Klebsiella pneumoniae KE-1 degrades endosulfan without formation of the toxic metabolite,endosulfan sulfate

For bioremediation of toxic endosulfan, endosulfan degradation bacteria, which do not form toxic endosulfan sulfate, were isolated from various soil samples using endosulfan as sole carbon and energy source. Among the 40 isolated bacteria, strain KE-1, which was identified as Klebsiella pneumoniae by physiological and 16S rDNA sequence analysis, showed superior endosulfan degradation activity. Analysis of culture pH, growth, free sulfate and endosulfan and its metabolites demonstrated that KE-1 biologically degrades 8.72 microg endosulfan ml(-1) day(-1) when incubated with 93.9 microg ml(-1) endosulfan for 10 days without formation of toxic endosulfan sulfate. Our results suggest that K. pneumoniae KE-1 degraded endosulfan by a non-oxidative pathway and that strain KE-1 has potential as a biocatalyst for endosulfan bioremediation.     

Monoclonal antibody of IgG isotype against a cross-reactive lipopolysaccharide epitope of Chlamydia and Salmonella Re chemotype enhances infectivity in L-929 fibroblast cells

A murine monoclonal antibody (MAb) 202D7 of IgG3 isotype recognizes a lipopolysaccharide (LPS) epitope of Chlamydia spp. and cross-reacts with the Re chemotype LPS of Salmonella and Escherichia coli. The antibody exhibits strong complement activating properties and stimulates phagocytosis of Salmonella enterica serovar Minnesota Re mutant by murine macrophages. Salmonella Re mutants are non-invasive for cell monolayers but still can enter and replicate in L-929 murine fibroblast cells. The entry of bacteria within the cells increases five-fold in the presence of MAb 202D7. The antibody mediates attachment and enhances five-fold the infectivity of Chlamydia pneumoniae into L-929 cells, which suggests a possible IgG-mediated mechanism of entry and survival of the pathogen in fibroblast cells.     

Characterisation of 20-kDa lectin-spermagglutinin from Arum maculatum that prevents Chlamydia pneumoniae infection of L-929 fibroblast cells

A novel lectin from the root of Arum maculatum was isolated by saline extraction and purified by cold ethanol precipitation and subsequent fractionation on Superose 6 column. The lectin named A. maculatum agglutinin is a non-glycosylated protein with 20-kDa molecular mass agglutinating human ejaculated spermatozoa, but not human erythrocytes. The agglutination was blocked in the presence of N-acetylneuraminic acid indicating that the lectin is sialoglycoprotein specific. Chlamydia pneumoniae strain AR-39 showed considerable potential to grow in murine L-929 fibroblast cells. Pretreatment of the cell monolayers with purified lectin reduced the entry and intracellular replication of C. pneumoniae. These results suggest that the isolated lectin prevents attachment by binding to a C. pneumoniae specific sialoglycoprotein receptor expressed on the surface of L-929 fibroblast cells.     

Crystal structure of a stress inducible protein from Mycoplasma pneumoniae at 2.85 Å resolution

Journal of Structural and Functional Genomics -     

Conservation of the mosaic structure of the four internal transcribed spacers and localisation of the rrn operons on the Streptococcus pneumoniae genome

The detection of heterogeneity of the 16S-23S ribosomal intergenic transcribed spacer (ITS) region has become rather common over the past years for identification and typing purposes of bacteria. The ITS not only varies in sequence and length, but also in number of alleles per genome and in their position on the chromosome together with the ribosomal clusters. The ITS characterisation has allowed discrimination of several species within a genus and variation in ITS sequences between the multiple rrn operons present within a genome may be as high or greater than between strains of the same species or subspecies. It is important to understand the variability of ITS sequences in a given genome to gain insights into bacterial physiology and taxonomy. The present study describes the possibility to type Streptococcus pneumoniae by PCR-ribotyping of the spacer region, the determination of the molecular structure of the ITS, and the determination of the number and localisation of rrn operons in this microorganism. Our results show that the genome of S. pneumoniae contains four ribosomal operons, showing the same genomic organisation among strains, each containing a single ITS allele of 270 bp. The ITS sequence presents a mosaic organisation of blocks highly conserved intra- and inter-species within the genus Streptococcus, giving no possibility for variations to arise.     

Anti-Chlamydia pneumoniae heat shock protein 10 antibodies in asthmatic adults

A multicenter prospective study was performed on 160 asthmatic adults suffering from acute episodes of bronchitis and 88 non-asthmatic controls, to investigate potential associations among Chlamydia pneumoniae infection and/or anti-C. pneumoniae heat shock protein 10 antibodies, and asthma. We used micro-immunofluorescence to detect serum anti-C. pneumoniae IgG, IgA and IgM antibodies and enzyme-linked immunosorbent assay to detect serum anti-Chsp10 peptide IgG antibodies. The serological prevalence of C. pneumoniae was 73.1%. An association was observed between the presence of anti-Chsp10 antibodies and adult onset asthma. The humoral immune responses were not confined to any particular region of the Chsp10 protein.     

Streptococcus pneumoniae heat shock protein 70 does not induce human antibody responses during infection

Mouse monoclonal antibodies (mAbs) were developed against Streptococcus pneumoniae in search for potential common pneumococcal proteins as vaccine antigens. mAb 230,B-9 (IgG1) reacted by immunoblotting with a 70-kDa protein which was isolated by immunoaffinity chromatography and subsequent preparative electrophoresis. N-terminal amino acid sequencing showed homology to that of heat shock protein 70 (hsp70). The hsp70 epitope reactive with mAb 230,B-9 was found in all the pneumococci examined as well as in other streptococci and enterococci. The epitope was not expressed in several other examined Gram-positive or -negative bacteria. Pneumococcal hsp70 has by other investigators been proposed to be a vaccine candidate. Binding experiments using flow cytometry showed that the epitope was not surface-exposed on live exponential phase grown S. pneumoniae. Human patient sera did not react with affinity-purified pneumococcal hsp70. Therefore the pneumococcal hsp70 does not seem to be of special interest in a vaccine formulation. The human sera contained antibodies to high molecular proteins co-purified with hsp70. Some of these proteins could be the pneumococcal surface protein A.     

Protective effect of beta-glucan against systemic Streptococcus pneumoniae infection in mice

The antimicrobial effect of soluble beta-1,3-D-glucan from Sclerotinia sclerotiorum (SSG) was examined in mice experimentally infected intraperitoneally (i.p.) with Streptococcus pneumoniae serotypes 4 and 6B. SSG was administered i.p. either 3 days before challenge or 3-48 h after challenge. The number of bacteria in blood samples and the mouse survival rates were recorded. Pre-challenge SSG administration protected dose-dependently against both S. pneumoniae type 4 and 6B infections. SSG injected 24 h post-challenge had a curative effect against type 6B but not type 4 pneumococcal infection. The data demonstrate that SSG administered systemically protects against pneumococcal infection in mice.     

Interaction with magnesium and ADP stabilizes both components of nitrogenase from Klebsiella pneumoniae against urea denaturation

The nitrogenase enzyme of Klebsiella pneumoniae consists of two separable proteins, each with multiple subunits and one or more oxygen sensitive metallocenters. The wild-type nitrogenase proteins are stable to electrophoresis in high concentrations of urea under anaerobic conditions. Addition of Mg+2 and ADP greatly increases the stability of the smaller Fe protein (from <4 to >6 M for full unfolding), an effect directly analogous to stabilization in p21ras induced by Mg+2 and GDP. Stabilization by Mg+2 is slight for the holo MoFe protein (from approximately 1.5 to approximately 2.4 M) but more dramatic for the apo protein form of the MoFe protein accumulated by certain Fe protein (nifH gene) mutants. The potent product inhibitor of nitrogenase function, MgADP, increases stability of the MoFe protein more than Mg+2 alone, to approximately 3.6 M, showing that nucleotides interact with the MoFe protein. Mutations of the nifM gene result in slower accumulation of less stable Fe protein, indicating that NifM is involved in correct folding of the Fe protein. Mutationally altered proteins are often difficult to purify for study because of their inherent instability, low expression level, or oxygen lability. Crude extracts of 11 different mutants of Fe protein (nifH gene) were examined by transverse urea gradient gels to rapidly screen for stabilizing interactions in the presence or absence of substrate or inhibitor analogs. Amino acid alterations D44N and R188C, at the interface of the dimer, in the vicinity of the nucleotide binding site(s), have significantly lower stability than the wild-type enzyme in the absence of Mg+2 but comparable stability in its presence, showing the importance of Mg+2 in the subunit interactions. Mutations N163S and E266K, in which residues normally involved in hydrogen bonding far from the active site were altered, are more labile than the wild-type even with Mg+2 added. Seven other mutants, though nonfunctional, did not appear altered in stability compared to the wild-type.     

Conversion of sucrose to isomaltulose by Klebsiella planticola CCRC 19112

An isomaltulose-producing bacterium was isolated and taxonomically characterized. Its morphological and biochemical properties conform best to those described for Klebsiella planticola. When cultured under optimal conditions, the organism simultaneously converted sucrose into both isomaltulose (α-D-glucopyranosyl-1,6-fructose) and trehalulose (α-D-glucopyranosyl-1,1-fructose) with substrate conversion rates of 80% and 15%, respectively. Sucrose and Bacto-tryptone were the most effective carbon and supplemental nitrogen sources, respectively, for producing cells of high isomaltulose-forming ability. None of several inorganic salts tested had any significant effect. The major product formed in the reaction mixture was verified to be isomaltulose by co-chromatography and IR spectroscopy. Received 21 April 1998/ Accepted in revised form 7 July 1998