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PubMed1980

[Differentiation of cholera and NAG vibrios by their virulence].

The method for differentiating cholera and NAG vibrios by their virulence is proposed. This method is based on the evaluation of the following characteristics: hemolytic activity and sensitivity to phages C and Xdf-5, the vibrios being cultivated in 1% peptone water with 1% of glycerol (for the determination of hemolytic activity) and in 1% peptone water with 1% of barium chloride (for the determination of sensitivity to phages).

Bacteriophage Typing↗
PubMed1978

[Experience in the isolation of salmonellae in foods of different risk categories (author's transl)].

146 samples of pancreatic powder, 208 samples of pasteurized egg albumen, both deep-frozen and spray-dried, and 355 samples of frozen poultry were investigated for the presence of salmonellae. Four pre-enrichment media (lactose broth, nutrient broth, buffered peptone water and tryptone soya broth) and two direct selective enrichment media (dulcitol selenite and tetrathionate) were used. For the pre-enrichment of frozen poultry, buffered peptone water and tryptone soya broth proved to be more satisfactory than lactose broth and nutrient broth. Pre-enrichment of freeze-dried material in a non-selective medium yielded more Salmonella than were obtained by a direct selective enrichment. 16 of 146 samples of freeze-dried pancreatic powder contained salmonellae. Accordingly, tissue and organs of slaughter animals designed for the production of pharmaceutical preparations require regular bacteriological control.

Culture Media↗
PubMedDate not supplied

[Identification of Vibrio cholerae O1 by flow cytometry].

A total of 72 peptonated water samples suspected of carrying Vibrio cholerae were assessed by laser flow cytometry (LFC) and compared with positive culture. We used a direct fluorescence technique using polyclonal (PolAb) and monoclonal antibodies (MoAb) conjugated to fluorescein. The PolAb were able to detect 33 positive samples. A clear difference among the 20 positive samples was found with only three V. cholerae O1 false negatives when MoAb were used whereas all 13 V. cholerae Non O1 samples were detected. The correlation index comparing control autofluorescence with peptonated water samples show a R = 0.69, versus 0.96 with pure V. cholerae O1 strains. Our data suggest that the LFC technique is able to recognize V. cholerae O1 from a mixture of microorganisms with high sensitivity and specificity in a few hours.

Antibodies, Bacterial↗
PubMedDate not supplied

[Polymerase chain reaction (PCR) for the identification of toxigenic Vibrio cholerae O1 in oysters].

PCR was made with ctx2 (CGG GCA GAT TCT AGA CCT CCT G) y ctx3 (CGA TGA TCT TGG AGC ATT CCC AC) primers for subunit A of cholera toxin, 30 cycles of temperature on samples of 50 g of oysters added in 450 ml of peptone alcaline water that were inoculated with 15 x 10(6), 0.75 x 10(6) and 0.15 x 10(6) CFU/ml of toxigenic 6707 V. cholerae O1 reference strain. The samples were tested by three microbiological methods: INDRE's method uses 1 x 10(-1) dilution of sample, two fold pass to peptone alcaline water pH 9 incubated 18 h and 6 h at 37 degrees C, the Food and Drugs Administration (FDA) method uses 10(-1) to 10(-6) dilutions of sample, 6 h incubation and reincubation for 18 h at 37 and 42 degrees C and the Mexican laboratories (LMD) with 10(-4) to 10(-3) dilutions, the samples were incubated for 6 h and then reincubated for 18 h at two temperatures 37 and 42 degrees C. The PCR by INDRE's method was positive with 3 x 10(2) CFU/ml/g oyster. In the FDA's method the PCR detected DNA in 10(-4) dilution with 3 x 10(1) CFU/ml/g oyster and in LMD's method the PCR was positive in 10(-3) with 3 CFU/ml/g oyster. The results of the PCR were obtained between 5-6 h, and later V. cholerae O1 was isolated by three microbiological methods. The PCR reproducibility was better on DNA sample diluted 1:4 and 10 microliters of sample increased from 1:1000 to 1:10000 the sensitivity of PCR.

Animals↗
PubMed1995

Effect of cultivation medium on some physicochemical parameters of outer bacterial membrane.

The changes of surface charge and hydrophobicity of the outer bacterial membrane in relation to utilization of n-hexadecane were studied. For this spectrophotometric study adsorption of methylene blue and transport of gentian violet were used. The decrease in the negative charge of the bacterial strains Pseudomonas putida CCM 3423, P. aeruginosa, and P. fluorescens CCM 2115, depended on the type of growth medium. The decrease of surface charge was in the order: meat extract peptone broth > mineral medium with glucose > mineral medium with n-hexadecane. The highest permeability of the bacterial membrane for gentian violet was determined in the case of P. fluorescens grown in meat extract peptone broth. This effect can be explained by a greater hydrophobicity of the bacterial surface for this strain. In other strains a lower permeability was observed. P. fluorescens showed a greater adherence to hexadecane.

Cell Membrane↗
PubMed1995

In vivo regulation of rat neutrophil apoptosis occurring spontaneously or induced with TNF-alpha or cycloheximide.

We previously demonstrated that human TNF-alpha induces rapid apoptosis of human neutrophils. To understand better the in vivo significance of neutrophil apoptosis, we examined spontaneous, recombinant human and mouse TNF-alpha- or cycloheximide-induced apoptosis of normal peripheral blood neutrophils (PBN), PBN from rats injected i.p. with proteose peptone or a streptococcus preparation, OK-432 (inflammatory PBN), peritoneally exudated neutrophils (PEN) obtained after a proteose peptone injection, and normal bone marrow neutrophils. The following observations were made. 1) Normal PBN responded to TNF-alpha, but PEN, normal bone marrow neutrophils, and inflammatory PBN at 12 h after stimulation did not. 2) The sensitivity to TNF-alpha of the inflammatory PBN started to decrease at 3 h, was lowest at 12 h, and was almost restored at 52 h after stimulation. 3) Spontaneous apoptosis of normal and inflammatory PBN reached 25% at 12 h after in vitro incubation, but that of PEN and normal bone marrow neutrophils was very low over this period. 4) The sensitivity to cycloheximide (6 h incubation) was high for normal PBN and bone marrow neutrophils, but low for PEN and inflammatory PBN after 12 h. 5) 125I-rhTNF-alpha binding of bone marrow neutrophils was significantly lower than that of normal and inflammatory PBN and PEN. 6) TNF-alpha-induced apoptosis of normal or inflammatory PBN and bone marrow neutrophils was enhanced by treatment with low doses of cycloheximide that alone were barely able to induce neutrophil apoptosis; however, apoptosis of PEN was not. The mechanisms and in vivo significance of these phenomena are discussed.

Animals↗
PubMedDate not supplied

Aspergillus fumigatus somatic antigens: comparison between two methods of preparation and immunochemical characterization.

Aspergillus fumigatus and other species of the same genus can cause different pathological processes amongst which can be found pulmonary hypersensitivity, intracavitary colonization and invasive and disseminated aspergillosis. The diagnosis of these processes is helped in a great way by the immunological response of the patient to Aspergillus antigens or by the detection of these as a metabolic product in blood and other fluids such as urine. To date, no firm agreement exists regarding the best preparation and standardization technique of aspergillar antigens, which is however the most important step towards the development of immunological diagnostic tests. In this study, two methods for the obtention of A. fumigatus somatic antigens have been compared, using 3 strains of different origins. The main differences between both antigenic extraction methods were the liquid media used for cultures (Czapek and Glucose peptone), the incubation times (4 weeks and 3 days respectively), the type of agitation during the incubation period (intermittent or continuous) and the dialysis and concentration method. The antigens obtained were standardized by the determination of the proteic and hydrocarbonate content, vertical electrophoresis in SDS PAGE, immunoprecipitation and immunoblotting against experimental antisera. From the comparison between both extraction methods, it was found that the highest proteic content was obtained with the somatic antigens of mycelium developed in Czapek for 4 weeks, whereas fungi cultivated in Glucose peptone had a higher glucide content. Vertical electrophoresis revealed the presence of 6 common fractions in all the antigens, whilst some fractions were in majority, only found in one antigen such as the MW 109 kDa, present in the GP 69 antigen.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMedDate not supplied

[Effectiveness of the use of nutrient media in isolation of Yersinia].

Three liquid nutrient media (phosphate buffer medium, 1% buffered peptone water pH 7.6-7.7, buffer casein yeast medium) and live solid media (Endo's, Serov's, and differential diagnostic medium with bromothymol blue, made in this country, and two foreign ones, agar deoxycholate and McConkie's) are compared in examinations of the material from 213 patients with acute intestinal diseases and from 460 small mammals. The results demonstrate a higher efficacy of bacteriologic investigations carried out in the new buffer casein yeast medium or 1% buffered peptone water, pH 7.6-7.8. The data are also indicative of the essential advantages of the differential diagnostic medium with bromothymol blue and prompt its wide practical use.

Acute Disease↗
PubMedDate not supplied

[Description of Streptomyces malvinense nov. sp. isolated from the soil of Islas Malvinas].

In the course of a study on antibiotic activity of microorganisms isolated from soil samples from Port Stanley, Islas Malvinas, Argentina, four thermophilic Actinomycetes were sudied. The four strains had the same morphological and physiological characteristics. The new species belonged to Seccion Rectiflexibilis genus Streptomyces and had the following characteristics: sporophores straight, round spores of 1,5 or 2 millimicron in diameter. The colonies in agar meat peptone were round, flat, mealy, with fimbriate edges, odorless, friable, easy to emulsify and the aerial mycelium was white to light gray. The sporulation was rapid (less of 24 hours at 50 degrees C), the microorganisms grew in media with organic nitrogen and his ability to utilize carbon was scarce, only a trace of growth was detected with manitol (20 days at 50 degrees C). No growth was observed in gliceril-asparagine agar and the utilization of glucose was scarce or absent. It produced coagulation and peptonization of milk and liquifaction of gelatin. Neither was starch hidrolized nor nitrate reduced. The optimun growth temperature was between 45 - 60 degrees C. Very resistant to high temperatures having a thermal death point of more that 2 hours at 100 degrees C. It produced SH2.

Argentina↗
PubMedDate not supplied

[Disc electrophoretic characteristics of the serum proteins of "Vitalact"-type "humanized" milk].

Disc-electrophoreograms in the polyacrylamide gel of the cow milk serumal proteins show 11 peaks belonging to immune globulins, the poteose-peptone fraction, the serum-globulin, alpha-lactoalbumin, beta-lactoglobulin and to glucoproteids. Under a high-temperature treatment of humanized milk (105 degrees----10 minutes) a significant denaturation of the serumal proteins, re-distribution and a fall in the amount of proteinic fractions were noted. Beta-lactoglobulins and immune globulins are most sensitive. With disc-eletrophoresis in the polyacrylamide gel of the human milk serumal proteins the densitograms distinctly demonstrate the presence of peaks that correspond to immunoglobulins, proteose-peptones, serum-albumins, alpha-lacto-albumins in the absence of the beta-lactoglobulins fraction. The cited data allow the method of disc-electrophoresis in the polyacrylamide gel to be employed for improving the fractional composition of the serumal proteins in the nutrients intended for nurslings and infants of the first year of life.

Animals↗
PubMedDate not supplied

[Effect of culture medium components on the accumulation of extracellular bacillary ribonucleases in culture fluid of recombinant strains of Escherichia coli].

The effect of the concentrations of peptone, yeast extract, and inorganic phosphate on the expression of genes of extracellular ribonucleases from Bacillus intermedius 7P (binase) and Bacillus amyloliquefaciens H2 (barnase) was studied in Escherichia coli cells transformed with plasmids containing the structural genes of binase or barnase under the control of their own or synthetic regulatory region or the structural binase gene under the control of the regulatory regions of the genes of barnase or Bacillus pumilus RNase. Inorganic phosphate inhibited the expression of the binase gene under the control of its own regulatory region or the regulatory region of the RNase Bp gene. In all other cases, inorganic phosphate produced no effect on the synthesis of RNases by E. coli cells. This difference in the effects of phosphate may be due to the presence of a nucleotide sequence similar to the E. coli Pho box in the promoters of the binase and RNase Bp genes and the absence of this sequence in the barnase gene promoter. It was shown that high peptone and yeast extract concentrations in the cultivation medium are required for good growth of the recombinant E. coli strains and the biosynthesis of RNases.

Bacillus↗
PubMed1976

Bacteriologic and vaccination studies in a field epizootic of infectious bovine keratoconjunctivitis in calves.

Infectious bovine keratoconjunctivitis (IBK) was enzootic in the beef cattle herds at Dixon Springs Agricultural Center, University of Illinois. The development of nonhemolytic and hemolytic Moraxella bovis flora in the eyes of 48 calves in a closed cow-calf herd was monitored from late May to October, 1972. The incidence of clinical IBK was recorded each week. In late May, nonhemolytic M bovis was isolated from 26% of calf eyes. The peak incidence of IBK was observed in early September, with 26% of the eyes affected. At that time, nonhemolytic M bovis was isolated from 10% of the eyes, and hemolytic M bovis from 58%. By late October, nonhemolytic M bovis was isolated from 25% of the eyes, and hemolytic M bovis from only 4%. In an attempt to increase the resistance of ocular tissue, 19 calves were vaccinated in each third eyelid with 0.5 ml of an autogenous M bovis bacterin in late May. Vaccination did not provide practical protection against the establishment of hemolytic M bovis in the eyes nor the development of clinical IBK. However, at the peak incidence of IBK, hemolytic M bovis was isolated from the eyes of 48% of vaccinated calves and 73% of nonvaccinated calves; clinical IBK was present in 21% of the eyes of vaccinated calves and 29% of nonvaccinated calves. In evaluating the in vitro proteolytic potential of M bovis isolates, 83 hemolytic and 5 nonhemolytic isolates peptonized litmus milk; 176 nonhemolytic isolates did not peptonize litmus milk. Hemolytic M bovis isolates were more pathogenic for mice than were nonhemolytic isolates. Of 60 mice, 53 (88%) died in 1 to 3 hours after intraperitoneal inoculation of hemolytic isolates; 8 of 32 (25%) mice died in 5 to 15 hours after inoculation of nonhemolytic isolates. Hemolytic M bovis isolates produced testicular swelling and scrotal necrosis after inoculation into the scrotal sac of rabbits; nonhemolytic isolates produced only mild transient testicular swelling.

Animals↗
PubMed1976

[Use of Nessler's reagent for recognition of lysine, ornithine, and arginine decomposition by gramnegative fermentative bacteria (author's transl)].

The reactions of lysine, ornithine and arginine decomposition are often difficult to read in Falkow's medium because either the decolorization of the indicator or the lack of sharp colour differences between positive and negative reactions. In such cases Nessler's reagent may be a useful aid. A volume of about 0.2 ml is added to the cultures after 4 days incubation through the mineral oil layer by means of a pipette. A positive reaction is indicated by an immediate white precipitation in case of lysine and ornithine decarboxylation, and a white or brownish precipitate which indicates arginine decomposition. A delayed opacity should be regarded as a negative reaction. Only unequivocal reactions should be considered. The specificity of the reactions was tested with pure substances of compounds which are formed by the decomposition of lysine, ornithine and arginine. Further studies of bacterial cultures in Falkow's medium and in a synthetic, amino acid containing medium without peptone gave identical results and showed that peptone derivates do not cause a false positive reaction with Nessler's reagent (Table 1). Comparative studies on 605 strains of Enterobacteriaceae and Vibrio in Falkow's medium with and without added Nessler's reagent gave corresponding results except some strains of Escherichia coli and Citrobacter freundii with different arginine reactions (Table 2). Strains of these species mostly decolorized the indicator thereby hindering the recognition of either a true positive or a true negative reaction. In these cases, however, the results obtained after addition of Nessler's reagent corresponded closely to the percentage of positive reactions cited in the literature.

Arginine↗
PubMed1965

ALPHA SARCIN, A NEW ANTITUMOR AGENT. II. FERMENTATION AND ANTITUMOR SPECTRUM.

Aspergillus giganteus MDH 18894 was found to produce alpha sarcin, a new antitumor agent inhibitory to a number of different animal tumors. This culture produced culture filtrates that inhibited mouse sarcoma 180 at dilutions up to 1:32. Optimal fermentation conditions were established for shake flasks and 30-liter and 100-gal (378.5-liter) fermentors. Results from a variety of media were reported; however, the medium which yielded the most reproducible filtrates consisted of 2% corn starch, 1.5% beef extract (Difco), 2% peptone (Difco), and 0.5% sodium chloride. Purified preparations of 800 sarcoma 180 dilution units per mg were inactive against bacteria, yeasts, molds, actinomycetes, and protozoa, but were cytotoxic to several strains of mammalian cells. Alpha sarcin inhibited sarcoma 180 in mice at a dose of 62.5 mug per kg per day. A variety of other animal tumors were also inhibited by this compound.

Animals↗
PubMed1964

PRODUCTION OF HYDROGEN SULFIDE BY STREPTOMYCETES AND METHODS FOR ITS DETECTION.

The ability of streptomycetes to produce hydrogen sulfide is generally used for taxonomic purposes. It was found that the previously used method, the blackening of Peptone Iron Agar, does not clearly indicate formation of hydrogen sulfide. It was shown that the blackening of a lead acetate strip is the most accurate indicator for H(2)S-producing streptomycetes. A great variety of organic and inorganic sulfur compounds were examined and compared, and the choice of the most suitable sulfur source and method for the detection of hydrogen sulfide is discussed.

Agar↗
PubMed1977

Lymphokine-induced secretion of plasminogen activator by murine macrophages.

Thioglycollate-stimulated macrophages are known to release a plasminogen activator (PA) into the medium. In this study it was investigated whether macrophages could be activated to release PA after exposure to lymphokines. Macrophage monolayers obtained by 24 h culture of proteose peptone-elicited murine exudate cells were incubated with lymphocyte culture supernatants. After 48 h the supernatants were replaced by serum-free medium and the macrophages were incubated for another 24-48 h. These supernatants were assayed for PA as measured by the lysis of 125I-labeled fibrin. The following results were obtained: (a) Supernatants of antigen or mitogen-stimulated spleen cells induced PA secretion by macrophages whereas control supernatants were ineffective. The same was found with supernatants of mitogen-stimulated lymph node cells. (b) PA secretion by macrophages seems to be induced by a rather narrow concentration range of lymphokines. (c) Lymphokine-induced PA secretion by macrophages is enhanced after phagocytosis of latex beads. The results show that PA secretion by activated macrophages can be considered as a parameter of immunoactivation.

Animals↗
PubMed1979

Macrophage-dependent, NK-cell-independent "natural" surveillance of tumors in syngeneic mice.

The present study, which was designed to further characterize the "natural" T-independent rejection of syngenetic tumours (Greenberg and Greene, 1976), has revealed the following points: (1) no detectable DBA/2 NK cell activity was demonstrated against the syngeneic tumour lines studied, and these tumours were indensitive to NK cells from high-activity strains; (2) in addition the tumour frequencies in old and young mice receiving small tumour inocula were identical, in contrast with the reported decline in NK cell activity with age, suggesting that the surveillance of small inocula of these tumours was NK-cell-independent; (3) injection of silica intraperitoneally enhanced the frequency of tumours in normal and immunodeficient AT x BM mice, suggesting that the rejection mechanism was macrophage-dependent; (4) the effects of silica injection were maximal if administered 3 days prior to tumour injection, indicating that the period of time in which the rejection mechanism must act was very limited; (5) silica markedly decreased the survival of AKR mice dying of spontaneous tumours, providing evidence that the effect of this agent was not limited to model systems but would influence the appearance of spontaneous tumours; (6) reticuloendothelial stimulants such as mycobacterium butyricum and proteose peptone decreased the tumour frequency of small tumour inocula, indicating that the effector mechanism can be stimulated; and (7) soluble tumour antigen enhanced the tumour frequency in normal and immunodeficient mice, suggesting that the specific receptor molecule of the surveillance mechanism was not thymus-dependent.

Animals↗
PubMed1984

Differential expression of macrophage effector functions: bactericidal versus tumoricidal activities.

Macrophage populations may be induced to express tumoricidal or bactericidal activities following exposure to certain stimuli. An understanding of the differences in the stimulatory mechanisms and in the characteristics of the macrophages they affect will be facilitated by comparing functional activities of various macrophage populations. The experiments described here were conducted to determine whether injection of a single stimulus necessarily drives cells to express both tumoricidal and bactericidal activities or whether selected reagents can drive cells to express one activity without expressing the other. The data show that a single population of mouse or hamster peritoneal exudate cells obtained following injection of proteose peptone is bactericidal for Listeria monocytogenes and for E. coli, but is not tumoricidal for TCMK-1, Ad2HE3, or mKS-A TU-5 target cells. In contrast, peritoneal exudate cells collected after injection of Bacillus Calmette Guerin (BCG) organisms are always highly tumoricidal, and either show no effect on Listeria monocytogenes or E. coli, or are at best bacteriostatic. Data indicate that the effector cells in these assays are macrophages, that the dissociation of tumoricidal and bactericidal activity occurs over a wide dose range, and that the tumoricidal capabilities are not artifacts of the assay system. These results suggest that a given macrophage population may preferentially express tumoricidal or bactericidal activities depending on the stimulus used.

Animals↗