bluuebunny/crossref_metadata_2025 huggingface dataset:
Rhodobacter capsulatus genes encoding form I ribulose-1,5-bisphosphate carboxylase/oxygenase (cbbLS) and neighbouring genes were acquired by a horizontal gene transfer
Authors: Paoli George C. , Soyer Ferda , Shively Jessup , Tabita F. Robert
Published: June 2009

Summary: Analysis of the nucleotide sequence of the form I ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) genes (cbbL and cbbS) of the non-sulfur purple bacterium Rhodobacter capsulatus indicated that the deduced amino acid sequence of the large subunit was not closely homologous to the large subunit from related organisms. Indeed, phylogenetic analysis suggested that the large subunit protein (CbbL) more closely resembled the enzyme from αlβlγ purple bacteria and cyanobacteria and is within a ‘green-like’ radiation of the RubisCO phylogenetic tree, well separated from CbbL of the related organism Rhodobacter sphaeroides. A cbbQ gene was discovered downstream of cbbS in Rh. capsulatus, a gene arrangement which also appears to be limited to certain organisms containing a ‘green-like’ RubisCO. Upstream, and divergently transcribed from cbbLSQ, is a gene (cbbR
I) that encodes a LysR-type transcriptional activator. Phylogenetic analysis of the deduced amino acid sequence of CbbRI also suggests that this protein is quite distinct from the Rh. sphaeroides CbbR protein, and is even distinct from the previously described CbbRII protein, the gene of which is upstream and divergently transcribed from the cbb
II operon of Rh. capsulatus. Interestingly, Rh. capsulatus CbbRI is more closely related to CbbR from bacteria whose RubisCO falls within the ‘greenlike’ radiation of the CbbL tree. These studies suggest that the cbbR
I-cbbL-cbbS-cbbQ genes were acquired by Rh. capsulatus via horizontal gene transfer from a bacterial species containing a ‘green-like’ RubisCO.
The two-component hybrid kinase regulator CaNIKl of Candida albicans
Authors: Srikantha Thyagarajan , Tsai Luong , Daniels Karla , Enger Lee , Highley Kumiko , Soll David R.
Published: June 2009

SUMMARY: Using degenerate primers of highly conserved regions of two-component response regulators for PCR amplification, a two-component response regulator was cloned from Candida albicans that is homologous to nik-l+ of Neurospora crassa. This two-component hybrid kinase, CaNIKl, also shows features of bacterial two-component response regulators, including a putative unorthodox second histidine kinase motif at the carboxy-terminal end. CaNIKl was expressed at low levels in both the white and opaque switch phenotypes and in the bud and hyphal growth forms of C. albicans strain WO-1, but in both developmental programmes, the level of transcript was modulated (levels were higher in opaque cells and in hyphae). Partial deletion of both CaNIKl alleles, by which the histidine autokinase- and ATP-binding domains were removed, did not inhibit either high-frequency phenotypic switching or the bud-hypha transition in high salt concentrations, but in both cases the efficiency of the developmental process was reduced.
Adaptation of Cornamonas testosteroni TAM1 to utilize phenol: organization and regulation of the genes involved in phenol degradatio
Authors: Arai Hiroyuki , Akahira Saiko , Ohishi Tohru , Maeda Michihisa , Kudo Toshiaki
Published: June 2009

SUMMARY: Comamonas testosteroni TAU1 was not able to grow on phenol as a sole carbon and energy source, but it gained the ability to utilize phenol after a 2-3-week incubation in a medium containing phenol. Phenol hydroxylase (PH) and catechol2,3-dioxygenase (C230) were highly induced by phenol in the adapted strain designated as strain P1, suggesting that phenol was degraded via the meta-pathway. Gene clusters for phenol degradation were isolated from both strains TAU1 and P1. The structural genes encoding multi- component PH and C230 (aphKLMNOPQB), and a regulatory gene of the NtrC family (aphR), were located in a divergent transcriptional organization. The cloned aphKLMNOPQl3 genes from either strain TAU1 or strain P1 produced active PH and C230 enzymes in strain TA441. No difference was found between the strains in the sequences of aphR and the intergenic promoter region of aphK and aphR. However, the transcriptional activities of the aphK and aphR promoters were higher in strain P1 than in strain TA441. The aphK-promoter activity was not observed in aphR mutant strains and these strains could not grow on phenol. The aphR mutant of strain P1 was able to grow on phenol after transformation with a recombinant aphR gene but strain TAM1 was not, suggesting that the expression of the aph genes is silenced by an unidentified repressor in strain TAU1 and that this repressor is modified in strain P1.
A Candida albicans chaperonin subunit (CaCct8p) as a suppressor of morphogenesis and Ras phenotypes in C. albicans and Saccharomyces cerevisiae
Authors: Rademacher Felicitas , Kehren Verena , Stoldt Volker R. , Ernst Joachim F.
Published: June 2009

SUMMARY: Saccharomyces cerewisiae and the pathogen Candida albicans can be induced to undergo morphogenesis from a yeast to a filamentous form. A C. albicansgene (CaCCT8) was identified encoding a subunit of the Cct chaperonin complex, whose expression prevents filament formation in both fungi without interfering with growth of the yeast form. In 5. cerewisiae, pseudohyphal growth induced by Ra2 119va, by overproduction of Phdlp or by expression of the C. albicans EFGl gene, was blocked by CaCct8p and its N-terminally deleted derivative CaCct8-Alp; in contrast, pseudohyphal induction by othe components (Cphlp, Cdc42p) could not be suppressed, indicating that morphogenesis per se is not inhibited. CaCCT8 expression also interfered with other Ra2p va119, phenotypes, including heat sensitivity, lack of glycogen accumulation and lack of sporulation. In C. albicans, overproduction of CaCct8p effectively blocked hyphal morphogenesis induced by starvation conditions and by serum. The results suggest that the activity of a component in the Ras2p signal transduction pathway is suppressed by excess chaperonin subunits. This component may be a novel folding target for the Cct complex. In agreement with this hypothesis, disruption of one of the two CaCC7'8 alleles in C. albicans led t o defective hyphal morphogenesis.
Truncation as a novel form of variation of the p50 gene in Mycoplasma hominis
Authors: Henrich Birgit , Lang Klaudia , Kitzerow Annette , MacKenzie Colin , Hadding Ulrich
Published: June 2009

SUMMARY: A characteristic feature of the mycoplasmas is the presence of variable surface proteins which may play an important role in the adaptation of the cell-wall-less organisms to their host environments. In addition, this antigen variation may be an important pathogenic property of the organism. The ubiquity of the gene encoding P50, an adhesin of Mycop/asrna hominis FBG, and its transcription were analysed in different isolates ofM. horninis. The p50 gene was present in all isolates tested. Based on Southern blot analysis and sequencing of the gene, the isolates could be classified into one of three
distinct groups. Within two groups specific truncations of the p50 gene occurred. The reduction of the gene size was confirmed in Northern blot analysis of representative isolates from each group, with a decrease in transcript length from 1.6 kb in group G-1 down to 076 kb in group 6-3. In addition to truncation, a coincidental duplication of some gene segments was detected. This work has provided evidence for the genetic basis of a further variation in the M. horninis P50 adhesin.
Characterization of Trichornonas vaginalis AP33 adhesin and cell surface interactive domains
Authors: Engbring J. A. , Alderete J. F.
Published: June 2009

SUMMARY: Adherence to host target cells is a critical step in establishing infection with the sexually transmitted pathogen Tiichomonas vaginalis. Four parasite surface proteins mediating attachment to vaginal epithelial cells have been identified. One surface protein, termed AP33, was characterized further to identify domains interactive with previously generated antibodies and with host surface sites. N- and C-terminal deletion subclones were generated and tested for reactivity with both mAb and rabbit antiserum against AP33, and were also examined for their ability to bind to host cells. Surprisingly, the rabbit antiserum known to inhibit cytoadherence recognized an epitope(s) contained within 72 residues in the N-terminal half of the protein. However, the mAb epitope was immunoreactive with a 28-amino-acid region near the C-terminus. Subsequent mapping of this region with overlapping peptides identified a nine-amino-acid sequence reactive with the mAb. Equally surprising, two domains interactive with host cell surfaces were identified at distinct parts of AP33: one in the N-terminal half of the protein, and the other within 24 residues in the C-terminal third. Further analysis of the C-terminal binding domain revealed that a peptide representing this area could inhibit T. vaginalis cytoadherence by 40%.
A lipid A-associated protein of Porphyromonas gingivalis, derived from the haemagglutinating domain of the RI protease gene family, is a potent stimulator of interleukin 6 synthesis
Authors: Sharp Lindsay , Poole Stephen , Reddi Krisanavane , Fletcher Julie , Nair Sean , Wilson Michael , Curtis Michael , Henderson Brian , Tabonal Peter
Published: June 2009

SUMMARY: There is evidence that the lipid A-associated proteins (LAPs) of enteric bacteria can induce the synthesis of interleukin 1 (IL-1) and therefore may be important virulence factors. Porphyromonas gingivdis is now recognized as a major pathogen in the chronic inflammatory periodontal diseases and it has previously been reported that a crude LAP fraction from this organism could induce IL-1 and interleukin 6 (IL-6) synthesis. In the present study the chemical and biological properties of the LAPs of this bacterium have been further characterized. Analysis by SDS-PAGE has shown that the LAPs comprise nine proteins of molecular masses 81,68,48,47,28,25,20,17 and 16 kDa. These LAPS, at concentrations as low as 100 ng mV, were shown to stimulate human gingival fibroblasts, human peripheral blood mononuclear cells and whole human blood t o produce the pro-inflammatory cytokine IL-6. The cytokine-inducing activity of the LAPs was reduced after heat-inactivation and trypsinization, suggesting that the activity was not due to contaminating LPS. To establish which proteins in this mixture were the active cytokine inducers, the LAPs were separated by electrophoresis on polyacrylamide gels. The majority of the activity was associated with the 17 kDa LAP. N-terminal sequence analysis demonstrated that this protein was homologous t o an internal region of a conserved adhesin domain contained within a family of P. gingivdis extracellular proteins including the RI protease, Lys-gingipain, porphypain and haemagglutinin A. In addition to a role in adherence, the adhesin domain(s) of these proteins may also have cytokine-inducing properties. Furthermore, it has also been shown that the previously observed degradation of cytokines by P. gingivelis may be attributable to the catalytic domain of the RI protease. Thus, different domains within the same molecule appear t o have opposing actions on pro-inflammatory cytokine levels and the balance between these two activities may influencethe cytokine status of the periodontiurn in patients with the common chronic inflammatory conditions known as the periodontal diseases.
Streptococcus pyogenes protein F promotes invasion of HeLa cells
Authors: Okada Nobuhiko , Tatsuno lchiro , Hanski Emanuel , Caparon Michael , Sasakawa Chihiro
Published: June 2009

SUMMARY: Although the Gram-positive bacterium Streptococcus pyogenes (group A streptococcus) has been considered an extracellular pathogen which adheres to human mucosal epithelium, the streptococcus possesses invasive capacity for cultured human epithelial cells. This study provides genetic and functional evidence supporting the conclusion that protein F is capable of mediating entry of S. pyogenes into HeLa cells. Using 111916 insertion mutagenesis or an isogenic 5. pyogenes strain with a defined mutation in the gene encoding protein F (prtF), it was observed that the invasive capacity was affected by the levels of surface-exposed protein F, but not by those of M protein. In addition, heterologous expression of protein F on Enterococcus faecalis conferred upon the bacteria an efficient invasive phenotype. Several assays demonstrated that both the fibronectin-binding domains of protein F, UR and RD2, were involved in host-cell invasion. In addition, coinfection experiments of HeLa cells with 5. pyogenes and an Escherichia coli K-12 strain expressing an afimbrial adhesin AFA-I showed that the uptake of S. pyogenes did notpermit internalization of the E. coli cells.
Analysis of the effect exerted by extracellular pH on the maltose regulon in Escherichia coli K-12
Authors: Alonzo Sylvie , Heyde Martine , Laloi Patrick , Portalier Raymond
Published: June 2009

Summary: The Escherichia coli maltose regulon consists of five operons under the control of the MalT transcriptional activator. lac operon fusions were constructed in vitro with the MalT-dependent promoter and with the malT promoter itself. -Galactosidase activity displayed by these fusions during growth at different external pH (pHo) revealed that growth at a pHo higher than 6 stimulates the transcription of malT- and MalT-controlled genes in the absence or presence of maltose. Using a malTp1 malTp10 promoter that is cAMP-CRP (cAMP receptor protein)-independent, it was demonstrated that CRP is essential for malT pHo regulation and that the pHo-dependent activity of malKp is a direct consequence of malT regulation. The pHo regulation displayed by a deleted but still functional malT promoter fused to lacZ demonstrates that this minimal promoter contains all the regulatory regions for establishing pHo regulation. In the absence of Mlc, a repressor of malT expression, the pHo regulation of malT was still effective. It is proposed that binding of cAMP-CRP at malTp may be affected by malTp topology induced by pHo or that a pHo-dependent effector may act in concert with the cAMP-CRP complex.
Production of a non-toxic site-directed mutant of Clostridium perfringens ε-toxin which induces protective immunity in mice
Authors: Oyston Petra C. F. , Payne Dean W. , Havard Helen L. , Williamson E. Diane , Titball Richard W.
Published: June 2009

A panel of ten site-directed mutants ofClostridium perfringensε-toxin was generated. All of the mutated proteins expressed inEscherichia coliwere recognized in immunoblots by a neutralizing mAb raised against wild-type native ε-toxin. The cytotoxicity of the site-directed mutated toxins was assayedin vitroagainst MDCK cells. One mutation resulting in loss of activity in the assay was identified. This non-toxic protein was derived by substituting a proline for the histidine at residue 106 of the toxin. Immunization of mice with the non-toxic mutated ε-toxin resulted in the induction of a specific antibody response and immunized mice were protected against 1000 LD50doses of wild-type recombinant ε-toxin.
Requirement for ubiquinone downstream of cytochrome(s) b in the oxygen-terminated respiratory chains of Escherichia coli K-12 revealed using a null mutant allele of ubiCA
Authors: balle Britta Sø , Poole Robert K.
Published: June 2009

AnEscherichia coliknockoutubiCAmutant has been constructed using a gene replacement method and verified using both Southern hybridization and PCR. The mutant, which was unable to synthesize ubiquinone (Q), showed severely diminished growth yields aerobically but not anaerobically with either nitrate or fumarate as terminal electron acceptors. Low oxygen uptake rates were demonstrated in membrane preparations using either NADH or lactate as substrates. However, these rates were greatly stimulated by the addition of ubiquinone-1 (Q-1). The rate of electron transfer to those oxidase components observable by photodissociation of their CO complexes was studied at sub-zero temperatures. In theubiCAmutant, the reduced form of haemoproteins - predominantly cytochromeb595-was reoxidized significantly faster in the presence of oxygen than in a Ubi+strain, indicating the absence of Q as electron donor. Continuous multiple-wavelength recordings of the oxidoreduction state of cytochrome(s)bduring steady-state respiration showed greater reduction in membranes from theubiCAmutant than in wild-type membranes. A scheme for the respiratory electron-transfer chain inE. coliis proposed, in which Q functions downstream of cytochrome(s)b.
A role for the MAP kinase gene MKC1 in cell wall construction and morphological transitions in Candida albicans
Authors: Navarro-García Federico , Alonso-Monge Rebeca , Rico Hortensia , Pla Jesús , Sentandreu Rafael , Nombela César
Published: June 2009

TheCandida albicans MKC1gene encodes a mitogen-activated protein (MAP) kinase, which has been cloned by complementation of the lytic phenotype associated withSaccharomyces cerevisiae slt2(mpk1) mutants. In this work, the physiological role of this MAP kinase in the pathogenic fungusC. albicanswas characterized and a role forMKC1in the biogenesis of the cell wall suggested based on the following criteria. First,C. albicans mkc1Δ/mkc1Δ strains displayed alterations in their cell surfaces under specific conditions as evidenced by scanning electron microscopy. Second, an increase in specific cell wall epitopes (O-glycosylated mannoprotein) was shown by confocal microscopy inmkc1Δ/mkc1Δ mutants. Third, the sensitivity to antifungals which inhibit (1,3)-β-glucan and chitin synthesis was increased in these mutants. In addition, evidence for a role for theMKC1gene in morphological transitions inC. albicansis presented based on the impairment of pseudohyphal formation ofmkc1Δ/mkc1Δ strains on Spider medium and on the effect of its overexpression onSacch. cerevisiaecolony morphology on SLADH medium. Using the two-hybrid system, it was also demonstrated thatMKC1is able to interact specifically withSacch. cerevisiaeMkk1p and Mkk2p, the MAP-kinase kinases of thePKC1-mediated route ofSacch. cerevisiae, and to activate transcription inSacch. cerevisiaewhen bound to a DNA-binding element. These results suggest a role for this MAP kinase in the construction of the cell wall ofC. albicansand indicate its potential relevance for the development of novel antifungals.
Expression of the second lysine decarboxylase gene of Escherichia coli
Authors: Lemonnier Marc , Lane David
Published: June 2009

Certain amino acids are substrates for two decarboxylase enzymes inEscherichia coli, one inducible by anaerobic growth at low pH and the other constitutive. In the case of lysine, an inducible decarboxylase (CadA) has been extensively characterized, but evidence for the existence of a second lysine decarboxylase is fragmentary and uncertain. This paper confirms that a second lysine decarboxylase is encoded by a locus (Idc) previously suggested to be a lysine decarboxylase gene on the basis of sequence comparisons. Overexpression of the cloned gene provided sufficient quantities of enzyme in cell-free extracts for preliminary examination of the properties of theIdcgene product, Ldc. The enzyme is active over a broad range of pH with an optimum at 7.6, much higher than that of CadA, about 5.5. The temperature optimum for both enzymes is similar, at about 52 °C, but Ldc is more readily inactivated by heat than CadA. Expression ofIdcfrom its own promoter was very weak for cells growing in a variety of media, although a low level of lysine decarboxylase was present in cells that carried theIdcregion on an oligo-copy plasmid when these were grown in minimal-glucose medium. Northern analysis of RNA extracted from such cells revealed a transcript whose length corresponded to that of theldcgene, suggesting thatldcis normally transcribed from a promoter immediately upstream. However, most of theldcmRNA was shorter, indicating degradation or premature termination. Theldcupstream sequence promoted transcription of alacZgene to which it was fused. Introduction of the upstream sequence as an insert in a multicopy vector increased transcription of the residentlacZfusion. The low level of expression in single copy, the emergence of expression when the gene is present at moderate copy number, and the derepression by the upstream sequencein transimply that this second lysine decarboxylase gene may not be constitutive but subject to specific repression by a factor which remains to be identified.
Oligopeptide permease in Borrelia burgdorferi: putative peptide-binding components encoded by both chromosomal and plasmid loci
Authors: Bono James L. , Tilly Kit , Stevenson Brian , Hogan Dan , Rosa Patricia
Published: June 2009

To elucidate the importance of oligopeptide permease forBorrelia burgdorferi, the agent of Lyme disease, a chromosomal locus inB. burgdorferithat encodes homologues of all five subunits of oligopeptide permease has been identified and characterized.B. burgdorferihas multiple copies of the gene encoding the peptide-binding component, OppA; three reside at the chromosomal locus and two are on plasmids. Northern analyses indicate that eachoppAgene is independently transcribed, although the three chromosomaloppAgenes are also expressed as bi- and tri-cistronic messages. Induction of one of the plasmid-encodedoppAgenes was observed following an increase in temperature, which appears to be an important cue for adaptive responsesin vivo. The deduced amino acid sequences suggest that all five borrelial OppA homologues are lipoproteins, but the protease-resistance of at least one of them in intact bacteria is inconsistent with outer-surface localization. Insertional inactivation of a plasmid-encodedoppAgene demonstrates that it is not essential for growth in culture.
Tandem organization and highly disparate expression of the two laccase genes lcc1 and lcc2 in the cultivated mushroom Agaricus bisporus
Authors: Smith M. , Shnyreva A. , Wood D. A. , Thurston C. F.
Published: June 2009

Two non-allelic laccase genes (lcc1 and lcc2) in Agaricus bisporus have been mapped to the same cosmid clone and are close together, in tandem. The intergenic region consists of 1562 bp between the stop codon of lcc1 and the start codon of lcc2. Differences between the 5′ non-coding regions of the two genes suggest the potential for their differential regulation. By employing competitive RT-PCR and specific primer pairs that discriminate between lcc1 and lcc2, it has been shown that the level of lcc2 mRNA is approximately 300 times higher than that of lcc1 mRNA in malt extract liquid cultures; in compost cultures lcc2 mRNA is almost 7000 times more abundant than lcc1 mRNA.
Repetitive sequences (RPSs) in the chromosomes of Candida albicans are sandwiched between two novel stretches, HOK and RB2, common to each chromosome
Authors: Chindamporn Ariya , Nakagawa Yoshiyuki , Mizuguchi Ikuyo , Chibana Hiroji , Doi Matsuko , Tanaka Kenji
Published: June 2009

A novel sequence designated HOK, which is next to the RPS, a repetitive sequence specific to Candida albicans, was cloned and sequenced. HOK hybridized with all of the chromosomes on which the RPSs were located, but did not hybridize with chromosome 3, which does not harbour any RPSs. Sequence determination revealed that a portion of HOK has significant homology with the B and C1 fragments of Ca3, which is used as a molecular epidemiological probe. A homology search of the deduced amino acids of HOK against the protein database showed partial homology with an isocitrate dehydrogenase of Saccharomyces cerevisiae, although an ORF large enough to encode the enzyme was not detected. To verify the existence of other sequences homologous with HOK, a portion of the HOK sequence was amplified using PCR. Sequence determination of the 41 clones from the PCR products resulted in at least six HOK-homologous clones. Another RPS-containing clone, RB2, was isolated from the Pstl-digested chromosome R or 1. It was determined that RB2a, one of the subclones from RB2, hybridized with all of the chromosomes, including chromosome 3, with which neither HOK nor RPS hybridized. The hybridization profile also showed that RPS is located between HOK and RB2a on chromosomes other than chromosome 3.
Bacterial alginate biosynthesis – recent progress and future prospects
Authors: Gacesa Peter
Published: June 2009

The extracellular polysaccharide alginate has been widely associated with chronic Pseudomonas aeruginosa infections in the cystic fibrosis lung. However, it is clear that alginate biosynthesis is a more widespread phenomenon. Alginate plays a key role as a virulence factor of plant-pathogenic pseudomonads, in the formation of biofilms and with the encystment process of Azotobacter spp.
Rapid identification of urinary tract infection bacteria using hyperspectral whole-organism fingerprinting and artificial neural networks
Authors: Goodacre Royston , Timmins Éadaoin M , Burton Rebecca , Kaderbhai Naheed , Woodward Andrew M. , Kell Douglas B. , Rooney Paul J.
Published: June 2009

Three rapid spectroscopic approaches for whole-organism fingerprinting-pyrolysis mass spectrometry (PyMS), Fourier transform infra-red spectroscopy (FT-IR) and dispersive Raman microscopy - were used to analyse a group of 59 clinical bacterial isolates associated with urinary tract infection. Direct visual analysis of these spectra was not possible, highlighting the need to use methods to reduce the dimensionality of these hyperspectral data. The unsupervised methods of discriminant function and hierarchical cluster analyses were employed to group these organisms based on their spectral fingerprints, but none produced wholly satisfactory groupings which were characteristic for each of the five bacterial types. In contrast, for PyMS and FT-IR, the artificial neural network (ANN) approaches exploiting multi-layer perceptrons or radial basis functions could be trained with representative spectra of the five bacterial groups so that isolates from clinical bacteriuria in an independent unseen test set could be correctly identified. Comparable ANNs trained with Raman spectra correctly identified some 80% of the same test set. PyMS and FT-IR have often been exploited within microbial systematics, but these are believed to be the first published data showing the ability of dispersive Raman microscopy to discriminate clinically significant intact bacterial species. These results demonstrate that modern analytical spectroscopies of high intrinsic dimensionality can provide rapid accurate microbial characterization techniques, but only when combined with appropriate chemometrics.
Genetic diversity in the Mycobacterium tuberculosis complex based on variable numbers of tandem DNA repeats
Authors: Frothingham Richard , Meeker-O’Connell Winifred A.
Published: June 2009

Genetic loci containing variable numbers of tandem repeats (VNTR loci) form the basis for human gene mapping and identification, forensic analysis and paternity testing. The variability of bacterial tandem repeats has not been systematically studied. Eleven tandem repeat loci in the M. tuberculosis genome were analysed. Five major polymorphic tandem repeat (MPTR) loci contained 15-bp repeats with substantial sequence variation in adjacent copies. Six exact tandem repeat (ETR) loci contained large DNA repeats with identical sequences in adjacent repeats. These 11 loci were amplified in 48 strains to determine the number of tandem repeats at each locus. The strains analysed included 25 wild-type strains of M. tuberculosis, M. bovis, M. africanum and M. microti and 23 substrains of the attenuated M. bovis BCG vaccine. One of the five MPTR loci and all six ETR loci had length polymorphisms corresponding to insertions or deletions of tandem repeats. Most ETR loci were located in intergenic regions where copy number may influence expression of downstream genes. Each ETR locus had multiple alleles in the panel. Combined analysis identified 22 distinct allele profiles in 25 wild-type strains of the M. tuberculosis complex and five allele profiles in 23 M. bovis BCG substrains. Allele profiles were reproducible and stable, as demonstrated by analyses of multiple isolates of particular reference strains obtained from different laboratories. VNTR typing may be generally useful for strain differentiation and evolutionary studies in bacteria.
Biological functions of UDP-glucose synthesis in Streptococcus mutans
Authors: Yamashita Yoshihisa , Tsukioka Yuichi , Nakano Yoshio , Tomihisa Kiyotaka , Oho Takahiko , Koga Toshihiko
Published: June 2009

A gene encoding glucose-1-phosphate uridylyltransferase (EC 2.7.7.9) was isolated fromStreptococcus mutans.A cell extract ofEscherichia coliexpressing the cloned gene exhibited glucose-1-phosphate uridylyltransferase activity. The enzyme catalyses the conversion of D-glucose 1-phosphate and UTP into UDP-D-glucose. Rabbit antiserum against the serotype-c-specific antigen did not react with autoclaved extracts from mutant cells in which the cloned gene was insertionally inactivated. The glucose content of the cell-wall preparation purified from the mutant was very much lowered, whereas there was no observable decrease in the content of rhamnose. When the mutant strain was grown in an acidic environment, its cell viability was much lower than that of the wild-type. These results suggest that UDP-D-glucose functions not only as an immediate precursor of the serotype-c-specific antigen ofS. mutans(as a glucose donor for side-chain formation), but is also important for the organism’s viability in environmental conditions of low pH.
Identification of two glycosylated components of Mycoplasma penetrans: a surface-exposed capsular polysaccharide and a glycolipid fraction
Authors: Neyrolles Olivier , Brenner Catherine , Prevost Marie-Christine , Fontaine Thierry , Montagnier Luc , Blanchard Alain
Published: June 2009

Among the wall-less mycoplasmas only a few species have been identified with a capsule at their cell surface. Mycoplasma penetrans is a recently identified mycoplasma with unique morphology, isolated from HIV-infected patients. Using transmission electron microscopy, it was found that M. penetrans is surrounded by capsular material 11 nm (strain GTU-54-6A1) to 30 nm (strain HF-2) thick, which can be stained with ruthenium red and labelled with cationized ferritin. The polysaccharide composition of this capsule was indicated by its staining with periodic acid-thiocarbohydrazide silver proteinate and the abolition of ruthenium red staining of the cell surface by neuraminidase treatment. In addition, proteinase K treatment of the M. penetrans cells resulted in removal of the capsule, suggesting that polypeptides may contribute in anchoring it to the membrane or in its stability. Two different types of glycosylated material were detected in mycoplasma extracts by SDS-PAGE and periodic acid-Schiff staining. The first component was a high-molecular-mass material, which was heat- and proteinase-K-labile and which probably constitutes the capsular polymer. The other component was a low-molecular-mass glycolipid fraction, which was proteinase-K-, heat- and EDTA-resistant. The identification of a capsule at the M. penetrans cell surface is of particular interest for a mycoplasma which has been shown to adhere to various host cells and to penetrate into their intracellular compartments. The capsule may have significance in the pathogenesis of disease associated with infection by this organism.
Environmentally controlled dimorphic cycle in a fission yeast
Authors: Sipiczki M. , Takeo K. , Yamaguchi M. , Yoshida S. , Miklos I.
Published: June 2009

The fission yeastSchizosaccharomyces pombeshows bipolar growth and is a convenient model for studying cell polarity and polar growth. This paper shows that the relatedSchiz. japonicusvar.japonicuscan switch to unipolar growth and can exist in both yeast and mycelial phases. On solid media, the yeast phase is unstable and prone to switch to the mycelial form, which shows unipolar growth by tip elongation. The hyphae can colonize the body of the substrate (true mycelium) or just its surface (pseudo-mycelium). The yeast-to-mycelium transition and the growth of the mycelium are regulated by a nutritional gradient and are associated with extensive vacuolation. The mycelium can convert into arthroconidia or return to the yeast phase in response to environmental changes. These environmentally controlled morphological transitions makeSchiz. japonicusvar.japonicusan attractive model for the investigation of cell polarity and morphogenesis.
Structural characteristics of halobacterial gas vesicles
Authors: Offner Sonja , Ziese Ulrike , Wanner Gerhard , Typke Dieter , Pfeifer Felicitas
Published: June 2009

Gas vesicle formation in halophilic archaea is encoded by a DNA region (the vac region) containing 14 different genes: gvpACNO and gvpDEFGHIJKLM. In Halobacterium salinarum PHH1 (which expresses the p-vac region from plasmid pHH1), gas vesicles are spindle shaped, whereas predominantly cylindrical gas vesicles are synthesized by the chromosomal c-vac region of H. salinarum PHH4 and the single chromosomal mc-vac region of Haloferax mediterranei. Homologous complementation of gvp gene clusters derived from the chromosomal c-vac region led to cylindrical gas vesicles in transformants and proved that the activity of the c-gvpA promoter depended on a gene product from the c-gvpE-M DNA region. Heterologous complementation experiments with transcription units of different vac regions demonstrated that the formation of chimeric gas vesicles was possible. Comparison of micrographs of wild-type and chimeric gas vesicles indicated that the shape was not exclusively determined by GvpA, the major structural protein of the gas vesicle wall. More likely, a dynamic equilibrium of several gvp gene products was responsible for determination of the shape. Transmission electron microscopy of frozen hydrated, wild-type gas vesicles showed moiré patterns due to the superposition of the front and back parts of the ribbed gas vesicle envelope. Comparison of projections of model helices with the moiré pattern seen on the cylindrical part of the gas vesicles provided evidence that the ribs formed a helix of low pitch and not a stack of hoops.
Taking a genetic scalpel to the Streptomyces colony
Authors: Chater Keith F.
Published: June 2009

1997 Fred Griffith Review Lecture

(Delivered at the 138th Meeting of the Society for General Microbiology, 2 September 1977)
Yersinia pseudotuberculosis and Yersinia pestis are more resistant to bactericidal cationic peptides than Yersinia enterocolitica
Authors: Bengoechea José-Antonio , Lindner Buko , Seydel Ulrich , Ramón Díaz , Ignacio Moriyón
Published: June 2009

The action of bactericidal polycationic peptides was compared in Yersinia spp. by testing peptide binding to live cells and changes in outer membrane (OM) morphology and permeability. Moreover, polycation interaction with LPS was studied by measuring the dependence of dansylcadaverine displacement and zeta potential on polycation concentration. When grown at 37 °C, Yersinia pestis and Yersinia pseudotuberculosis bound less polymyxin B (PMB) than pathogenic or non-pathogenic Yersinia enterocolitica, regardless of virulence plasmid expression. Y. pseudotuberculosis OMs were unharmed by PMB concentrations causing extensive OM blebbing in Y. enterocolitica. The permeability to Iysozyme caused by PMB was greater in Y. enterocolitica than in Y. pseudotuberculosis or Y. pestis and differences increased at 37 °C. Similar observations were made with other polycations using a polymyxin/novobiocin permeability assay. With LPS of cells grown at 26 °C, polycation binding was highest for Y. pseudotuberculosis and lowest for Y. pestis, with Y. enterocolitica yielding intermediate results which were lower for pathogenic than for non-pathogenic strains. With LPS of cells grown at 37 °C, polycation binding remained unchanged for Y. pastis and pathogenic Y. enterocolitica, increased for non-pathogenic Y. enterocolitica and decreased for Y. pseudotuberculosis to Y. pestis levels. Polycation binding related in part to differences in charge density (zeta potential) of LPS aggregates, suggesting similar effects at bacterial surfaces. It is suggested that species and temperature differences in polycation resistance relate to infection route, invasiveness and intracellular multiplication of Yersinia spp.
Phytase activity of anaerobic ruminal bacteria
Authors: Yanke L. J. , Bae H. D. , Selinger L. B. , Cheng K. J.
Published: June 2009

Phytase catalyses the release of phosphate from phytate (myo-inositol hexakisphosphate), the predominant form of phosphorus in cereal grains, oilseeds and legumes. The presence of phytase activity was investigated in 334 strains of 22 species of obligately anaerobic ruminal bacteria. Measurable activities were demonstrated in strains of Selenomonas ruminantium, Megasphaera elsdenii, Prevotella ruminicola, Mitsuokella multiacidus and Treponema spp. Strains isolated from fermentations with cereal grains proved to have high activity, and activity was particularly prevalent in S. ruminantium, with over 96% of the tested strains being positive. The measured phytase activity was found exclusively associated with the bacterial cells and was produced in the presence of approximately 14 mM phosphate. The most highly active strains were all S. ruminantium, with the exception of the one Mitsuokella multiacidus strain examined. Phytase activity varied greatly among positive strains but activities as high as 703 nmol phosphate released (ml culture)-1 were measured for a S. ruminantium strain and 387 nmol phosphate released (ml culture)-1 for the Mitsuokella multiacidus strain.
Characterization of a second cell-associated Arg-specific cysteine proteinase of Porphyromonas gingivals and identification of an adhesin-binding motif involved in association of the prtR and prtK proteinases and adhesins into large complexes
Authors: Slakeski Nada , Bhogal Peter S. , O’Brien-Simpson Neil M. , Reynolds Eric C.
Published: June 2009

Porphyromonas gingivalis has been associated with the development of adult periodontitis and cysteine proteinases with Arg- and Lys-specific activity have been implicated as major virulence factors. In a cell sonicate of P. gingivalis W50, a complex of non-covalently associated proteins has been previously characterized. This complex is composed of a 45 kDa Arg-specific, calcium-stabilized cysteine proteinase (PrtR45), a 48 kDa Lys-specific cysteine proteinase (PrtK48) and seven sequence-related adhesins designated PrtR44, PrtR15, PrtR17, PrtR27, PrtK39, PrtK15 and PrtK44, with all proteins being encoded by the two genes prtR and prtK. It has been proposed that these non-covalently associated complexes form extracellularly after autolytic processing of the PrtR and PrtK polyproteins, with the adhesins binding to the proteinases (PrtR45 and PrtK48) and autoaggregating. Another form of the cell-associated, Arg-specific, calcium-stabilized cysteine proteinase is described here. Designated PrtRII50, it is a discrete 50 kDa protein with no adhesin-association and has enzymic characteristics and an inhibitor/activator profile almost identical to PrtR45. The PrtRII50 proteinase is encoded as a preproprotein by a second gene, prtRII, with high sequence similarity to PrtR except that it lacks the C-terminal adhesin domains. A comparison of the deduced amino acid sequence of PrtRII50 with that of the adhesin-associated proteinases PrtR45 and PrtK48 revealed that PrtRII50 does not contain a C-terminal motif that is conserved in PrtR45 and PrtK48. Related motifs are also found in the adhesin domains of PrtR and PrtK. It is proposed that this conserved motif is an adhesin-binding motif (ABM) involved in association of the PrtR and PrtK proteinases and adhesins into large complexes, as the PrtR-PrtK proteinase-adhesin complex inactivated by N-α-p-tosyl-L-lysine chloromethyl ketone (TLCK) was shown to bind specifically to a synthetic peptide corresponding to the conserved motif in a competitive binding assay.
Reiterated repeat region variability in the ciliary adhesin gene of Mycoplasma hyopneumoniae
Authors: Wilton Jody L. , Scarman Anthony L. , Walker Mark J. , Djordjevic Steven P.
Published: June 2009

Mycoplasma hyopneumoniae is a highly prevalent pathogen which colonizes the ciliated epithelial lining of the porcine respiratory tract. Expression libraries constructed from genomic DNA of the non-pathogenic strain M. hyopneumoniae J were screened with porcine hyperimmune antiserum against M. hyopneumoniae. One clone expressed a 28 kDa protein which was also reactive with monospecific antiserum raised against a putative M. hyopneumoniae-specific 94 kDa antigen derived from strain J. Trypsin digestion of whole M. hyopneumoniae cells showed the 94 kDa antigen to be surface-accessible. DNA sequence analysis of the gene encoding the 94 kDa antigen revealed greater than 90% homology to two adhesin genes, encoding P97 and Mhp1, cloned from pathogenic strain 232 and strain P5722 of M. hyopneumoniae, respectively. Two regions of repetitive DNA sequence were identified in the gene encoding the 94 kDa antigen. The first encoded the deduced amino acid sequence A(T)-K-P-E(V)-A(T) arranged as nine tandem repeats (RR1). The second region of repetitive DNA sequence encoded the deduced amino acid sequence G-A(E,S)-P-N(S)-Q-G-K-K-A-E arranged as five tandem repeats (RR2). Comparison of the three M. hyopneumoniae adhesin genes revealed that the genes encoding P97 and Mhp1, and the strain J gene encoding the 94 kDa antigen contained 15, 12 and 9 tandem repeats, respectively, in RR1, and 4, 5 and 5 tandem repeats, respectively, in RR2. Southern hybridization analysis of EcoRI-digested genomic DNA probed with an 820 bp fragment spanning RR1 and RR2 identified a strongly hybridizing fragment ranging in size from 2.15 to 2.30 kb among seven geographically diverse strains of M. hyopneumoniae but failed to hybridize with DNA from four strains of Mycoplasma hyorhinis or Mycoplasma flocculare strain Ms42. PCR primers flanking the DNA sequence encoding RR1 and RR2 were used to amplify DNA from the seven strains of M. hyopneumoniae and DNA sequence analysis of the amplification products showed that the number of tandem amino acid repeats in RR1 varied considerably between strains. RR1 from M. hyopneumoniae strains YZ, Beaufort, Sue, OMZ407 and C1735/2 comprised 11, 15, 12, 15 and 8 tandem copies, respectively, of the 5-aa repeat whilst RR2 comprised 4, 3, 4, 3 and 4 tandem copies, respectively, of the 10-aa repeat. Two putative integrin binding sites (L-E-T and R-X-X-X-D) were identified in the 94 kDa ciliary adhesin. Variability in the number of amino acid repeats in RR1 amongst strains of M. hyopneumoniae may influence ciliary binding.
Rhizobium leguminosarum contains a group of genes that appear to code for methyl-accepting chemotaxis proteins
Authors: Yost Christopher K. , Rochepeau Patrice , Hynes Michael F.
Published: June 2009

Methyl-accepting chemotaxis proteins (MCPs) play important roles in the chemotactic response of many bacteria. Oligonucleotide primers designed to amplify the conserved signalling domain of MCPs by PCR were used to identify potential MCP-encoding genes in Rhizobium leguminosarum. Using a PCR-derived probe created from these primers a genomic library of R. leguminosarum VF39SM was screened; at least five putative MCP-encoding genes (termed mcpB to mcpF) were identified and isolated from the library. One of these putative genes (mcpC) is located on one of the indigenous plasmids of VF39SM. Fifteen different cosmids showing homology to an mcpD probe were also isolated from a genomic library. The complete DNA sequences of mcpB, mcpC and mcpD were obtained. All three genes code for proteins with characteristics typical of MCPs. However, the protein encoded by mcpB has a relatively large periplasmic domain compared to that in other MCPs. Partial DNA sequences of mcpE and mcpF had strong similarity to sequences from the methylation domains of known MCPs. Mutants defective in mcpB, mcpC, mcpD or mcpE were created using insertional mutagenesis strategies. Mutation of mcpB resulted in impairment of chemotaxis to a wide range of carbon sources on swarm plates; phenotypes for the other three mutants have yet to be elucidated. The mcpB, mcpC and mcpD mutants were tested for loss of nodulation competitiveness. When co-inoculated with the wild-type, the mcpB and mcpC mutants formed fewer nodules than the wild-type, whereas the mcpD mutant was just as competitive as the wild-type. The results overall suggest that R. leguminosarum possesses mcp-like genes, and that at least some of these play a role in early steps in the plant-microbe interaction.
Mycobacterial linear plasmids have an invertron-like structure related to other linear replicons in actinomycetes
Authors: Picardeau Mathieu , Vincent Véronique
Published: June 2009

The authors previously identified large plasmids in Mycobacterium xenopi, M. branderi and M. celatum which appeared to have a linear topology. This study has confirmed the presence of such linear plasmids in mycobacteria, including M. avium, and demonstrated that the ends of these replicons are covalently bound with protein(s), suggesting an invertron-like structure. The termini of one 25 kb plasmid, designated pCLP, from M. celatum were cloned and the first 500 bp of each terminus were sequenced. The termini of this plasmid show the characteristic features of invertrons with terminal inverted repeats of 45 bp (with imperfect matches) and several palindromic sequences. Moreover, similarity existed in the structure and terminal nucleotide sequence of pCLP and the termini of linear replicons of Streptomyces and Rhodococcus species, indicating a conservation of these linear extrachromosomal elements within the Actinomycetales.
High exogenous concentrations of phenoxyacetic acid are crucial for a high penicillin V productivity in Penicillium chrysogenum
Authors: Henriksen Claus M. , Nielsen Jens , Villadsen John
Published: June 2009

A high-penicillin-yielding strain of Penicillium chrysogenum was grown in continuous culture on a chemically defined medium with glucose as the growth-limiting component. The cultivations were operated at a constant dilution rate of 0.05 h-1and the feed concentration of the penicillin V sidechain precursor phenoxyacetic acid was varied between 0 and 6.5 g l-1. Subsequent formation of penicillin V and by-products related to the penicillin biosynthetic pathway was monitored at steady state. It was established that the concentration of phenoxyacetic acid in the growth medium had to be kept high to obtain a high productivity of penicillin V. The specific production rate of penicillin V as a function of the phenoxyacetic acid concentration followed Michaelis--Menten-type kinetics, from which an overall apparent Kmvalue of 42 mM for the incorporation of intracellular phenoxyacetic acid into penicillin V could be obtained. High phenoxyacetic acid concentrations tended to lower the formation of the by-products 6-aminopenicillanic acid and 8-hydroxypenillic acid. Furthermore the undesirable loss of the pathway intermediate isopenicillin N into the extracellular medium was lowered, whereas the opposite effect was observed for the pathway intermediate δ-(L-α-aminoadipyl)-L-cysteinyl-D-valine and the by-product 6-oxo-piperidine-2-carboxylic acid, the δ-lactam form of α-aminoadipic acid.