Biomedical subjects

Joachim Reidl

Publications and source records attributed to Joachim Reidl.

8 recordsLinked to original sources

PubMed2003

Porin OmpP2 of Haemophilus influenzae shows specificity for nicotinamide-derived nucleotide substrates.

Haemophilus influenzae has an absolute requirement for NAD (factor V) because it lacks all biosynthetic enzymes necessary for de novo synthesis of that cofactor. Therefore, growth in vitro requires the presence of NAD itself, NMN, or nicotinamide riboside (NR). To address uptake abilities of these compounds, we investigated outer membrane proteins. By analyzing ompP2 knockout mutants, we found that NAD and NMN uptake was prevented, whereas NR uptake was not. Through investigation of the properties of purified OmpP2 in artificial lipid membrane systems, the substrate specificity of OmpP2 for NAD and NMN was determined, with KS values of approximately 8 and 4mm, respectively, in 0.1 m KCl, whereas no interaction was detected for the nucleoside NR and other purine or pyrimidine nucleotide or nucleoside species. Based on our analysis, we assume that an intrinsic binding site within OmpP2 exists that facilitates diffusion of these compounds across the outer membrane, recognizing carbonyl and exposed phosphate groups. Because OmpP2 was formerly described as a general diffusion porin, an additional property of acting as a facilitator for nicotinamide-based nucleotide transport may have evolved to support and optimize utilization of the essential cofactor sources NAD and NMN in H. influenzae.

Bacterial Outer Membrane Proteins↗

PubMed2003

Aerobic growth deficient Haemophilus influenzae mutants are non-virulent: implications on metabolism.

We investigated aerobic metabolism in Haemophilus influenzae to better understand its essential physiological growth pathways. We describe the isolation and characterization of transposon insertions leading to knockout mutations in lpdA, encoding dihydrolipoamide dehydrogenase. H. influenzae Rd lpdA::Tn10d-cat mutants were unable to grow aerobically and an H. influenzae type b lpdA::Tn10d-cat mutant was significantly attenuated in an infant rat infection model. Since LpdA is a functional subunit of both pyruvate dehydrogenase (aceEF) and alpha-ketoglutarate dehydrogenase (sucAB) the phenotype of the lpdA mutant was further explored by creating separate knockout mutants in the sucAB and aceEF loci. DeltaaceEF and deltasucAB mutants were both significantly attenuated in virulence in the infant rat, but only the sucAB mutant was able to grow aerobically. We therefore conclude that the ability for aerobic growth is critical for invasive disease, and furthermore that a TCA cycle enzyme, alpha-ketoglutarate dehydrogenase, appears to contribute a key metabolic function in vivo, but is not required for growth under laboratory conditions.

Aerobiosis↗

PubMed2003

Nicotinamide ribosyl uptake mutants in Haemophilus influenzae.

The gene for the nicotinamide riboside (NR) transporter (pnuC) was identified in Haemophilus influenzae. A pnuC mutant had only residual NR uptake and could survive in vitro with high concentrations of NR, but could not survive in vivo. PnuC may represent a target for the development of inhibitors for preventing H. influenzae disease.

Bacterial Proteins↗

PubMed2003

Transposon Tn10.

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DNA Transposable Elements↗

PubMed2002

Vibrio cholerae and cholera: out of the water and into the host.

The facultative human pathogen Vibrio cholerae can be isolated from estuarine and aquatic environments. V. cholerae is well recognized and extensively studied as the causative agent of the human intestinal disease cholera. In former centuries cholera was a permanent threat even to the highly developed populations of Europe, North America, and the northern part of Asia. Today, cholera still remains a burden mainly for underdeveloped countries, which cannot afford to establish or to maintain necessary hygienic and medical facilities. Especially in these environments, cholera is responsible for significant mortality and economic damage. During the last three decades, intensive research has been undertaken to unravel the virulence properties and to study the epidemiology of this significant human pathogen. More recently, researchers have been elucidating the environmental lifestyle of V. cholerae. This review provides an overview of the current knowledge of both the host- and environment-specific physiological attributes of V. cholerae.

Cholera↗

PubMed2002

Role of Vibrio cholerae O139 surface polysaccharides in intestinal colonization.

Since the first occurrence of O139 Vibrio cholerae as a cause of cholera epidemics, this serogroup has been investigated intensively, and it has been found that its pathogenicity is comparable to that of O1 El Tor strains. O139 isolates express a thin capsule, composed of a polymer of repeating units structurally identical to the lipopolysaccharide (LPS) O side chain. In this study, we investigated the role of LPS O side chain and capsular polysaccharide (CPS) in intestinal colonization by with genetically engineered mutants. We constructed CPS-negative, CPS/LPS O side chain-negative, and CPS-positive/LPS O side chain-negative mutants. Furthermore, we constructed two mutants with defects in LPS core oligosaccharide (OS) assembly. Loss of LPS O side chain or CPS resulted in a approximately 30-fold reduction in colonization of the infant mouse small intestine, indicating that the presence of both LPS O side chain and CPS is important during the colonization process. The strain lacking both CPS and LPS O side chain and a CPS-positive, LPS O side chain-negative core OS mutant were both essentially unable to colonize. To characterize the role of surface polysaccharides in survival in the host intestine, resistance to several antimicrobial substances was investigated in vitro. These investigations revealed that the presence of CPS protects the cell against attack of the complement system and that an intact core OS is necessary for survival in the presence of bile.

Animals↗

PubMed2002

Comparative and genetic analyses of the putative Vibrio cholerae lipopolysaccharide core oligosaccharide biosynthesis (wav) gene cluster.

We identified five different putative wav gene cluster types, which are responsible for the synthesis of the core oligosaccharide (OS) region of Vibrio cholerae lipopolysaccharide. Preliminary evidence that the genes encoded by this cluster are involved in core OS biosynthesis came from analysis of the recently released O1 El Tor V. cholerae genome sequence and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of O1 El Tor mutant strains defective in three genes (waaF, waaL, and wavB). Investigations of 38 different V. cholerae strains by Southern blotting, PCR, and sequencing analyses showed that the O1 El Tor wav gene cluster type is prevalent among clinical isolates of different serogroups associated with cholera and environmental O1 strains. In contrast, we found differences in the wav gene contents of 19 unrelated non-O1, non-O139 environmental and human isolates not associated with cholera. These strains contained four new wav gene cluster types that differ from each other in distinct gene loci, providing evidence for horizontal transfer of wav genes and for limited structural diversity of the core OS among V. cholerae isolates. Our results show genetic diversity in the core OS biosynthesis gene cluster and predominance of the type 1 wav gene locus in strains associated with clinical cholera, suggesting that a specific core OS structure could contribute to V. cholerae virulence.

Amino Acid Sequence↗

PubMed2002

Vibrio cholerae phage K139: complete genome sequence and comparative genomics of related phages.

In this report, we characterize the complete genome sequence of the temperate phage K139, which morphologically belongs to the Myoviridae phage family (P2 and 186). The prophage genome consists of 33,106 bp, and the overall GC content is 48.9%. Forty-four open reading frames were identified. Homology analysis and motif search were used to assign possible functions for the genes, revealing a close relationship to P2-like phages. By Southern blot screening of a Vibrio cholerae strain collection, two highly K139-related phage sequences were detected in non-O1, non-O139 strains. Combinatorial PCR analysis revealed almost identical genome organizations. One region of variable gene content was identified and sequenced. Additionally, the tail fiber genes were analyzed, leading to the identification of putative host-specific sequence variations. Furthermore, a K139-encoded Dam methyltransferase was characterized.

Bacteriophages↗