spores contained a greater percentage of calcium than was found in vegetative cells. Cells which had undergone "abortive" sporulation contained the same percentage of calcium in this fraction as homologous vegetative cells.

2021 ◽  
Vol 9 (3) ◽  
pp. 667
Author(s):  
Zhiwei Tu ◽  
Peter Setlow ◽  
Stanley Brul ◽  
Gertjan Kramer

Bacterial endospores (spores) are among the most resistant living forms on earth. Spores of Bacillus subtilis A163 show extremely high resistance to wet heat compared to spores of laboratory strains. In this study, we found that spores of B. subtilis A163 were indeed very wet heat resistant and released dipicolinic acid (DPA) very slowly during heat treatment. We also determined the proteome of vegetative cells and spores of B. subtilis A163 and the differences in these proteomes from those of the laboratory strain PY79, spores of which are much less heat resistant. This proteomic characterization identified 2011 proteins in spores and 1901 proteins in vegetative cells of B. subtilis A163. Surprisingly, spore morphogenic protein SpoVM had no homologs in B. subtilis A163. Comparing protein expression between these two strains uncovered 108 proteins that were differentially present in spores and 93 proteins differentially present in cells. In addition, five of the seven proteins on an operon in strain A163, which is thought to be primarily responsible for this strain’s spores high heat resistance, were also identified. These findings reveal proteomic differences of the two strains exhibiting different resistance to heat and form a basis for further mechanistic analysis of the high heat resistance of B. subtilis A163 spores.


1992 ◽  
Vol 38 (1) ◽  
pp. 38-44 ◽  
Author(s):  
Sonia Senesi ◽  
Giulia Freer ◽  
Giovanna Batoni ◽  
Simona Barnini ◽  
Anna Capaccioli ◽  
...  

Spores of the strain NCIB 8122 of Bacillus cereus have been depleted of coats by treatment with 0.1% sodium dodecyl sulfate – 200 mM 2-mercaptoethanol – 0.5 M NaCl (pH 9.6). The coat-depleted spores did not show any decrease in viability, heat resistance, refractility, dipicolinic acid content, or specific activities of several protoplastic enzymes. The germinative response of the coat-depleted spores to adenosine and several analogues thereof was found qualitatively similar to that obtained with intact spores. However, germination kinetics appeared to be affected by coat removal, since germination rate measured as loss of refractility was eight times slower even at inducer concentrations 10-fold higher than those required to promote optimal germination response of intact spores. Loss of heat resistance, on the other hand, was hardly affected by coat removal. These results suggest that, even though spore coats are not essential for the triggering reaction, they are required for a rapid evolution of the later events in the germination process. Key words: adenosine analogues, germination-triggering reaction, spore coats, coat-depleted spores, Bacillus cereus.


1992 ◽  
Vol 46 (2) ◽  
pp. 357-364 ◽  
Author(s):  
E. Ghiamati ◽  
R. Manoharan ◽  
W. H. Nelson ◽  
J. F. Sperry

UV resonance Raman spectra of Bacillus cereus, Bacillus megaterium, and Bacillus subtilis endospores have been excited at 222.7,230.7,242.5, and 251.1 nm, and spectra have been compared with those of vegetative cells. The resonance Raman spectra of aqueous solutions of dipicolinic acid and calcium dipicolinate have been measured at the same wavelengths. Spectra of endospores and their corresponding germinated spores show only modest differences when excited at 222, 231, and 251 nm. However, very substantial differences appear when excitation occurs at 242 nm. Difference spectra obtained at 242 nm by subtracting spectra of germinated spores of Bacillus cereus from spectra of their corresponding endospores are attributed almost entirely to dipicolinate. Vegetative cells and endospores show large spectral dissimilarities at all exciting wavelengths. These spectral differences, which vary strongly with exciting wavelength, appear to be the result of large differences in the amounts and composition of proteins and nucleic acids, especially ribosomal RNA. The very substantial resonance enhancement of Raman spectra has been obtained from aqueous solutions of pure dipicolinic acid and of sodium and calcium dipicolinate salts, as well as spores at the various exciting wavelengths. The strong enhancement of dipicolinate spectra in spores, however, was noted only with 242-nm excitation. Consequently, only with 242-nm light was it possible to selectively and sensitively excite and study calcium dipicolinate in spores. Resonance enhancement of the dipicolinate spectra with 242-nm excitation appears due primarily to resonance interactions with n-π* electronic transitions associated with the pyridine ring and/or the carboxylate group.


1970 ◽  
Vol 118 (1) ◽  
pp. 81-87 ◽  
Author(s):  
R. J. White ◽  
P. W. Kent

1. After incubation of Escherichia coli with N-iodo[1,2-14C2]acetylglucosamine, 95–99% of the 14C taken up by whole cells is located in a cold-trichloroacetic acid-soluble fraction. Two major components of this fraction are S-carboxymethylcysteine and S-carboxymethylglutathione. The same compounds accumulate during incubation with iodo[14C]acetate but not with iodo[14C]acetamide. The amount of 14C associated with a cold-trichloroacetic acid-insoluble fraction are comparable for all three alkylating agents. After incubation with iodo[14C]acetamide, 50% of the label bound to whole cells is recoverable in a cold-trichloroacetic acid-insoluble fraction. 2. Uptake and incorporation of 14C from [U-14C]glycerol is blocked at an early stage by N-iodoacetylglucosamine. No specific inhibition of macromolecular synthesis could be demonstrated. 3. Mutants selected for resistance to iodoacetate are partially resistant to iodoacetate and N-iodoacetylglucosamine, but show no resistance to iodoacetamide. 4. Mutants selected for resistance to N-iodoacetylglucosamine are not resistant to iodoacetate or iodoacetamide, and are defective in their ability to grow on N-acetylglucosamine. Resistance to N-iodoacetylglucosamine is not absolute, and depends on the presence of glucose or certain other sugars; there is no resistance during growth on maltose, glycerol or succinate. 5. Absolute resistance can be achieved by selecting for a second mutation conferring resistance during growth on maltose; double mutants isolated by this procedure are unable to grow on N-acetylglucosamine and grow poorly on glucosamine. Resistant single mutants have a slightly diminished uptake of N-acetyl[1-14C]glucosamine, but in resistant double mutants the uptake of both [1-14C]glucosamine and N-acetyl[1-14C]glucosamine is severely diminished. 6. These observations are consistent with the presence of two permeases for N-acetylglucosamine, one that also permits uptake of glucosamine and another that allows entry of methyl 2-acetamido-2-deoxy-α-d-glucoside. N-Iodoacetylglucosamine can gain entry to the cell by both permeases.


1991 ◽  
Vol 260 (3) ◽  
pp. R525-R532
Author(s):  
J. L. Beverly ◽  
B. J. Hrupka ◽  
D. W. Gietzen ◽  
Q. R. Rogers

Diffusion or metabolism of the dietary limiting amino acid (DLAA) in the prepyriform cortex (PPC) may account for the time lag between injection of the DLAA into the PPC and the increase in intake of an amino acid-imbalanced diet. Results from the injection of [3H]Leu +/- [14C]Thr (DLAA) into the PPC indicated rapid (less than 15 min) and limited diffusion (85-90% of recovered label was less than or equal to 1 mm from the injection site). 3H and 14C decreased in the trichloroacetic acid (TCA)-soluble fraction and increased in the TCA-insoluble fraction during the first 1.5 h and remained constant in the TCA-insoluble fraction 1.5-6 h after injection. An increase (approximately 50%) in 3H in the TCA-insoluble fraction was found less than or equal to 30 min after injection of the DLAA. There was no affect of the DLAA on 3H in the TCA-soluble fraction. These results indicated that a change in metabolism within the PPC may be responsible for the delay in onset of the feeding response after injection of the DLAA into the PPC.


2017 ◽  
Vol 64 ◽  
pp. 195-201 ◽  
Author(s):  
Alizée Guérin ◽  
Claire Dargaignaratz ◽  
Thierry Clavel ◽  
Véronique Broussolle ◽  
Christophe Nguyen-the

1960 ◽  
Vol 6 (2) ◽  
pp. 213-224 ◽  
Author(s):  
Samuel H. Black ◽  
Tadayo Hashimoto ◽  
Philipp Gerhardt

Spores of Bacillus cereus strain terminalis formed "endotrophically" by transferring granular vegetative cells to distilled water were found to be relatively susceptible to heat and deficient in dipicolinic acid. Calcium ions alone, added in low concentration shortly after the cells were placed in water, could completely relieve these abnormalities. Although the water-formed spores were sensitive to heat, they were as fully resistant as normal spores to gamma radiation or phenol.


1963 ◽  
Vol 9 (2) ◽  
pp. 251-258 ◽  
Author(s):  
Elisabeth Ann Pelcher ◽  
H. P. Fleming ◽  
Z. John Ordal

A replacement technique, whereby spores of Bacillus cereus were produced by transferring vegetative cells into non-nutritive solutions, was employed for the investigation of the relationship between calcium, dipicolinic acid (DPA), and heat resistance in spores. Replacement solutions of distilled water, fresh medium, calcium, strontium, barium, nickel, and other metals were employed. Spores produced in distilled water were heat sensitive and contained low levels of calcium and DPA. Heat-resistant spores with a normal complement of calcium and DPA were formed only in fresh medium or in solutions containing suitable levels of calcium. Calcium and DPA were present in the spores in nearly equimolar quantities. Of the other metals tested, only strontium stimulated DPA synthesis above the level of the spores produced in water. Spores of intermediary heat resistance but low DPA content were formed in solutions of barium and nickel.


2015 ◽  
Vol 35 (03) ◽  
pp. 300
Author(s):  
Reny Mailia ◽  
Bara Yudhistira ◽  
Yudi Pranoto ◽  
Saiful Rochdyanto ◽  
Endang Sutriswati Rahayu

Characteristics of tofu with higher a (0.89 to 0.90) and protein levels of 8% or more, made tofu to be a suitable medium for bacterial growth. This leads to out to be very easy to damage due to bacterial contamination. Contamination of bacteria is commonly found in the tofu because of contamination in the process making of tofu. Source of contamination can come out from the raw material, during the process of making tofu and hygienic sanitation level during processing. Generally, this study aimed to determine the level of contamination of Escherichia coli, Staphylococcus aureus, Bacillus cereus and spore-forming bacteria in the process of making tofu and study the properties of heat resistance of eachisolate. Phases of of the study started with the isolation and identification and then quantitative analysis of Escherichiawcoli, Staphylococcus aureus, Bacillus cereus and spore-forming bacteria in the tofu process from raw materials to end product, tofu, comprised from water and soybean, slurry, soymilk cooking, curd, whey and tofu. Isolates originating from the cooking process and the coagulation process was for testing the heat resistance (D value and Z value). D and Z values were calculated using linear regression. Escherichia coli found in the water, soybeans, soybean slurry, curd and tofu, the number 10 =4,83 min and the value of Z = 22.73°C. Staphylococcus aureus found in soybeans and curd, showed the number of 101-102 CFU/g. Escherichia coli GMP isolate had D60°C CFU/g. The Staphylococcus aureus GMP4 isolate, had D60°C 1=2.72 min and the value of Z = 18.87°C. The Staphylococcus aureus GMP 6 isolate, had D=2.54min and the value of Z = 18.18°C. Bacillus cereus found in the water, soybean, soybean slurry, soymilk cooking, curdand tofu, showed the number 102-103CFU/g. Bacillus cereus vegetative cells SK 2 had D=5.43 min and the value of Z = 22.72°C. Bacillus cereus vegetative cells SK 4 had D60°C 60°C =5.95 min and the value of Z = 22.22°C. Spore-forming bacteria found in water, soybean, soybean slurry from the grinding process, the process cooking of soymilk, the process of clotting, whey and tofu, showed the number of 102CFU/g.Keywords: Tofu, Escherichia coli, Staphylococcue aureus, Bacillus cereus, a spore forming bacteria, heat resistance 60°C ABSTRAKKarakteristik tahu dengan a0,89-0,90 dan kadar protein 8% atau lebih, menjadikan tahu sebagai media yang cocok bagipertumbuhan bakteri. Hal ini menyebabkan tahu menjadi sangat mudah rusak karena cemaran bakteri. Penelitian ini bertujuan untuk mengetahui tingkat cemaran Escherichia coli, Staphylococcus aureus, Bacillus cereus dan Bakteri pembentuk spora pada proses pembuatan tahu dan mempelajari sifat ketahanan panas dari masing-masing cemaran. Tahapan penelitian dimulai dari pengamatan proses pembuatan tahu, isolasi dan identifikasi dan analisa kuantitatif cemaran Escherichia coli, Staphylococcus aureus, Bacillus cereus dan bakteri pembentuk spora pada proses pembuatantahu. Isolat yang berasal dari proses pemasakan dan proses penggumpalan digunakan untuk pengujian ketahanan panasdengan melihat nilai D dan Z menggunakan regresi linier. Escherichia coli ditemukan pada air, kedelai, bubur kedelai, gumpalan tahu dan tahu, dengan jumlah 10w1-10CFU/g. Isolat Escherichia coli dari proses penggumpalan (GMP), nilaiD60°C 2=4,83 menit dan nilai Z=22,73°C. Staphylococcus aureus ditemukan pada kedelai, gumpalan tahu dan tahu, dengan jumlah 10=2,72 menit dan nilai Z =18,87°C. Untuk isolat Staphylococcus aureus GMP 6, nilai D1CFU/g.  Isolat Staphylococcus aureus GMP 4, memiliki nilai D60°C60°C =2,54 menit dan nilai Z =18,18°C. Bacillus cereus ditemukan pada air,kedelai, bubur kedelai, sari kedelai masak, gumpalan tahu dan tahu, dengan jumlah 102-10CFU/g. Sel vegetatif Bacilluscereus yang berasal dari sari kedelai (SK) 2, memiliki nilai D60°C3=5,43 menit dan nilai Z =22,72°C. Untuk sel vegetatif Bacillus cereus SK 4, memiliki nilai D60°C=5,95 menit dan nilai Z =22,22°C. Bakteri pembentuk spora ditemukan pada air, kedelai, bubur kedelai pada proses penggilingan, sari kedelai masak, gumpalan tahu, kecutan dan tahu, dengan jumlah 10CFU/g.Kata kunci: Tahu, Escherichia coli, Staphylococcue aureus, Bacillus cereus, bakteri pembentuk spora, ketahananpanas2    


1969 ◽  
Vol 15 (11) ◽  
pp. 1313-1317 ◽  
Author(s):  
D. H. Ashton ◽  
L. C. Blankenship

Two soluble reduced nicotinamide adenine dinucleotide (NADH2) oxidases purified from extracts of Bacillus cereus T spores were compared with vegetative ceil soluble NADH2 oxidase. The minor spore component and vegetative cell soluble NADH2 oxidase reacted equally well with riboflavin or flavin mononucleotide (FMN), were inhibited by 15 mM dipicolinic acid (DPA), and possessed similar thermal inactivation characteristics at 80 °C. Activity of the major spore component was stimulated by a factor of 3.6 when riboflavin replaced FMN as the coenzyme. The major spore component was not inhibited by DPA and resisted heat treatments which inactivated vegetative cell soluble NADH2 oxidase. These observations indicate that the minor spore component and vegetative cell soluble NADH2 oxidase are identical while the major spore component is a distinct protein.


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