ALBERT

Notes: . In vitro excystation of Spironucleus muris cysts, purified by sequential sucrose and Percoll gradients from mouse feces, was studied. Three in vitro excystation procedures, used for Giardia, were assessed to determine the most useful method. Excystation was monitored by light microscopy and subsequently characterized by transmission and scanning electron microscopy. Spironucleus muris excysted routinely at a level greater than 90% when induced in Hanks’ balanced salt solution containing sodium bicarbonate at pH 2.0 and transferred to Tyrodes’ salt solution as an excystation medium. Similarly, high rates of excystation were recorded after induction of S. muris cysts in 0.1 M potassium phosphate buffer (pH 7.0) with sodium bicarbonate and excystation in trypticase-yeast extract-iron medium (TYI medium) or phosphate-buffered saline. A lower rate and percentage of excystation were observed after induction of S. muris cysts in an aqueous hydrochloric acid solution (pH 2.0) followed by excystation in TYI medium. All excystation methods produced extremely active S. muris trophozoites with normal morphology. Nonexcysting S. muris cysts have a wall composed of an outer fibrous and an inner membranous portion. Following induction, numerous vesicles appeared in the peritrophic space. Excystation began by the cyst wall opening at one pole, and the anterior part of the trophozoite protruding from the cyst wall. The trophozoite emerged progressively from the cyst wall and the empty cyst wall appeared to collapse. Excysted trophozoites exhibited normal morphological features of S. muris trophozoites isolated from the mouse intestine.

Notes: Abstract The list of carboxydotrophic bacteria is constantly growing and we have found that burning charcoal piles harbor an especially rich CO-oxidizing microflora. The newly isolated Streptomyces thermoautotrophicus UBT1 is particularly interesting as it is thermophilic, capable of chemolithoautotrophic growth with CO or H2 plus CO2 and incapable of using organic substrates. Molybdenum is essential for CO-autotrophic growth.Some species of carboxydotrophic bacteria can denitrify under heterotrophic conditions yielding N2 (e.g. Pseudomonas carboxydoflava) or N2O (e.g. Pseudomonas carboxydohydrogena); others perform nitrate respiration (e.g. Azomonas B1). P. carboxydohydrogena could grow at the expense of H2 plus CO2 using nitrate as electron acceptor.In intact cells of Pseudomonas carboxydovorans, CO dehydrogenase has the ability of dissociating from and rebinding to the cytoplasmic membrane. That process can be simulated in vitro by removing CO dehydrogenase from cytoplasmic membranes and rebinding it to depleted membranes. Reconstitution of the enzyme onto depleted membranes requiring di- or trivalent cations, was specific for membranes from CO-grown bacteria and led to reactivation of respiratory activities with CO.A complex consisting of 1 molecule of CO dehydrogenase and 2 molecules of cytochrome b561 could be isolated from cytoplasmic membranes of P. carboxydovorans solubilized with dodecyl β-d-maltoside. Within the complex as well as in assays containing purified CO dehydrogenase and cytochrome b561 the latter could serve as an electron acceptor. CO dehydrogenase had hydrogenase activity, and its KM of only 5 mM H2 suggested a role in the formation of H2. P. carboxydovorans OM5 contains the 128-kilo-base pairs (kb) plasmid pHCG3 which is essential for CO- and H2-lithoautotrophic growth. Evidence for the existence of pHCG3-coded structural genes of CO dehydrogenase was obtained from dot blot hybridizations employing synthetic oligodeoxynucleotides as heterologous probes for the detection of the S- and M-subunit genes. Employing appropriate probe genes encoding membrane-bound hydrogenase, ribulose biphosphate carboxylase and phosphoribulokinase were also identified on plasmid pHCG3.

Notes: Abstract The bioenergetic properties of the thermoacidophilic archaebacterium Sulfolobus acidocaldarius are reviewed and discussed under the aspect whether this archaebacterium conserved energy by oxidative phosphorylation and how the involved catalysts are related to those from eubacteria and eukaryotes. The thermodynamic parameters contributing to the proton-motive force and the efficiency of proton pumping are presented. The major components of the electron transport system are identified and a novel type of heme-aa3 containing terminal oxidase is described, oxidizing reduced caldariella quinone. The properties of an F1-analogous ATPase and of a DCCD-binding proteolipid from the plasmamembrane of Sulfolobus are discussed as likely components of an F0F1-analogous ATP-synthase. The structural and functional properties of this and other archaebacterial ATPases are compared to each other and with respect to evolutionary relations.

Notes: Abstract The pathogenic Neisseria have exploited the processes of horizontal DNA transfer and genetic recombination as mechanisms for the generation of extensive protein variation and modulation of gene expression. Localized recombinations have been well documented in members of multigene families as have alterations in short repetitive sequences. Here we report an analysis of the chromosomal structure of a defined lineage of Neisseria gonorrhoeae strain MS 11 pilin variants. This study reveals the occurrence of large rearrangements, including the amplification of a 26 kb region and an inversion involving more than a third of the chromosome. Additionally, a restriction site polymorphism that correlates with pilin expression has been observed. These findings highlight the flexibility of the gonococcal genome.

Notes: Abstract We previously identified and genetically characterized several factors essential for the natural competence of transformation in Neisseria gonorrhoeae. Here we analyse the sequential action of these factors and dissect the overall transformation process into three distinct steps, (i) the sequence-specific uptake of transforming DNA into a DNase-resistant state, (ii) the transfer of DNA to the cytosol and (iii) the processing and recombination of the incoming with the resident DNA. While two pilus-associated factors, PilE and PilC, were previously implicated in the early DNA uptake event, we show here that three competence factors unrelated to pilus biogenesis, ComA, ComL and Tpc, are not essential for DNA uptake and rather act in a subsequent step. The respective mutants, however, lack the characteristic nucleolytic processing observed with the incoming DNA in both wild-type and non-transformable RecA-deficient N. gonorrhoeae, indicating that they are blocked in the processing and/or the delivery of DNA to the cytoplasm. A hypothetical model proposing a sequential action of the known gonococcal competence factors is presented.