La distribution des gènes ermA, B, C, D et F de résistance aux macrolides, lincosamides et streptogramines de type B (MLSB), chez 105 souches d'enteronactéries hautement résistantes aux MLSB, a été étidiée par hybridation sur colonies. Le gène ermC a été détecté chez une souche de Escherichia coli (BM2576), et il apparaît qu'un minimum de 3 classes de gènes confèrent la résistance aux MLSB chez les entérobactéries. Ces résultats confirment l'existence de transfert d'information génétique dans les conditions naturelles des cocci à Gram-positif vers les bactéries à Gram-négatif. Dans le cas de ermC, il pourrait s'agir d'un transfert ancien avec adaptation subséquente au nouvel hôte.
The activity of two enzymes involved in acetone production in Clostridium acetobutylicum, acetoacetate decarboxylase and coenzyme A transferase, was studied under acidogenic or solventogenic conditions. Acetoacetate decarboxylase activity was low under acidogenic conditions and after pyruvate addition. Under the same conditions, coenzyme A transferase activity was high. A mutant which lacked acetoacetate decarboxylase activity but was positive for coenzyme A transferase activity was isolated.
Ten isolates of Bacillus sphaericus from Ghana, very toxic to mosquito larvae, have been identified as belonging to serotype H6. These isolates can be represented by the head-group strain IAB59. They form crystals at the sporulation stage. Their larvicidal effect on Culex pipiens and Anopheles stephensi larvae is as high as that of the most toxic strains already known, e.g. 1593 and 2362 (serotype H5a,5b) and 2297 (serotype H25). Spore-crystal extracts of all these strains contain a 43-Kd polypeptide immunologically related to the 43-Kd polypeptide from strain 2362 described by other authors.
The genetic determinants resposible for the adherence of Escherichia coli to uroepithelial cells have been identified in recent years by genetic and molecular methods. Specific DNA probes for each of the three operons which have been cloned so far (pap, afa, sfa/foc operons) have been used in colony hybridization experiments to detect the presence of each of these operons in the chromosomal DNA of 443 strains of E. coli; 186 strains were frompatients with urinary tract infections (pyelonephritis, 106 strains; cystitis, 59; asymptomatic bacteriuria, 21) and 257 were strains from the stools of healthy subjects (61) or from patients with various enteral infections (196). E.coli strains harbouring the pap operon were found more frequently in the urine of patients with pyelonephritis (ifp < 0.001) and cystitis (p < 0.01) than in control stools. The presence of two operons (pap+afa) or (pap+sfa/foc) was only observed in uropathogenic strains. (p < 0.02).
The bactericidal and fungicidal effects of five disinfectants and one combination of two disinfectants were tested using a modified Kelsey-Sykes method in which living microorganisms suspended in a sterilized yeast suspension («dirtyå conditions) and in sterile distilled water («cleanå conditions) were added to the disinfectants in three stages. Six bacteria and two fungal organisms were employed as test microbes.
The extracellular and intracellular activities of 4 antituberculous drugs (rifampicin, streptomycin, ansamycin and kanamycin) and 7 beta-lactam antibiotics (penicillin G, amoxycillin, carbenicillin, ticarcillin, ampicillin, cloxacillin and imipenem) were determined against the bacteria of the Mycobacterium avium complex (MAC). The extracellular activities were determined against 14 strains by the broth-dilution method, whereas the intracellular activities were determined against type strain ATCC 15769 multiplying actively inside murine J-774 macrophages. Six of the beta-lactam drugs (except the beta-lactamase-stable drug imipenem) were also used in association with the beta-lactamase inhibitor clavulanic acid. Our results showed that the screening of MAC strains against beta-lactams may be useful, and that correlation of usual in vitro minimal inhibitory concentrations with the intracellular activities of the same drugs against pathogenic mycobacteria, whenever feasible, would be the method of choice to designate the therapeutic protocols against these intracellular pathogens.
The interest of the image analysis procedure is the time-saving in automated planktonic bacterial counting and sizing, with the possibility of manual visual field control at all times. Bacterial biomass (in number and volume) and bacterial projected area histograms were determined with a microcomputer. Performance limits of image-analysed epifluorescence microscopy were: camera sensitivity, considering the very low fluorescence levels on stained bacteria; pixel-μm conversion factor, and the impossibility of the apparatus distinguishing between bacteria and fluorescent small particles. This method is not of interest for counting sediment bacteria.
A rapid, simple and preparative method is described for the recovery of the seven highest molecular weight proteins (HMWP) from Mycoplasma pneumoniae membrane. The yield of proteins obtained was approximately 90%. The method involved the separation of M. pneumoniae proteins by socium dodecyl sulphate polycrylamide gel electrophoresis (SDS-PAGE), followed by electroelution of HMWP.
Quarante-deux souches de Legionella de référence représentatives de 35 espéces ou sérogroupes et 53 souches isolées soit de malades (14) soit de l'environment (39), ont été étudiées.
The specificity of Mycobacterium tuberculosis anti-PGL-Tb1 antiserum prepared in rabbits was evaluated by an enzyme-linked immunosorbent assay. It was found that the antiserum immunoreacted with its homologous antigen but not with purified phenolic glycolipids from M. bovis BCG (mycoside B), M. kansasii (mycoside A), M. leprae (PGL-I) or M. marinum (mycoside G).
DNA of type strain Campylobacter pylori NCTC 11637 and 32 other strains of C. pylori recovered from gastric biopsy specimens was examined by thermal denaturation for its guanine-plus-cytosine (GC) content.
A high-productivity process has been developed for the production of mature human interleukin-1 beta (IL-1 beta) from recombinant Escherichia coli strains. Conditions were found that allow high IL-1 beta expression levels in high cell density cultures. Improved fed-batch fermentation strategies are described which include maintenance of glucose and acetate concentrations below 1 g/l and sparging the fermentor with an O2-enriched air supply. Using the E. coli tryptophan promoter control of transcription, a 2.2 g/l production level of IL-1 beta was achieved in E. coli B at cell densities of 55 g dry weight per litre. Another genetic construction involving the bacteriophage lambda cIts-PR expression cassette allowed a similar IL-1 beta production level (1.9 g/l) in E. coli E103S, albeit at a lower cell density (30 g/l). A simplified procedure allowing the purification of fully active IL-1 beta is also presented.
Two Legionella-like organisms were isolated from cooling-tower water samples in Czechoslovakia. They were presumptively identified as legionellae by their growth on buffered charcoal-yeast extract agar (BCYE) containing L-cysteine and their absence of growth on BCYE without L-cysteine. Both strains contained predominantly branch-chained cellular fatty acids and were therefore definitively placed in the genus Legionella. They were serologically distinct from other described Legionella species and were shown by DNA studies to constitute two new Legionella species, Legionella moravica (type strain 316-36; ATCC 43877) and Legionella brunensis (type strain 441-1; ATCC 43878). Two Legionella-like organisms were isolated from cooling-tower water samples in Czechoslovakia. They were presumptively identified as legionellae by their growth on buffered charcoal-yeast extract agar (BCYE) containing L-cysteine and their absence of growth on BCYE without L-cysteine. Both strains contained predominantly branch-chained cellular fatty acids and were therefore definitively placed in the genus Legionella. They were serologically distinct from other described Legionella species and were shown by DNA studies to constitute two new Legionella species, Legionella moravica (type strain 316-36; ATCC 43877) and Legionella brunensis (type strain 441-1; ATCC 43878).
Extracellular D-glucose oxidation by 5 enterobacterial species was studied with the purpose of selecting conditions useful for taxonomic studies. Extracellular production of gluconate from 14C-glucose by bacterial cells was evidenced by DEAE-cellulose paper chromatography. Escherichia coli oxidized glucose only when pyrroloquinoline quinone (PQQ) was added, whereas Serratia marcescens, Yersinia frederiksenii, Erwinia cypripedii and Cedecea lapagei oxidized D-glucose without added PQQ. 2-Deoxyglucose was found to be an excellent non-metabolized analogue of D-glucose in oxidation experiments. D-glucose oxidation was inhibited by KCN, p-chloromercuribenzoic acid and carbonyl cyanide m-chlorophenylhydrazone; and activated by p-benzoquinone. Iodoacetate had no action. Comparative cellulose thin-layer chromatography including 2-ketogluconate and 2,5-diketogluconate (produced by Janthinobacterium lividum) as standards, showed that gluconate was oxidized to 2-ketogluconate by S. marcescens and E. cypripedii, and 2-ketogluconate was oxidized to 2,5-diketogluconate by E. cypripedii. The diversity of D-glucose oxidation products in the Enterobacteriaceae could have some taxonomic applications.
The envelope structure of Branhamella catarrhalis was studied by electron microscopy and compared with that of other bacteria of the family Neisseriaceae, such as Moraxella lacunata subsp. liquefaciens and Neisseria gonorrhoeae. Negative staining of B. catarrhalis showed a mamilliform surface similar to that of Moraxella. On thin sections, the cell wall appeared to be made up of a wavy outer membrane tightly linked to a straight peptidoglycan layer. Spicule-like structures protruded from the cell surface. Ruthenium red staining revealed that they contained polysaccharides. While the outer polysaccharide layer of N. gonorrhoeae was unstable after repeated subcultures in vitro, this layer remained stable in B. catarrhalis and in Moraxella lacunata subsp. liquefaciens. La structure des enveloppes de Branhamella catarrhalis a été étudiée en microscopie électronique et comparée à celle des enveloppes d'autres bactéries de la famille des Neisseriaceae, telles que Moraxella lacunata subsp. liquefaciens et Neisseria gonorrhoeae. La coloration négative de B. catarrhalis a montré une surface mamelonnée semblable à celle de Moraxella. En coupe fine, la paroi cellulaire est constituée d'une membrane externe ondulée formant des arches en étroite relation avec la couche rigide du peptidoglycane. Des structures en forme de spicules irradient à partir de la surface cellulaire et semblent responsables des phénomènes d'agglutination bactérienne normalement recontrés chez cette bactérie. La coloration au rouge de ruthénium montre que ces structures contiennent des polyosides. Tandis que la couche externe de polyosides de N. gonorrhoeae montre une grande instabilité, cette couche demeure stable aussi bien chez B. catarrhalis que chez Moraxella lacunata subsp. liquefaciens, dans des conditions identiques.
We devised a procedure to construct strains of Escherichia coli which expose at their surface a foreign antigen genetically inserted into LamB, an outer membrane protein. In particular, we showed that amino acid residues 93-103 of poliovirus type 1 capsid polypeptide VP1, which correspond to the C3 neutralization epitope, when inserted into two different external loops of LamB (after residues 153 and 374 of the mature protein), yielded the synthesis of stable hybrid proteins named, respectively, 153-C3 and 374-C3. The poliovirus epitope was accessible to monoclonal antibody C3 at the cell surface. In the present work, these two hybrid proteins were injected into rabbits by the intravenous route in the form of live recombinant bacteria, and the humoral response to the poliovirus epitope was studied. With construction 153-C3, the subcutaneous route was also assayed using solubilized hybrid protein. The C3 viral sequence inserted in the two different regions of LamB were found to be immunogenic. Different types of antibodies specific to the C3 peptide were raised with the two construction: anti-peptide and antiviral particle antibodies. These first results indicate that the LamB presentation vector system constitutes a mode of peptide coupling which may lead to the elaboration of a new type of live vaccine.
A rapid co-agglutination test using monospecific antisera was developed for the serological typing of enteropathogenic strains of Yersinia enterocolitica. A total of 70 bacterial strains (17 reference strains and 53 clinical isolates) were examined. Absorption of immune sera against srotypes O:3, O:8 and O:9 with their heterologous antigens (S-LPS) was necessary to avoid the appearance of different cross-reactions, as observed by co-agglutination. The proteins present in the S-LPS preparations obtained from each serotype seemed to be responsible for such cross-reactions. Results obtained with a total of 57 clinical isolates belonging to other members of the family Enterobacteriaceae indicated a high specificity of the assay.
A microtechnique is described which permits simple evaluation of the activity of the enzyme adenosine phoshorylase (AdoP), present in all mycoplasmas tested thus far. The good solubility and stability of AdoP and the sensitivity of the assay should be advantageous in detecting mycoplasmas in biological samples such as animal sera, cell cultures and vaccines.
