Biomedical subjects
Publications and source records attributed to R Beaudet.
In a 38-year-old man with recurrent chest pain 14 months following percutaneous transluminal coronary angioplasty (PTCA) of the left anterior descending coronary artery, two aneurysms were noted at previous PTCA sites without evidence of restenosis. Although the precise mechanism of formation of these aneurysms is not known, it is possible that medial dissection and weakening of the artery provoked aneurysm formation.
Outer membrane proteins (OMP) from Neisseria meningitidis cells of groups A, B, C and Y were extracted with CaCl2. Sodium dodecyl sulphate polyacrylamide gel electrophoresis of each extract showed a multibanded pattern characterized by the presence of three to four major proteins. Immunization of mice with individual extracts induced homologous bactericidal reactions. In addition, the extracts from groups B and C induced heterologous bactericidal reactions (B-C, C-B). The sera of mice immunized with a mixture of group A, B, and Y extracts showed a high bactericidal titre against the three N. meningitidis groups forming the mixture. Moreover, this bactericidal activity had a large spectrum against different strains from groups A, B, Y and also C. After immunization of mice with the individual extracts, each of the sera was shown by immunoblotting to contain antibodies reacting with some of the corresponding proteins. Mice immunized with the mixture showed a significant reduction of bacteraemia following challenge with either of N. meningitidis groups A, B or C. These results suggest that a mixture of OMP from different N. meningitidis groups may have the potential for a meningococcal polyvalent vaccine.
A bacteriophage for Corynebacterium glutamicum strain LP-6 was isolated from swine waste. It belongs to the Siphoviridae family or Bradley morphologic group B, has a narrow host range, and is sensitive to chloroform and resistant to carbon tetrachloride. The phage is unstable (96% inactivation) in swine waste stored for 4 months at 22 C. The DNA has a molecular weight of approximately 20 Md, cohesive ends, and numerous restriction endonuclease sites. The phage differs from other known C glutamicum phages.
A gonococcal inhibitor produced by Staphylococcus haemolyticus was separated into three components by reverse-phase h.p.l.c. The amino acid composition analysis of each of the three components indicated extensive similarities. N-Terminal sequence analysis of all three components allowed the identification of the first 27-30 residues of each. The complete primary structure of each component was determined from the sequence analysis of trypic peptides and peptides generated by mild acid hydrolysis. Each component is composed of 44 amino acid residues, with evidence suggesting the presence of an N-terminal formylmethionine residue in each. The components I, II and III have respectively 33, 29 and 33 identical amino acid residues in their sequences, which represents 75%, 65.9% and 75% homology. These components contain a high proportion of hydrophobic amino acids, and their hydrophobicity profiles are closely related. Also, each of the three components contains a positively charged residue (lysine) as the third residue, followed by a core of hydrophobic residues. These results suggest that the three components are possible signal sequences of one or more secreted or membrane-associated proteins.
The inhibitory substance produced by Staphylococcus haemolyticus strain no. 7 acts on growing as well as resting gonococcal cells, as shown by reductions in viable counts. The optical density of these cell suspensions was only slightly reduced. The inhibitor caused lysis of gonococcal spheroplasts at 24 degrees C and 37 degrees C, but was much less active at 4 degrees C. Acting on intact gonococcal cells, the inhibitor caused a temperature-dependent release of radioactive cytoplasmic material. Electronmicroscopy showed that treated suspensions contained ghost cells with the cell envelope relatively intact. Our results suggest that the inhibitor may act on the cytoplasmic membrane of the gonococcal cell causing cytoplasmic leakage and, eventually, death.
Phenol, p-cresol, and volatile fatty acids (VFA; acetic, propionic, isobutyric, butyric, isovaleric, and valeric acids) were used as odor indicators of swine waste. Aeration of the waste allowed the indigenous microorganisms to grow and degrade these malodorous substances. The time required for degradation of these substances varied according to the waste used, and it was not necessarily related to their concentrations. Using a minimal medium which contained one of the malodorous compounds as sole carbon source, we have selected from swine waste microorganisms that can grow in the medium. The majority of these microorganisms were able to degrade the same substrate when inoculated in sterilized swine waste but with an efficiency varying from one strain to the other. None of these strains was able to degrade all malodorous substances studied. Within 6 days of incubation these selected strains degraded the following: Acinetobacter calcoaceticus, phenol and all VFA; Alcaligenes faecalis, p-cresol and all VFA; Corynebacterium glutamicum and Micrococcus sp., phenol, p-cresol, and acetic and propionic acids; Arthrobacter flavescens, all VFA. On a laboratory scale, the massive inoculation of swine waste with C. glutamicum or Micrococcus sp. accelerated degradation of the malodorous substances. However, this effect was not observed with all of the various swine wastes tested. These results suggest that an efficient deodorization process of various swine wastes could be developed at the farm level based on the aerobic indigenous microflora of each waste.
The conventional catalase test only gives qualitative or semi-quantitative information of the amount of catalase activity produced. Using such a method, we have selected two strong catalase activity producers (Neisseria gonorrhoeae G-10, Staphylococcus epidermidis 66) and a weak producer (Neisseria meningitidis 34702). Quantitative determination of the catalase activity produced by these strains was done by the disk-flotation method using the Catalasemeter. The production of catalase activity in liquid culture by N. gonorrhoeae, S. epidermidis and N. meningitidis occurred mainly during the logarithmic growth phase. The maximum concentration of catalase activity found in these cultures was, for both strong producers, 160 units (U)/ml while it was around 20 U/ml for the weak producer. Some of the other bacteria tested showed high concentrations of catalase activity (130-162 U/ml) and these were: N. gonorrhoeae strain PR 77112 (penicillin resistant), Staphylococcus strain 7 (coagulase negative), Micrococcus sp. strains 2 and 42.
Coagulase-negative staphylococci are known to produce bactericidal and bacteriostatic antigonococcal activities. Out of 12 staphylococci, Staphylococcus haemolyticus (four isolates) was identified as the principal source of bactericidal activity, whereas Staphylococcus epidermidis (six isolates) was primarily responsible for producing bacteriostatic activity. A comparison of the bacteriostatic substance produced by S. epidermidis isolate 66 with the bactericidal substance produced by S. haemolyticus isolate 7, which had been previously purified, showed that they were similar lipoproteins or lipid-associated proteins. However, these two inhibitors migrated differently under electrophoresis on agarose gel. The protein component of the bacteriostatic inhibitor was more difficult to separate from the lipid component when chromatographed on Ultrogel AcA 54 in the presence of urea (4 M) than the protein component of the bactericidal inhibitor. The protein component of both types of inhibitors was responsible for the antigonococcal activity and was dissociable into subunits of approximately 1,400 daltons. However, these protein components had different migration patterns on agarose gel. The bacteriostatic substance displayed a bactericidal effect when dissociated from its lipidic component suggesting that the lipids might play a role in the type of inhibitory activity produced. All of the bactericidal and bacteriostatic substances contained in the different crude preparations were antigenically related.
A strain of Streptococcus faecalis isolated from the urogenital flora was selected for its ability to inhibit the in vitro growth of Neisseria gonorrhoeae. Initially, the inhibitory activity was demonstrated on solid medium only when the inhibitor and the target strains were growing simultaneously, such as in the spot-lawn and flip-flop agar overlay methods. The antigonococcal effect was not due to a shift in the pH or depletion of nutrient in the medium. This activity was not produced in liquid medium nor could it be extracted in a soluble form from either the solid medium or the streptococcal cells. The production of the inhibitory activity could not be enhanced by ultraviolet irradiation or treatment with mitomycin C. The composition of the medium was found to affect the size of the inhibitory zone produced. The inhibitory activity showed a wide antigonococcal spectrum and was susceptible to trypsin and pronase but resistant to alpha- and beta-amylases and catalase. This activity passed through a filtering membrane and was also dialyzable and had an apparent molecular weight of less than 1,000. The addition of bovine serum albumin to solid medium enabled us to show an inhibition even when the producer and the target strains were grown sequentially, thus suggesting that part of the difficulty of studying such inhibitory activity could be due to its instability.
Thrombus obstruction of a prosthetic heart valve is usually treated surgically. We report a well-documented case of an obstructed mitral prosthetic valve where fibrinolytic treatment was successful. Furthermore the thrombus formation probably had occurred 6 months earlier. Thus fibrinolysis appears to be a safe alternative to surgery although late occurrence of thrombosis may be possible.
The purified antigonococcal substance produced by Staphylococcus haemolyticus no. 7 has shown a broad antigonococcal spectrum and a narrow antibacterial spectrum. The inhibitor produced in vitro was also active in the guinea pig subcutaneous chamber. The inhibitor has shown hemolytic activity; the human, horse, and mouse erythrocytes were the most susceptible. Hemolytic and antigonococcal activities were inhibited in the presence of phosphatidylcholine. The amino acid composition of the antigonococcal substance was characterized by the absence of proline, tyrosine, histidine, cysteine, and tryptophan. The molecular weight was found to be 2,565, and the major isoelectric points were 4.8 and 4.9 in the presence of 8 M urea and 4.6 without urea. The inhibitor has some properties similar to those of the delta toxin of Staphylococcus aureus, although the two substances are different based mainly on their chemical characteristics. Also an antiserum directed against the gonococcal inhibitor did not give a precipitation line with the delta toxin, indicating that the two substances are antigenically unrelated.
Among anaerobic bacteria normally found in the urogenital flora, Eubacterium limosum was found to inhibit the in vitro growth of Neisseria gonorrhoeae. The antigonococcal activity produced by E. limosum was soluble in methanol and in a chloroform--methanol mixture (30:70). The fraction soluble in chloroform--methanol (30:70) yielded eight absorbance peaks when chromatographed on Bio-Gel P-2 and the inhibitory activity was found in the first two peaks. This activity was not absorbed on DEAE Sephacel and was eluted with distilled water in a peak considered as peak 1, on which preliminary characterization was done. The inhibitory activity of peak 1 was found to be heat and pH resistant and not susceptible to proteases, lipase, or amylases. When peak 1 was chromatographed on cellulose paper using a butanol--acetic acid (4:1) solvent system, eight different spots were detected upon spraying the paper with ninhydrin. No spot was detected with anthrone, bromothymol, nor Sudan black reagents used for the detection of carbohydrates and lipids. Based on sodium dodecyl sulfate-urea polyacrylamide gel electrophoresis and gel chromatography on Sephadex G-25, peak 1 appeared either as a diffuse band and as a single peak, respectively. The molecular weight of the inhibitory complex was estimated to be 2400. All these results suggest that the antigonococcal activity produced by E. limosum is composed of more than one low molecular weight amino compound.
Seven streptococci isolated from the normal urogenital flora were selected for the production of an inhibitory activity toward Neisseria gonorrhoeae on solid medium. This activity was abolished when catalase was added to the medium in the case of Streptococcus mitis isolates 22, 25, 74 and Streptococcus sanguis II isolate 70 suggesting the involvement of hydrogen peroxide. The inhibitory activities produced by the other isolates (Streptococcus faecalis) were trypsin sensitive (isolates 12 and 14) or stable (isolate 59). Hydrogen peroxide production by S. sanguis II was quantitatively evaluated by reference to a standard curve representing the inhibitory effect of various concentrations of commercial H2O2 on N. gonorrhoeae growth. After 22 h of incubation a yield of 5.3 m mol H2O2 I-1 was reached on solid medium while in liquid medium values of 2.8 and 7.3 m mol H2O2 I-1 were obtained, respectively, in non-agitated and agitated cultures after 8 h of incubation.
The induction of antituberculous immunity by crude ribosomal fractions isolated from Mycobacterium tuberculosis strain H37Ra, M. bovis strain BCG, and M. smegmatis was studied in CF-1 mice. Levels of antituberculous immunity similar to that induced by live BCG were induced by the BCG and H37Ra ribosomal fractions whereas that isolated from M. smegmatis was found to be inactive. Electrophoresis of the three ribosomal fractions in sodium dodecyl sulfate - polyacylamide gels followed by differential staining showed the two active ribosomal fractions to be similar in their proteins, carbohydrate-containing substances, and lipid profiles. The inactive smegmatis ribosomal fraction differed mainly from the active ones on the basis of its carbohydrate-containing substances profile and by the absence of lipids. The polysaccharides and the ribosomes present in the H37Ra ribosomal fractions were purified by affinity chromatography on concanavalin A - Sepharose 4B. Each purified preparation showed no or only low antituberculous activity when injected separately, but when mixed together a high protection was observed. The formation of complexes between the ribosomes and the polysaccharide fraction was suggested and appears to be necessary for the induction of antituberculous immunity.
The dynamics of in vivo multiplication of 10 BCG substrains were studied in three different strains of mice: CF-1, B6D2F1/J and C57BL/6. For each substrain, the number of CFU recovered from the spleen of mice previously injected intravenously with BCG was determined weekly for 5 consecutive weeks. On the basis of the statistical analysis of the results, the BCG substrains were classified for each strain of mice in three groups of virulence: high, intermediate and low. The Russian and Japanese substrains were found to be the most virulent, and the Glaxo and Tice 946 BL (I2) the least virulent substrains. No correlation could be established between in vivo multiplication and antitumour activity of each BCG substrain as estimated in preventive treatment against three tumour systems: Ehrlich carcinoma in CF-1 mice, L1210 leukaemia in B6D2F1/J mice, and B16 melanoma in C57BL/6 mice.
Myocardial infarction is usually caused by a thrombus occurring on a significant coronary lesion. A 60-year-old male was admitted with an acute evolving anterior myocardial infarction. Three hours after the beginning of chest pain, the electrocardiogram showed ST-segment elevation in the anterior and lateral leads which persisted despite intravenous nitroglycerin (100 mcg/min). One hour later, an angiogram showed complete obstruction at the origin of the left anterior descending artery (LAD). After intracoronary streptokinase (250,000 units) the LAD opened and a 90% obstruction was seen at its origin. However, an anterior myocardial infarction occurred. One month later, an angiogram showed a slight irregularity at the origin of the LAD. Thus, this case demonstrates that 1) a myocardial infarction may occur with a near normal coronary artery, and 2) a thrombus may occur at the site of a slight coronary irregularity.
Coagulase-negative staphylococcus no. 7, isolated from the urogenital flora, was grown on semisolid brain heart infusion medium. Supernatants were obtained by centrifuging frozen and thawed media which had supported the growth of the staphylococci at 37 degrees C. The kinetic of production revealed that the antigonococcal activity was detected at the end of the logarithmic phase of growth and that the maximum activity was obtained after 24 h of incubation. Production of inhibitory activity was detected in cultures grown between 35 and 39 degrees C and in a pH range of 6.9 to 9.4. The inhibitory substance was purified by methanol extraction, acetone fractionation, dialysis, and chromatography on Ultrogel AcA 54. The characterization of the inhibitor showed that it was a lipoprotein or a lipid-associated-protein and that the protein component could be separated from the lipids when chromatographed on Ultrogel AcA 54 in the presence of urea. The inhibitory activity was associated with the protein component which had a molecular weight of approximately 15,900. In polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and urea, the protein component was dissociated in a subunit estimated to be approximately 1,400 daltons.
In the course of a study dealing with the biodegradation of 4-chlorobiphenyl by strain B-206, we noticed that the gram-negative bacterium accumulated different metabolic intermediates depending on the nitrogen source of the medium. Hence, in the presence of nitrate, strain B-206 produced four compounds which were identified as 2- and 4-hydroxy-4'-chlorobiphenyl and 2- and 4-hydroxy-mononitro-4'-chlorobiphenyl. The accumulation of these compounds in the culture medium indicated the presence of a monooxygenase in strain B-206 leading to the production of arene oxide intermediates. The possible transformation of 4-chlorobiphenyl to an arene oxide by this bacterial strain is a matter of concern because of the high reactivity of these arene oxides with biological material.