Voen Med Zh, 1999 Oct, 320(10), 45 - 8, 96 {The methods of traditional medicine in the treatment of tuberculosis}; Petrosian FK et al.; The authors sum up the results of phyto-, aromatherapy and marmot fat complex treatment of 64 patients with pulmonary tuberculosis . In a two-month treatment course in 81.8-95.6% of patients the symptoms of tuberculosis intoxication had disappeared, the body mass increased and the bacteria discharge stopped . It proves that traditional medicine means and methods can be recommended for the rehabilitation periods of tuberculosis patients. Mol Biol Evol, 1999 Dec, 16(12), 1696 - 710 Using protein structural information in evolutionary inference: transmembrane proteins; Lio P et al.; We present a model of amino acid sequence evolution based on a hidden Markov model that extends to transmembrane proteins previous methods that incorporate protein structural information into phylogenetics . Our model aims to give a better understanding of processes of molecular evolution and to extract structural information from multiple alignments of transmembrane sequences and use such information to improve phylogenetic analyses . This should be of value in phylogenetic studies of transmembrane proteins: for example, mitochondrial proteins have acquired a special importance in phylogenetics and are mostly transmembrane proteins . The improvement in fit to example data sets of our new model relative to less complex models of amino acid sequence evolution is statistically tested . To further illustrate the potential utility of our method, phylogeny estimation is performed on primate CCR5 receptor sequences, sequences of l and m subunits of the light reaction center in purple bacteria, guinea pig sequences with respect to lagomorph and rodent sequences of calcitonin receptor and K-substance receptor, and cetacean sequences of cytochrome b. Z Gastroenterol, 1999 Nov, 37(11), 1105 - 8 Lymphocytic colitis, induced by ticlopidine; Feurle GE et al.; Lymphocytic colitis is a chronic inflammatory colonic disease characterized by watery diarrhea and a dense infiltration of the colonic mucosa with lymphocytes . The etiology is unknown but an immune reaction to various immunostimulatory agents including pathogenic or commensal bacteria, products of bacterial metabolism of dietary degradation, or antigens derived directly from the diet, and autoimmune phenomena are discussed . We observed a patient with all features of lymphocytic colitis characterized by a prominent intraepithelial T-cell component . The colitis resolved completely when therapy with ticlopidine--an agent inhibiting platelet aggregation--was stopped . This observation suggests that medical history concerning drug ingestion may reveal the etiology of lymphocytic colitis and allows cure of this otherwise difficult to treat disorder. Infect Immun, 2000 Jan, 68(1), 387 - 90 Evaluation of Mycobacterium tuberculosis genes involved in resistance to killing by human macrophages; Miller BH et al.; A coinfection assay was developed to examine Mycobacterium tuberculosis genes suspected to be involved in resistance to killing by human macrophages . THP-1 macrophages were infected with a mixture of equal numbers of recombinant Mycobacterium smegmatis LR222 bacteria expressing an M . tuberculosis gene and wild-type M . smegmatis LR222 bacteria expressing the xylE gene . At various times after infection, the infected macrophages were lysed and the bacteria were plated . The resulting colonies were sprayed with catechol to determine the number of recombinant colonies and the number of xylE-expressing colonies . M . smegmatis bacteria expressing the M . tuberculosis glutamine synthetase A (glnA) gene or open reading frame Rv2962c or Rv2958c demonstrated significantly increased survival rates in THP-1 macrophages relative to those of xylE-expressing bacteria . M . smegmatis bacteria expressing M . tuberculosis genes for phospholipase C (plcA and plcB) or for high temperature requirement A (htrA) did not. Infect Immun, 2000 Jan, 68(1), 360 - 7 Isolates of Chlamydia trachomatis that occupy nonfusogenic inclusions lack IncA, a protein localized to the inclusion membrane; Suchland RJ et al.; The chlamydiae are obligate intracellular pathogens that occupy a nonacidified vacuole, termed an inclusion, throughout their developmenal cycle . When an epithelial cell is infected with multiple Chlamydia trachomatis elementary bodies, they are internalized by endocytosis into individual phagosomal vacuoles that eventually fuse to form a single inclusion . In the course of large-scale serotyping studies in which fluorescent antibody staining of infected cells was used, a minority of strains that had an alternate inclusion morphology were identified . These variants formed multiple nonfusogenic inclusions in infected cells, with the number of independent inclusions per cell varying directly with the multiplicity of infection . Overall the nonfusogenic phenotype was found in 1.5% (176 of 11,440) of independent isolates . Nonfusing variants were seen in C . trachomatis serovars B, D, D-, E, F, G, H, Ia, J, and K . The nonfusing phenotype persisted through repeated serial passage, and the phenotype was consistent in four mammalian host cell lines . Fluorescence microscopy and immunoblotting with antisera directed at proteins in the C . trachomatis inclusion membrane revealed that one such protein, IncA, was not detected in the inclusion membrane in each tested nonfusogenic strain . The distributions of other chlamydial proteins, including one additional Inc protein, were similar in wild-type and variant strains . The incA coding and upstream regions were amplified and sequenced from the prototype serovar D and two nonfusing serovar D((s)) strains . Three nucleotide changes were discovered in the D((s)) incA gene, leading to two amino acid changes within the predicted D((s)) IncA sequence . These studies demonstrate a subgroup of variant C . trachomatis isolates that form nonfusing inclusions; the variant phenotype is associated with the absence of detectable IncA and with an altered incA sequence that modifies the characteristic hydrophobic domain of the IncA protein. Biochem Biophys Res Commun, 1999 Dec 29, 266(3), 690 - 8 Forty years of cytochrome P450; Omura T; The term "cytochrome P450" first appeared in literature in 1962 . It was a microsomal membrane-bound hemoprotein without known physiological functions at that time and was characterized by a unique 450-nm optical absorption peak of its carbon monoxide-bound form, which was originally reported as the spectrum of a novel "microsomal carbon monoxide-binding pigment" in 1958 . Elucidation of its function as the oxygenase in 1963 triggered a rapid expansion of research on this hemoprotein . Annual numbers of the published papers dealing with cytochrome P450, which were listed in Biological Abstracts, increased from 60 in 1970 to 500 in 1980, 900 in 1990, and 1500 in 1997 . Cytochrome P450 is now regarded as the collective name of a large family of hemoproteins, "cytochrome P450 superfamily, "which seems to have diversified from a single ancestral protein to many forms during the course of biological evolution and is distributed widely among various forms of life from animals and plants to fungi and bacteria . Multicellular eukaryotic organisms including animals and plants have about 100 or more P450 genes in their genomes, and those many P450 genes are expressed tissue specifically and developmental stage specifically, indicating their diverse physiological functions . In mammals, various P450s participate in the biosynthesis and metabolism of sterols and steroid hormones and the metabolism of various lipid biofactors including eicosanoids, vitamin D3, and retinoids . Oxidative metabolism of foreign hydrophobic compounds as the first step of their excretion from the animal body is apparently another major function of cytochrome P450, which protects animals from noxious foreign compounds, man-created and natural . Eur J Immunol, 1999 Dec, 29(12), 4037 - 42 Dendritic cells up-regulate immunoproteasomes and the proteasome regulator PA28 during maturation; Macagno A et al.; Dendritic cells (DC) are highly specialized professional antigen presenting cells which are pivotal for the initiation and control of the cytotoxic T cell response . Upon stimulation by cytokines, bacteria, or CD40L DC undergo a maturation process from an antigen-receptive state to a state of optimal stimulation of T cells . We investigated the composition of proteasomes of DC derived from human peripheral blood monocytes before and after stimulation by CD40L, LPS, or proinflammatory cytokines (TNF-alpha + IL-6 + IL-1beta) . Immunoprecipitation of proteasomes and analysis on two-dimensional gels revealed that during maturation the inducible proteasome subunits LMP2, LMP7, and MECL-1 are up-regulated and that the neosynthesis of proteasomes is switched exclusively to the production of immunoproteasomes containing these subunits . The proteasome regulator PA28 is markedly up-regulated in mature DC and in addition a so - far unidentified 21-kDa protein co-precipitates with the proteasome in LPS - stimulated DC . These changes in proteasome composition may be functionally linked to special properties of DC like MHC class I up-regulation or cross-priming . Our findings imply that the spectrum of class I-bound peptides may change after DC maturation which could be relevant for the design of DC - based vaccines. Caries Res, 2000 Jan-Feb, 34(1), 88 - 93 The effects of milk and kappa-casein on salivary pellicle formed on hydroxyapatite discs in situ; Vacca Smith AM et al.; The effects of milk and kappa-casein rinses on the salivary pellicle formed on hydroxyapatite discs carried in the mouth were studied . SDS-PAGE analyses revealed an increase in the number of proteins deposited onto the discs carried after the water and milk rinses only . Scanning electron microscopy studies revealed the deposition of an amorphous material, small, micelle-like structures, cocci and rods on discs carried in the mouth after the water rinse . Large, micelle-like structures were seen on discs carried in the mouth after the milk and kappa-casein rinses; bacteria were not seen . Glucosyltransferase (Gtf) activity on discs carried in the mouth after the milk and kappa-casein rinses were 45+/-5 and 67+/-2% lower than the activity of Gtf on discs carried in the mouth after a water rinse, respectively . These data suggest that milk and kappa-casein may influence pellicle formation in vivo. J Biol Chem, 1999 Dec 24, 274(52), 36866 - 75 Identification and functional characterization of a novel, tissue-specific NAD(+)-dependent isocitrate dehydrogenase beta subunit isoform; Kim YO et al.; To understand the interactions and functional role of each of the three mitochondrial NAD(+)-dependent isocitrate dehydrogenase (IDH) subunits (alpha, beta, and gamma), we have characterized human cDNAs encoding two beta isoforms (beta(1) and beta(2)) and the gamma subunit . Analysis of deduced amino acid sequences revealed that beta(1) and beta(2) encode 349 and 354 amino acids, respectively, and the two isoforms only differ in the most carboxyl 28 amino acids . The gamma cDNA encodes 354 amino acids and is almost identical to monkey IDHgamma . Northern analyses revealed that the smaller beta(2) transcript (1.3 kilobases) is primarily expressed in heart and skeletal muscle, whereas the larger beta(1) mRNA (1.6 kilobases) is prevalent in nonmuscle tissues . Sequence analysis of the IDHbeta gene indicates that the difference in the C-terminal 28 amino acids between beta(1) and beta(2) proteins results from alternative splicing of a single transcript . Among the various combinations of human IDH subunits co-expressed in bacteria, alphabetagamma, alphabeta, and alphagamma combinations exhibited significant amounts of IDH activity, whereas subunits produced alone and betagamma showed no detectable activity . These data suggest that the alpha is the catalytic subunit and that at least one of the other two subunits plays an essential supporting role for activity . Substitution of beta(1) with beta(2) in the co-expression system lowered the pH optimum for IDH activity from 8.0 to 7.6 . This difference in optimal pH was analogous to what was observed in mouse kidney and brain (beta(1) prevalent; optimal pH 8.0) versus heart (beta(2) prevalent; pH 7.6) mitochondria . Experiments with a specially designed splicing reporter construct stably transfected into HT1080 cells indicate that acidic conditions favor a splicing pattern responsible for the muscle- and heart-specific beta(2) isoform . Taken together, these data indicate a regulatory role of IDHbeta isoforms in determining the pH optimum for IDH activity through the tissue-specific alternative splicing. J Bacteriol, 1999 Dec, 181(24), 7647 - 9 The periplasmic 9.6-kilodalton c-type cytochrome of Geobacter sulfurreducens is not an electron shuttle to Fe(III); Lloyd JR et al.; Geobacter sulfurreducens contains a 9.6-kDa c-type cytochrome that was previously proposed to serve as an extracellular electron shuttle to insoluble Fe(III) oxides . However, when the cytochrome was added to washed-cell suspensions of G . sulfurreducens it did not enhance Fe(III) oxide reduction, whereas similar concentrations of the known electron shuttle, anthraquinone-2,6-disulfonate, greatly stimulated Fe(III) oxide reduction . Furthermore, analysis of the extracellular c-type cytochromes in cultures of G . sulfurreducens demonstrated that the dominant c-type cytochrome was not the 9.6-kDa cytochrome, but rather a 41-kDa cytochrome . These results and other considerations suggest that the 9.6-kDa cytochrome is not an important extracellular electron shuttle to Fe(III) oxides. J Bacteriol, 1999 Dec, 181(24), 7629 - 33 Conditional sigma factor expression, using the inducible acetamidase promoter, reveals that the Mycobacterium tuberculosis sigF gene modulates expression of the 16-kilodalton alpha-crystallin homologue; Manabe YC et al.; A chemically inducible acetamidase promoter-sigF fusion gene was integrated into the chromosome of Mycobacterium bovis BCG . Two-dimensional protein gel analysis permitted the identification of a number of protein spots whose expression was SigF related . One spot upregulated by inappropriate induction of sigF expression corresponded to the 16-kDa antigen alpha-crystallin. Biochem Biophys Res Commun, 1999 Dec 20, 266(2), 366 - 70 Lysozyme association with nucleic acids; Steinrauf LK et al.; Lysozyme is well known for the ability to hydrolyze the cell wall of bacteria . Based on the similarity of structure between lysozyme and histones as seen from the results of X-ray crystal structure determinations, we have postulated that binding to nucleic acids may be another biological function of lysozyme . We have therefore begun a systematic study of the interactions of lysozyme and related molecules with nucleic acids, and present here a preliminary report . Binding to DNA and RNA has been demonstrated from gel electrophoresis, enzyme activity, and coprecipitation studies . We suggest that this function of lysozyme will provide an explanation why Lee-Huang et al . (1999) {Proc . Natl . Acad . Sci . USA 96, 2678-2681} were able to call lysozyme a "killer protein" against the AIDS virus, and may provide a new avenue of research on AIDS therapy . Biochemistry, 1999 Dec 14, 38(50), 16424 - 31 Hydrodynamic radii of native and denatured proteins measured by pulse field gradient NMR techniques; Wilkins DK et al.; Pulse field gradient NMR methods have been used to determine the effective hydrodynamic radii of a range of native and nonnative protein conformations . From these experimental data, empirical relationships between the measured hydrodynamic radius (R(h)) and the number of residues in the polypeptide chain (N) have been established; for native folded proteins R(h) = 4.75N (0.29)A and for highly denatured states R(h) = 2.21N (0.57)A . Predictions from these equations agree well with experimental data from dynamic light scattering and small-angle X-ray or neutron scattering studies reported in the literature for proteins ranging in size from 58 to 760 amino acid residues . The predicted values of the hydrodynamic radii provide a framework that can be used to analyze the conformational properties of a range of nonnative states of proteins . Several examples are given here to illustrate this approach including data for partially structured molten globule states and for proteins that are unfolded but biologically active under physiological conditions . These reveal evidence for significant coupling between local and global features of the conformational ensembles adopted in such states . In particular, the effective dimensions of the polypeptide chain are found to depend significantly on the level of persistence of regions of secondary structure or features such as hydrophobic clusters within a conformational ensemble. Parasitol Res, 1999 Dec, 85(12), 974 - 6 Scanning electron microscopy of the surface coat of Blastocystis hominis; Zaman V et al.; Scanning electron microscopy of Blastocystis hominis showed that its outer coat has a fibrillar structure and individual fibrils may extend up to 5 microm from the periphery of the parasite . The surface coat remains intact during cell division . Bacteria are often seen adhering to it, but for the first time a trophozoite of Chilomastix mesnili was also seen in this position . It is postulated that breakdown of attached organisms may provide nutrients for Blastocystis. Dis Aquat Organ, 1999 Nov 8, 38(2), 115 - 23 Differential expression of the virulence-associated protein p57 and characterization of its duplicated gene msa in virulent and attenuated strains of Renibacterium salmoninarum; O'Farrell CL et al.; Virulence mechanisms utilized by the salmonid fish pathogen Renibacterium salmoninarum are poorly understood . One potential virulence factor is p57 (also designated MSA for major soluble antigen), an abundant 57 kDa soluble protein that is predominately localized on the bacterial cell surface with significant levels released into the extracellular milieu . Previous studies of an attenuated strain, MT 239, indicated that it differs from virulent strains in the amount of surface-associated p57 . In this report, we show overall expression of p57 in R . salmoninarum MT 239 is considerably reduced as compared to a virulent strain, ATCC 33209 . The amount of cell-associated p57 is decreased while the level of p57 in the culture supernatant is nearly equivalent between the strains . To determine if the lowered amount of cell-associated p57 was due to a sequence defect in p57, a genetic comparison was performed . Two copies of the gene encoding p57 (msa1 and msa2) were found in 33209 and MT 239, as well as in several other virulent isolates . Both copies from 33209 and MT 239 were cloned and sequenced and found to be identical to each other, and identical between the 2 strains . A comparison of msa1 and msa2 within each strain showed that their sequences diverge 40 base pairs 5' to the open reading frame, while sequences 3' to the open reading frame are essentially identical for at least 225 base pairs . Northern blot analysis showed no difference in steady state levels of msa mRNA between the 2 strains . These data suggest that while cell-surface localization of p57 may be important for R . salmoninarum virulence, the differences in localization and total p57 expression between 33209 and MT 239 are not due to differences in msa sequence or differences in steady state transcript levels. N M Dent J, 1998 Jul, 49(3), 12 - 3 Why do I need a root canal? Goodis C. Bacteria invading the pulp cause pulpal necrosis and the need for endodontic treatment . If bacteria are present after treatment, the root canal may fail . For the root canal to be successful in necrotic cases or retreating failing cases, the bacteria in the root canal walls must be removed and the canal sterilized with calcium hydroxide. Toxicol Lett, 1999 Nov 22, 110(3), 145 - 75 Methylglyoxal in living organisms: chemistry, biochemistry, toxicology and biological implications; Kalapos MP; Despite the growing interest towards methylglyoxal and glyoxalases their real role in metabolic network is still obscure . In the light of developments several reviews have been published in this field mainly dealing with only a narrow segment of this research area . In this article a trial is made to present a comprehensive overview of methylglyoxal research, extending discussion from chemistry to biological implications by reviewing some important characteristics of methylglyoxal metabolism and toxicity in a wide variety of species, and emphasizing the action of methylglyoxal on energy production, free radical generation and cell killing . Special attention is paid to the discussion of alpha-oxoaldehyde production in the environment as a potential risk factor and to the possible role of this a-dicarbonyl in diseases . Concerning the interaction of methylglyoxal with biological macromolecules (DNA, RNA, proteins) an earlier review (Kalapos, Toxicology Letters, 73, 1994, 3-24) means a supplementation to this paper, thus hoping the avoidance of unnecessary bombast . The paper arrives at the conclusion that since the early stage of evolution the function of methylglyoxalase pathway has been related to carbohydrate metabolism, but its significance has been changed over the thousands of years . Namely, at the beginning of evolution methylglyoxalase path was essential for the reductive citric acid cycle as an anaplerotic route, while in the extant metabolism it concerns with the detoxification of methylglyoxal and plays some regulatory role in triose-phosphate household . As there is a tight junction between methylglyoxal and carbohydrate metabolism its pathological role in the events of the development of diabetic complications emerges in a natural manner and further progress is hoped in this field . In contrast, significant advancement cannot be expected in relation to cancer research. Arch Med Res, 1999 Sep-Oct, 30(5), 360 - 7 Mutation of highly conserved arginine residues disrupts the structure and function of annexin V; Campos B et al.; BACKGROUND: Annexins are a family of structurally related proteins that bind to phospholipid membranes in a Ca(2+)-dependent manner . Annexins are characterized by highly conserved canonical domains of approximately 70 amino acids . Annexin V contains four such domains . Each of these domains has a highly conserved arginine (R) . METHODS: To evaluate the role of the conserved arginines in the molecular structure of annexin V, negatively charged amino acids were substituted for arginines at positions R43, R115, R199, and R274 using site-directed mutagenesis . RESULTS: Mutants R199D and R274E were rapidly degraded when expressed in bacteria, and were not further characterized . R43E exhibited an electrophoretic mobility similar to the wild-type protein, while R115E migrated significantly in a slower fashion, suggesting a less compact conformation . R43E and R115E exhibited much greater susceptibility to proteolytic digestion than the wild type . While Ca(2+)-dependence for phospholipid binding was similar in both mutants (half-maximal 50-80 microM Ca2+), R43E and R115E exhibited a 6- and 2-fold decrease in phospholipid affinity, respectively . Consistent with the different phospholipid affinities of the annexins, a phospholipid-dependent clotting reaction, the activated partial thromboplastin time (aPTT), was significantly prolonged by the wild-type protein and mutants R115E and R115A . The aPTT was unaffected by R43E . CONCLUSIONS: Our data suggest that mutation of these highly conserved arginine residues in each of the four canonical domains of annexin have differential effects on the phospholipid binding, tertiary structure, and proteolytic susceptibility of annexin V . The site I mutation, R43E, produced a large decrease in phospholipid affinity associated with an increase in proteolytic susceptibility . The site II mutation, R115E, produced a small change in phospholipid binding but a significant modification of electrophoretic mobility . Our data suggest that highly conserved arginine residues are required to stabilize the tertiary structure of annexin V by establishing hydrogen bonds and ionic bridges. Ther Umsch, 1999 Nov, 56(11), 631 - 9 {Meningitis (I)--differential diagnosis; aseptic and chronic meningitis}; Leib SL et al.; Meningitis is the most common serious manifestation of infection of the central nervous system . Inflammatory involvement of the subarachnoid space with meningeal irritation leads to the classical triad of headache, fever, and meningism, and to a pleocytosis of the cerebrospinal fluid (CSF) . Meningitis is clinically categorized into an acute and chronic disease based on the acuity of symptoms . Acute meningitis develops over hours to days, while in chronic meningitis symptoms evolve over days or even weeks . Aseptic meningitis, in which no bacterial pathogen can be isolated by routine cultures, can mimic bacterial meningitis, but the disease has a much more favorable prognosis . Many cases of aseptic meningitis are caused by viruses, primarily enteroviruses, but bacteria and noninfectious etiologies also cause meningitis with negative cultures . Symptoms of meningeal inflammation with CSF pleocytosis that persist for more than 4 weeks define the chronic meningitis syndrome . The diagnosis is based on the patient history, clinical evidence of meningitis, CSF examination, and often imaging studies . The differential diagnosis is broad, and the predominant CSF cell type can provide clues as to the underlying disease . Empiric therapy is primarily based on the age of the patient, with modifications if there are positive findings on CSF gram stain or if the patient presents with special risk factors . In patients with chronic meningitis, a definite diagnosis is often not available or delayed for days, in which case empiric therapy may have to be initiated . It is important to cover the treatable causes of meningitis, for which the outcome is poor if treatment is delayed. Microbiol Immunol, 1999, 43(9), 821 - 7 Identification and semiquantitation of Mycobacterium avium using a competitive PCR method; Hashimoto T et al.; A competitive PCR method with standard DNA (MIMIC) was developed for the rapid detection and semiquantitation of Mycobacterium avium (M . avium) using primers specific for the alpha antigen sequence of the bacteria . DNA from both M . avium and Mycobacterium marinum was amplified by polymerase chain reaction (PCR), but only M . avium could be detected by subsequent blotting confirmation with a probe specific for the bacteria . With the PCR and subsequent dot blot hybridization, as little as 10 fg of the M . avium DNA could be detected, equivalent to about 2 cells of the mycobacteria . In addition, we could distinguish 10(5) CFUs of M . avium from 10(4) CFUs or less by competitive PCR using a MIMIC . The present competitive PCR test enabled rapid identification and semiquantitation of M . avium, and could be used clinically to monitor disease severity and response to treatment of human M . avium disease. Syst Appl Microbiol, 1999 Sep, 22(3), 466 - 71 Improved technique for the isolation of magnetotactic spirilla from a freshwater sediment and their phylogenetic characterization; Schuler D et al.; An improved technique for the isolation of magnetotactic bacteria was used for the axenic cultivation of microaerophilic magnetotactic spirilla . Magnetotactic bacteria were first separated from non-magnetic contaminants by exploiting their active migration along magnetic field lines by a capillary "racetrack" method . The purified magnetic cells were then inoculated into a two-layer isolation medium with opposing oxygen and sulfide gradients . Several strains of magnetotactic spirilla were isolated from a freshwater sediment sample using this method . Based on their morphology, physiology and comparative analysis of almost complete 16S rRNA gene sequences, all newly isolated strains were identified as members of the genus Magnetospirillum . While five of the isolates were closely related to previously described species (> 99% sequence similarity), two isolates appear to represent a third phylogenetic cluster within the genus Magnetospirillum. Syst Appl Microbiol, 1999 Sep, 22(3), 387 - 92 Diversity and relationships of Bradyrhizobia from Amphicarpaea bracteata based on partial nod and ribosomal sequences; Sterner JP et al.; Partial sequences of three nod genes (nodC, nodD1, and nodA 5' flanking region) and of 16S and 23S rDNA were obtained from isolates of Bradyrhizobium sp . associated with the native North American legume Amphicarpaea bracteata . Isolates from Amphicarpaea had identical sequences in the three nod gene regions, but differed from all other Bradyrhizobium taxa at > 10% of nucleotide sites . Parsimony analysis of all nod gene segments indicated a phylogenetic relationship of these bacteria to B . elkanii, with B . japonicum diverging prior to the diversification of these taxa . All Bradyrhizobium isolates from Amphicarpaea were also identical to B . elkanii in the size of the intervening sequence (IVS) in the 5' region of the 23S rRNA gene, while B . japonicum had an IVS length variant with 29 additional nucleotides . Parsimony analysis of both 16S and 23S partial rDNA sequences grouped Bradyrhizobium sp . isolates from Amphicarpaea into a clade together with B . elkanii, consistent with the relationships inferred from nod sequences. J Appl Microbiol, 1999 Nov, 87(5), 683 - 8 Prevalence of mycobacteria in a swimming pool environment; Leoni E et al.; A study was performed to evaluate the prevalence of non-tubercular mycobacteria in swimming pool environments . The bacteria in question were found in 88.2% of pool water samples . The most frequent species were Mycobacterium gordonae (73.5% of samples; range 1-840 cfu 100 ml - 1), M . chelonei (38.2% 2-360 cfu 100 ml - 1) and M . fortuitum (35.3% 2-250 cfu 100 ml - 1) . The same species were also recovered from the water at the different phases of the treatment cycle, with relative percentages similar to those of the pool water . Shower floors and pool edges also presented high concentrations of the mycobacteria (100% of samples) and M . marinum was isolated from the surfaces of pool edges on two occasions (4.5% of samples) . The swimming pool environment provides a suitable habitat for the survival and reproduction of mycobacteria . Although mycobacteria are common in swimming pools, human mycobacterial disease associated with their use is rare . Apart from superficial infections with M . marinum, the risk of more serious diseases in subjects with weakened immune systems should not be underestimated, given the widespread presence of mycobacteria that are possible opportunistic pathogens and the direct contact bathers have with the water and aerosol. Biochim Biophys Acta, 1999 Dec 27, 1473(2-3), 363 - 75 Analysis of the nifHDK operon and structure of the NifH protein from the unicellular, diazotrophic cyanobacterium, Cyanothece strain sp . ATCC 51142(1); Colon-Lopez MS et al.; Cyanothece sp . ATCC 51142 is a unicellular, diazotrophic cyanobacterium that demonstrates diurnal rhythms for photosynthesis and N(2) fixation, with peaks of O(2) evolution and nitrogenase activity approximately 12 h out of phase . We cloned and sequenced the nifHDK operon, and determined that the amino acid sequences of all three proteins were highly conserved relative to those of other cyanobacteria and bacteria . However, the Fe-protein, encoded by the nifH gene, demonstrated two differences from the related protein in Azotobacter vinelandii, for which a 3-D structure has been determined . First, the Cyanothece Fe-protein contained a 37 amino acid extension at the N-terminus . This approximately 4 kDa addition to the protein appeared to fold as a separate domain, but remained a part of the active protein, as was verified by migration on acrylamide gels . In addition, the Cyanothece Fe-protein had amino acid differences at positions involved in formation of the Fe-protein dimer-dimer contacts in A . vinelandii nitrogenase . There were also changes in residues involved with interaction between the Fe-protein and the MoFe-protein when compared with A . vinelandii . Since the Cyanothece Fe-protein is quickly degraded after activity, it is suggested that the extension and the amino acid alterations were somehow involved in this degradative process. Nippon Rinsho, 1999 Oct, 57(10), 2205 - 10 {Vitamin B12}; Watanabe F et al.; Vitamin B12 is unique among all the vitamins in that it contains not only a complex organic molecule but also an essential trace element, cobalt . Vitamin B12 is synthesized in some bacteria but not in animals and plants . Intestinal absorption and subsequent plasma transport of vitamin B12 are mediated by specific vitamin B12-binding proteins and their receptors in mammals . Vitamin B12 taken up by the cells is enzymatically converted to coenzyme forms of vitamin B12, methyl- and adenosyl-vitamin B12, which function as coenzymes of methionine synthase (EC 2.1.13), involved in methionine biosynthesis, and methylmalonyl-CoA mutase (EC 5.4.99.2), involved in oxidation of odd-numbered fatty acids and amino acids (valine, isoleucine, and threonine), respectively . Chemical properties, physiological function, and intracellular metabolism of vitamin B12 are summarized in this section. Nucleic Acids Res, 2000 Jan 1, 28(1), 169 - 70 tmRDB (tmRNA database); Zwieb C et al.; The tmRNA database (tmRDB) is maintained at the University of Texas Health Science Center at Tyler, Texas, and is accessible on the WWW at URL +html . A tmRDB mirror site is located on the campus of Auburn University, Auburn, Alabama, reachable at the URL Since April 1997, the tmRDB has provided sequences of tmRNA (previously called 10Sa RNA), a molecule present in most bacteria and some organelles . This release adds 17 new sequences for a total of 60 tmRNAs . Sequences and corresponding tmRNA-encoded tag peptides are tabulated in alphabetical and phylo-genetic order . The updated tmRNA alignment improves the secondary structures of known tmRNAs on the level of individual basepairs . tmRDB also provides an introduction to tmRNA function in trans-translation (with links to relevant literature), a limited number of tmRNA secondary structure diagrams, and numerous three-dimensional models generated interactively with the program ERNA-3D. Nucleic Acids Res, 2000 Jan 1, 28(1), 33 - 6 The COG database: a tool for genome-scale analysis of protein functions and evolution; Tatusov RL et al.; Rational classification of proteins encoded in sequenced genomes is critical for making the genome sequences maximally useful for functional and evolutionary studies . The database of Clusters of Orthologous Groups of proteins (COGs) is an attempt on a phylogenetic classification of the proteins encoded in 21 complete genomes of bacteria, archaea and eukaryotes . The COGs were constructed by applying the criterion of consistency of genome-specific best hits to the results of an exhaustive comparison of all protein sequences from these genomes . The database comprises 2091 COGs that include 56-83% of the gene products from each of the complete bacterial and archaeal genomes and approximately 35% of those from the yeast Saccharomyces cerevisiae genome . The COG database is accompanied by the COGNITOR program that is used to fit new proteins into the COGs and can be applied to functional and phylogenetic annotation of newly sequenced genomes. Zhonghua Min Guo Wei Sheng Wu Ji Mian Yi Xue Za Zhi, 1996 May, 29(2), 108 - 15 Comparison of antigen binding capabilities of various membrane filters and filter papers in dot immunoassay; Lin MY et al.; Antigen binding activities of 25 kinds of filter papers, including nitrocellulose (NC), nylon or polyvinylidine difluoride (PVDF), in the binding of 5 viruses, 3 bacteria, 2 mycoplasmas and 1 chicken serum protein antigens in dot immunoassay were compared . Immobilon affinity membrane type D (IAM-D) was found best in binding viral antigens, followed by Ultrabind SV-450 (SV-450) . SV-450 was found best in binding bacterial antigen, followed by Ultrabind US-450 (US-450), IAM-D and NC 0.45 micron . IAM-D was the best in binding mycoplasma antigen, followed by Ultrabind HP, US-450, NC 0.2 micron . Overall, IAM-D had the best capability in the binding of the antigens. Mycoses, 1999, 42(9-10), 525 - 8 Identification of dermatophytes by Fourier transform infrared spectroscopy (FT-IR); Bastert J et al.; Fourier transform infrared spectroscopy is an established method in the routine diagnosis of various micro-organisms, including bacteria and yeasts, on a species level . Its possible value in the diagnostics of dermatophytes was analysed using three clinical isolates each of the three most frequently found species, namely Trichophyton rubrum, Trichophyton mentagrophytes and Microsporum canis . The results encourage further work to establish a library which would allow the use of this method in the clinical setting . This might help to make repeated subcultures, which are money- and time-consuming, redundant. Hum Biol, 1999 Dec, 71(6), 901 - 14 Deletion polymorphism in the human COL1A2 gene: genetic evidence of a non-African population whose descendants spread to all continents; Mitchell RJ et al.; We report the frequencies of a deletion polymorphism at the alpha 2 (1) collagen gene (COL1A2) and argue that this distribution has major implications for understanding the evolution of modern humans immediately after their exodus from sub-Saharan Africa as well as their subsequent spread to all continents . The high frequency of the deletion in non-African populations and its complete absence in sub-Saharan African groups suggest that the deletion event occurred just before or shortly after modern humans left Africa . The deletion probably arose shortly after the African exodus in a group whose descendants were among the ancestors of all contemporary populations, except for sub-Saharan Africans . This, of course, does not imply that there was a single migration out of Africa . The GM immunoglobulin haplotype GM*A,X G displays a similar distribution to that for the COL1A2 deletion, and these 2 polymorphisms suggest that the exodus from Africa may not have been a rapid dispersion to all other regions of the world . Instead, it may have involved a period of time for the savanna-derived gene pool to adapt to novel selective agents, such as bacteria, viruses, and/or environmental xenobiotics found in both animal and plant foods in their new environment . In this context these polymorphisms are indicators of the evolution that occurred before the diaspora of these populations to the current distribution of modern peoples. Dis Aquat Organ, 1999 Oct 11, 38(1), 23 - 31 Production of the 57 kDa major surface antigen by a non-agglutinating strain of the fish pathogen Renibacterium salmoninarum; Senson PR et al.; The major surface antigen of Renibacterium salmoninarum, p57, is associated with cell autoagglutination and implicated as a virulence factor in fish infections . An autoagglutinating strain, JD24, caused 92% mortality when 2 x 10(7) cells were injected intraperitoneally into rainbow trout Oncorhynchus mykiss, while a non-agglutinating strain, MT 239, produced only 7% mortality after 100 d . The p57 antigen was present in the supernates of broth cultures of both strains when examined by western immunoblotting, and the gene for p57 was detected in both strains by PCR . Electron microscopy of cryopreserved thin sections showed an amorphous layer associated with the cell surface of JD24 which was not seen with MT 239 . While p57 from JD24 could reassociate with cells of both strains, p57 from MT 239 failed to restore haemagglutination activity to either strain . Biotinylation of bacterial surfaces demonstrated the presence of a carbohydrate component of p57 from JD24 which was absent from the p57 produced by MT 239 . The higher virulence of JD24 may depend not only on the production of p57, but also its direct association with the bacterial cell surface. Microb Pathog, 1999 Dec, 27(6), 419 - 27 Live Bartonella henselae enhances endothelial cell proliferation without direct contact; Maeno N et al.; The proliferation of human umbilical vein endothelial cells (HUVECs) cocultivated with live B . henselae was enhanced in a bacterial dose-dependent manner, and the stimulatory effect was specific to vascular endothelial cells . The inactivation of B . henselae by UV or heat treatment abolished its stimulatory activity, suggesting that live bacteria is necessary for the growth stimulation effect . To investigate the role of direct contact, live B . henselae were separated from HUVECs by a filter membrane (Millicell-CM insert) . Even under this condition, an enhanced proliferation of HUVECs was observed . However, no morphological changes in the HUVECs were apparent compared to the B . henselae -infected cells . Furthermore, we isolated a nonpiliated strain of B . henselae that is unable to attach to and enter into endothelial cells . The nonpiliated strain possessed the ability to stimulate the proliferation of cocultivated HUVECs the same as the piliated strain . Moreover, the culture supernatants of B . henselae were also able to induce HUVEC proliferation . Our results indicate that the stimulation of HUVEC proliferation by B . henselae is mediated by soluble factor(s) secreted from the bacteria . Am J Respir Crit Care Med, 1999 Dec, 160(6), 2048 - 55 Enhanced in vivo human immunodeficiency virus-1 replication in the lungs of human immunodeficiency virus-infected persons with Pneumocystis carinii pneumonia; Koziel H et al.; The relationship of serum human immunodeficiency virus-1 (HIV-1) RNA levels to HIV-1 RNA levels in other compartments, such as the lungs, is not well characterized . The purpose of this study was to determine the viral burden of HIV-1 in the lungs by comparing HIV-1 RNA in cell-free bronchoalveolar lavage fluid (BALF) with that in serum . Specimens were examined from 77 HIV-seropositive adults (CD4(+) cell counts: 0 to 700 cells/mm(3); 48% receiving prescribed antiretroviral agents), comprising 43 asymptomatic individuals who were compared with 34 persons with active lung disease caused by Pneumocystis carinii (n = 26), bacteria (n = 3), Mycobacterium avium complex (n = 2), Nocardia sp . (n = 1), Aspergillus sp . (n = 1), or pulmonary Kaposi's sarcoma (n = 1) . For serum HIV-1 RNA, the proportion of subjects with detectable levels and the mean values were similar for asymptomatic individuals and persons with active lung disease (85% versus 86%, respectively) (6.64 x 10(4) versus 1 . 81 x 10(5) HIV-1 RNA copies/ml; p = 0.13) . In contrast, HIV-1 RNA in BALF was more often detected (16% versus 62%; p = 0.001), and mean values were higher (1.04 x 10(5) versus 3.31 x 10(6) HIV-1 RNA copies/ml; p = 0.032), in subjects with active lung disease than in asymptomatic subjects, independent of early or advanced clinical stages of HIV-related disease . For both study groups, HIV-1 RNA levels in BALF exceeded those in serum in 56% of cases by up to 66-fold, and did not correlate with local levels of tumor necrosis factor-alpha, granulocyte-macrophage colony-stimulating factor, or interleukin-16 . HIV-1 proviral DNA in cells from BALF was detected in up to 86% of subjects, more frequently in persons with advanced HIV disease (p = 0.0496), and often involved > 10% of BALF cells, but did not correlate with HIV-1 RNA detected in BALF . These data provide evidence for active HIV-1 replication in the lungs . HIV-1 replication is compartmentalized relative to serum, may be restricted, is independent of HIV-1 proviral DNA and clinical stage of HIV, and may be influenced by pulmonary disease such as P . carinii pneumonia or by other local or lung-specific factors . The lungs represent a large reservoir for HIV-1, and may present a source of persistent HIV-1 replication even during periods of apparent clinical latency of HIV-1 infection. Biophys J, 1999 Dec, 77(6), 2920 - 9 Protein-assisted pericyclic reactions: an alternate hypothesis for the action of quantal receptors; Radding W et al.; The rules for allowable pericyclic reactions indicate that the photoisomerizations of retinals in rhodopsins can be formally analogous to thermally promoted Diels-Alder condensations of monoenes with retinols . With little change in the seven-transmembrane helical environment these latter reactions could mimic the retinal isomerization while providing highly sensitive chemical reception . In this way archaic progenitors of G-protein-coupled chemical quantal receptors such as those for pheromones might have been evolutionarily plagiarized from the photon quantal receptor, rhodopsin, or vice versa . We investigated whether the known structure of bacteriorhodopsin exhibited any similarity in its active site with those of the two known antibody catalysts of Diels-Alder reactions and that of the photoactive yellow protein . A remarkable three-dimensional motif of aromatic side chains emerged in all four proteins despite the drastic differences in backbone structure . Molecular orbital calculations supported the possibility of transient pericyclic reactions as part of the isomerization-signal transduction mechanisms in both bacteriorhodopsin and the photoactive yellow protein . It appears that reactions in all four of the proteins investigated may be biological analogs of the organic chemists' chiral auxiliary-aided Diels-Alder reactions . Thus the light receptor and the chemical receptor subfamilies of the heptahelical receptor family may have been unified at one time by underlying pericyclic chemistry. Biochem J, 1999 Dec 15, 344 Pt 3, 625 - 31 Neuropathy target esterase; Glynn P; Neuropathy target esterase (NTE) is an integral membrane protein present in all neurons and in some non-neural-cell types of vertebrates . Recent data indicate that NTE is involved in a cell-signalling pathway controlling interactions between neurons and accessory glial cells in the developing nervous system . NTE has serine esterase activity and efficiently catalyses the hydrolysis of phenyl valerate (PV) in vitro, but its physiological substrate is unknown . By sequence analysis NTE has been found to be related neither to the major serine esterase family, which includes acetylcholinesterase, nor to any other known serine hydrolases . NTE comprises at least two functional domains: an N-terminal putative regulatory domain and a C-terminal effector domain which contains the esterase activity and is, in part, conserved in proteins found in bacteria, yeast, nematodes and insects . NTE's effector domain contains three predicted transmembrane segments, and the active-site serine residue lies at the centre of one of these segments . The isolated recombinant domain shows PV hydrolase activity only when incorporated into phospholipid liposomes . NTE's esterase activity appears to be largely redundant in adult vertebrates, but organophosphates which react with NTE in vivo initiate unknown events which lead, after a delay of 1-3 weeks, to a neuropathy with degeneration of long axons . These neuropathic organophosphates leave a negatively charged group covalently attached to the active-site serine residue, and it is suggested that this may cause a toxic gain of function in NTE. J Agric Food Chem, 1999 Oct, 47(10), 4478 - 83 Inheritance of morphological characters and glycoalkaloids in potatoes of somatic hybrids between dihaploid Solanum acauleand tetraploid Solanum tuberosum; Kozukue N et al.; Steroidal glycoalkaloids occur in potatoes and are reported to impart resistance to phytopathogens including bacteria, fungi, and insects . Because glycoalkaloids can be passed to progenies during breeding programs designed to develop improved potatoes, it is of importance to determine the quality of desired characteristics and the composition of glycoalkaloids of new somatic hybrids . The objective of this study was to determine the appearance, size, and shape (morphological characters) as well as the glycoalkaloid content of potato tubers of somatic hybrids between tetraploid Solanum tuberosum cv . Dejima (2n = 4x = 48 chromosomes) and the dihaploid clone ATDH-1 (2n = 2x = 24 chromosomes) induced by anther culture from Solanum acuale-T (acl-T, 2n = 4x = 48 chromosomes) . Tuber size and shape in somatic hybrids were in accord with those of cv . Dejima, whereas the tuber skin color resembled that of ATDH-1 . Thin-layer chromatography, high-performance liquid chromatography, and gas-liquid chromatography/mass spectrometry studies showed that the two steroidal glycoalkaloids (alpha-chaconine and alpha-solanine) were present in the tubers of S . tuberosum, whereas acl-T and ATDH-1 tubers were found to contain alpha-tomatine and demissine . The concentrations of total glycoalkaloids in both acl-T and ATDH-1 was >100 mg/100 g of fresh weight tuber cortex, much higher than in S . tuberosum . All somatic hybrids, except one clone, contained four glycoalkaloids (alpha-chaconine, alpha-solanine, alpha-tomatine, and demissine) derived from the fusion parents . The lack of alpha-tomatine in the remaining clone may be due to somaclonal variation . The results show that character expression is influenced by ploidy level and that total glycoalkaloid levels in most somatic hybrids were intermediate between those of the fusion parents . The possible significance of these findings for plant breeding and food safety is discussed. J Virol, 2000 Jan, 74(1), 1 - 7 Hepatitis B virus core gene mutations which block nucleocapsid envelopment; Koschel M et al.; Recently we generated a panel of hepatitis B virus core gene mutants carrying single insertions or deletions which allowed efficient expression of the core protein in bacteria and self-assembly of capsids . Eleven of these mutations were introduced into a eukaryotic core gene expression vector and characterized by trans complementation of a core-negative HBV genome in cotransfected human hepatoma HuH7 cells . Surprisingly, four mutants (two insertions {EFGA downstream of A11 and LDTASALYR downstream of R39} and two deletions {Y38-R39-E40 and L42}) produced no detectable capsids . The other seven mutants supported capsid formation and pregenome packaging/viral minus- and plus-strand-DNA synthesis but to different levels . Four of these seven mutants (two insertions {GA downstream of A11 and EHCSP downstream of P50} and two deletions {S44 and A80}) allowed virion morphogenesis and secretion . The mutant carrying a deletion of A80 at the tip of the spike protruding from the capsid was hepatitis B virus core antigen negative but wild type with respect to virion formation, indicating that this site might not be crucial for capsid-surface protein interactions during morphogenesis . The other three nucleocapsid-forming mutants (one insertion {LS downstream of S141} and two deletions {T12 and P134}) were strongly blocked in virion formation . The corresponding sites are located in the part of the protein forming the body of the capsid and not in the spike . These mutations may alter sites on the particle which contact surface proteins during envelopment, or they may block the appearance of a signal for the transport or the maturation of the capsid which is linked to viral DNA synthesis and required for envelopment. Mol Biochem Parasitol, 1999 Oct 25, 104(1), 121 - 30 Substrate depletion upregulates uptake of myo-inositol, glucose and adenosine in Leishmania; Seyfang A et al.; Leishmania flagellates undergo a digenetic life cycle in the gut of the sandfly insect vector and in macrophage phagolysosomes of the mammalian host . This involves vast changes of the environment to which the parasite has to adapt, including temperature, pH and concentration of nutrients between different types of meals of the insect vector or within the enclosed intracellular environment of the phagolysosome . The regulation of transporters for important organic substrates in Leishmania donovani, Leishmania mexicana and Leishmania enriettii has been investigated . A pronounced upregulation of inositol (25-fold), adenosine (11-fold) or glucose (5-fold) uptake activities was found when cells were depleted of the respective substrates during culture . Inositol-depleted cells showed a half-maximal uptake rate at nanomolar inositol concentration . Depletion of inositol only affected inositol uptake but did not affect uptake of glucose analog or proline in control experiments, indicating the specificity of the mechanism(s) underlying transport regulation . Adenosine-depleted cells showed an approximately 10-fold increase in both adenosine and uridine uptake, both mediated by the L . donovani nucleoside transporter 1 (LdNT1), but no change in guanosine uptake, which is mediated by the L . donovani nucleoside transporter 2 (LdNT2) . These results suggest that extracellular adenosine concentration specifically regulates LdNT1 transport activity and does not affect LdNT2 . The data imply that upregulation of transport activities by substrate depletion is a general phenomenon in protozoan flagellates, which is in remarkable contrast to bacteria where upregulation typically follows an increase of extracellular organic substrate . Hence, the parasites can maximize the uptake of important nutrients from the host even under limiting conditions, whereas bacteria often have dormant stages (spores) to overcome unfavorable environmental conditions or are heterotrophic for organic substrates. Curr Opin Clin Nutr Metab Care, 1999 Sep, 2(5), 399 - 404 The gut: the 'motor' of multiple organ dysfunction syndrome? Nieuwenhuijzen GA, Goris RJ. Abnormal colonization, gut-origin infections, and bacterial translocation are all signs of gut dysfunction that may be implicated in the pathogenesis of multiple organ dysfunction syndrome (MODS) . This review summarizes and updates relevant experimental and clinical data that have attempted to correlate these phenomena with the development of MODS and to answer whether or not the gut is the 'motor' of MODS . The presented data suggest that, in some patients, gut dysfunction may precede the development of MODS . However, in most patients, this relationship is less obvious . The gut may still be one of the motors of MODS; however, it does not appear that this motor is fueled by the systemic spread of bacteria . Bacteria may play a role on a local gut-associated level in initiating and perpetuating the production of local inflammatory mediators that may produce distant organ injury. Proc Natl Acad Sci U S A, 1999 Dec 7, 96(25), 14412 - 7 The DNMT3B DNA methyltransferase gene is mutated in the ICF immunodeficiency syndrome; Hansen RS et al.; DNA methylation is an important regulator of genetic information in species ranging from bacteria to humans . DNA methylation appears to be critical for mammalian development because mice nullizygous for a targeted disruption of the DNMT1 DNA methyltransferase die at an early embryonic stage . No DNA methyltransferase mutations have been reported in humans until now . We describe here the first example of naturally occurring mutations in a mammalian DNA methyltransferase gene . These mutations occur in patients with a rare autosomal recessive disorder, which is termed the ICF syndrome, for immunodeficiency, centromeric instability, and facial anomalies . Centromeric instability of chromosomes 1, 9, and 16 is associated with abnormal hypomethylation of CpG sites in their pericentromeric satellite regions . We are able to complement this hypomethylation defect by somatic cell fusion to Chinese hamster ovary cells, suggesting that the ICF gene is conserved in the hamster and promotes de novo methylation . ICF has been localized to a 9-centimorgan region of chromosome 20 by homozygosity mapping . By searching for homologies to known DNA methyltransferases, we identified a genomic sequence in the ICF region that contains the homologue of the mouse Dnmt3b methyltransferase gene . Using the human sequence to screen ICF kindreds, we discovered mutations in four patients from three families . Mutations include two missense substitutions and a 3-aa insertion resulting from the creation of a novel 3' splice acceptor . None of the mutations were found in over 200 normal chromosomes . We conclude that mutations in the DNMT3B are responsible for the ICF syndrome. Microb Comp Genomics, 1999, 4(3), 187 - 201 Computational analysis of the polymorphic membrane protein superfamily of Chlamydia trachomatis and Chlamydia pneumoniae; Grimwood J et al.; Whole sequence genome analysis is invaluable in providing complete profiles of related proteins and gene families . The genome sequences of the obligate intracellular bacteria Chlamydia trachomatis and Chlamydia pneumoniae both encode proteins with similarity to several 90-kDa Chlamydia psittaci proteins . These proteins are members of a large superfamily, C . trachomatis with 9 members and C . pneumoniae with 21 members . All polymorphic membrane protein (Pmp) are heterogeneous, both in amino acid sequence and in predicted size . Most proteins have apparent signal peptide leader sequences and hence are predicted to be localized to the outer membrane . The unifying features of all proteins are the conserved amino acid motifs GGAI and FXXN repeated in the N-terminal half of each protein . In both genomes, the pmp genes are clustered at various locations on the chromosome . Phylogenetic analysis suggests six related families, each with at least one C . trachomatis and one C . pneumoniae orthologue . One of these families has seen prolific expansion in C . pneumoniae, resulting in 13 protein paralogues . The maintenance of orthologues from each species suggests specific functions for the proteins in chlamydial biology. Biochemistry, 1999 Dec 7, 38(49), 16347 - 58 Characterization of unique DNA-binding and transcriptional-activation functions in the carboxyl-terminal extension of the zinc finger region in the human vitamin D receptor; Hsieh JC et al.; The vitamin D receptor (VDR) binds 1,25-dihydroxyvitamin D(3) and mediates its actions on gene transcription by heterodimerizing with retinoid X receptors (RXRs) on direct repeat (DR+3) vitamin D responsive elements (VDREs) located in target genes . The VDRE binding function of VDR has been primarily ascribed to the zinc finger region (residues 24-87) . To define the minimal VDRE binding domain for human VDR (hVDR), a series of C-terminally truncated hVDR mutants (Delta134, Delta113, Delta102, Delta90, Delta84, Delta80, and Delta60) was generated and expressed in bacteria . Only the Delta134 and Delta113 mutants bound the VDRE (predominantly as monomers), suggesting that, in addition to the conserved zinc finger region of hVDR, as many as 25 amino acids in a C-terminal extension (CTE) participate in DNA binding . Site-directed mutagenesis of conserved charged residues in full-length hVDR was then performed to dissect the functional significance of the CTE (residues 88-112) in the context of the complete hVDR-RXR-VDRE interaction . Functional assays revealed that E98K/E99K, R102A/K103A/R104A, and K109A/R110A/K111A mutant hVDRs possessed dramatically reduced DNA binding and transcriptional activities, whereas distinct point mutants, such as K103A, bound to DNA normally but lacked transcriptional activity . Therefore, the boundary for the minimal DNA-binding domain in hVDR extends C-terminal of the zinc fingers to Lys-111, with clusters of highly conserved charged amino acids playing a crucial role in binding to the DR+3 element . Further, individual residues in this region (e.g., Lys-103) may lie on the opposing face of a DNA-binding alpha-helix, where they could contact transcriptional coactivators or basal transcription factors. J Immunol, 1999 Dec 15, 163(12), 6748 - 55 The CD14 ligands lipoarabinomannan and lipopolysaccharide differ in their requirement for Toll-like receptors; Means TK et al.; Mammalian Toll-like receptor (TLR) proteins are new members of the IL-1 receptor family that participate in activation of cells by bacteria and bacterial products . Several recent reports indicate that TLR proteins mediate cellular activation by bacterial LPS via a signaling pathway that is largely shared by the type I IL-1 receptor . We previously showed that Chinese hamster ovary (CHO) fibroblasts engineered to express CD14 (CHO/CD14) were responsive to LPS, but not to a distinct CD14 ligand, mycobacterial lipoarabinomannan (LAM) . These CHO/CD14 cells were subsequently found to possess a frame-shift mutation within the TLR2 gene which resulted in their inability to express functional TLR2 protein . Thus, we hypothesized that TLR2, but not TLR4, was necessary for LAM signaling . In this paper we show that CHO/CD14 cells engineered to express functional TLR2 protein acquired the ability to be activated by LAM . Similarly, overexpression of TLR2 in murine macrophages conferred enhanced LAM responsiveness . Together, our data demonstrate that the distinct CD14 ligands LAM and LPS utilize different TLR proteins to initiate intracellular signals . These findings suggest a novel receptor signaling paradigm in which the binding of distinct ligands is mediated by a common receptor chain, but cellular activation is initiated via distinct signal-transducing chains that confer ligand specificity . This paradigm contrasts with many cytokine receptor complexes in which receptor specificity is conferred by a unique ligand-binding chain but cellular activation is initiated via shared signal-transducing chains. Nat Struct Biol, 1999 Dec, 6(12), 1126 - 31 Crystal structure of transhydrogenase domain III at 1.2 A resolution; Prasad GS et al.; The nicotinamide nucleotide transhydrogenases (TH) of mitochondria and bacteria are membrane-intercalated proton pumps that transduce substrate binding energy and protonmotive force via protein conformational changes . In mitochondria, TH utilizes protonmotive force to promote direct hydride ion transfer from NADH to NADP, which are bound at the distinct extramembranous domains I and III, respectively . Domain II is the membrane-intercalated domain and contains the enzyme's proton channel . This paper describes the crystal structure of the NADP(H) binding domain III of bovine TH at 1.2 A resolution . The structure reveals that NADP is bound in a manner inverted from that previously observed for nucleotide binding folds . The non-classical binding mode exposes the NADP(H) nicotinamide ring for direct contact with NAD(H) in domain I, in accord with biochemical data . The surface of domain III surrounding the exposed nicotinamide is comprised of conserved residues presumed to form the interface with domain I during hydride ion transfer . Further, an adjacent region contains a number of acidic residues, forming a surface with negative electrostatic potential which may interact with extramembranous loops of domain II . Together, the distinctive surface features allow mechanistic considerations regarding the NADP(H)-promoted conformation changes that are involved in the interactions of domain III with domains I and II for hydride ion transfer and proton translocation. Proteins, 1999 Nov 1, 37(2), 303 - 9 Multifunctional enzymes and evolution of biosynthetic pathways: retro-evolution by jumps; Roy S; A likely scenario of evolution of biosynthetic pathways is believed to have occurred by retro-evolution through recruitment of existing enzymes rather than generation of de novo classes . It had been proposed that such retro-evolution occurred in steps as a response to depletion of an essential metabolite and availability of another related substance in the environment . In this article, I argue that because of instability of many such extant intermediates, it is unlikely that retro-evolution had occurred in steps . I further propose that such evolution in many cases has taken place by jumps, i.e., by recruitment of a multifunctional enzyme capable of catalyzing several steps at a time, albeit inefficiently . I further speculate that in some cases one primordial multienzyme may have catalyzed the whole sequence of reaction of a biosynthetic pathway, i.e., the pathway may have evolved by a single leap . Gene duplications and further evolution to more efficient enzymes led to extant pathways . Such a mechanism predicts that some or all enzymes of a pathway must have descended from a common ancestor . Sequence and structural homologies among extant enzymes of a biosynthetic pathway have been examined. Appl Environ Microbiol, 1999 Dec, 65(12), 5607 - 11 Isolation of Terrabacter sp . strain DDE-1, which metabolizes 1, 1-dichloro-2,2-bis(4-chlorophenyl)ethylene when induced with biphenyl; Aislabie J et al.; Terrabacter sp . strain DDE-1, able to metabolize 1,1-dichloro-2, 2-bis(4-chlorophenyl)ethylene (DDE) in pure culture when induced with biphenyl, was enriched from a 1-1-1-trichloro-2, 2-bis(4-chlorophenyl)ethane residue-contaminated agricultural soil . Gas chromatography-mass spectrometry analysis of culture extracts revealed a number of DDE catabolites, including 2-(4'-chlorophenyl)-3,3-dichloropropenoic acid, 2-(4'-chlorophenyl)-2-hydroxy acetic acid, 2-(4'-chlorophenyl) acetic acid, and 4-chlorobenzoic acid. Appl Environ Microbiol, 1999 Dec, 65(12), 5541 - 5 Estimation of methanogen biomass by quantitation of coenzyme M; Elias DA et al.; Determination of the role of methanogenic bacteria in an anaerobic ecosystem often requires quantitation of the organisms . Because of the extreme oxygen sensitivity of these organisms and the inherent limitations of cultural techniques, an accurate biomass value is very difficult to obtain . We standardized a simple method for estimating methanogen biomass in a variety of environmental matrices . In this procedure we used the thiol biomarker coenzyme M (CoM) (2-mercaptoethanesulfonic acid), which is known to be present in all methanogenic bacteria . A high-performance liquid chromatography-based method for detecting thiols in pore water (A . Vairavamurthy and M . Mopper, Anal . Chim . Acta 78:363-370, 1990) was modified in order to quantify CoM in pure cultures, sediments, and sewage water samples . The identity of the CoM derivative was verified by using liquid chromatography-mass spectroscopy . The assay was linear for CoM amounts ranging from 2 to 2,000 pmol, and the detection limit was 2 pmol of CoM/ml of sample . CoM was not adsorbed to sediments . The methanogens tested contained an average of 19.5 nmol of CoM/mg of protein and 0.39 +/- 0.07 fmol of CoM/cell . Environmental samples contained an average of 0.41 +/- 0.17 fmol/cell based on most-probable-number estimates . CoM was extracted by using 1% tri-(N)-butylphosphine in isopropanol . More than 90% of the CoM was recovered from pure cultures and environmental samples . We observed no interference from sediments in the CoM recovery process, and the method could be completed aerobically within 3 h . Freezing sediment samples resulted in 46 to 83% decreases in the amounts of detectable CoM, whereas freezing had no effect on the amounts of CoM determined in pure cultures . The method described here provides a quick and relatively simple way to estimate methanogenic biomass. Science, 1999 Dec 3, 286(5446), 1893 - 7 Quality control mechanisms during translation; Ibba M et al.; Translation uses the genetic information in messenger RNA (mRNA) to synthesize proteins . Transfer RNAs (tRNAs) are charged with an amino acid and brought to the ribosome, where they are paired with the corresponding trinucleotide codon in mRNA . The amino acid is attached to the nascent polypeptide and the ribosome moves on to the next codon . The cycle is then repeated to produce a full-length protein . Proofreading and editing processes are used throughout protein synthesis to ensure the faithful translation of genetic information . The maturation of tRNAs and mRNAs is monitored, as is the identity of amino acids attached to tRNAs . Accuracy is further enhanced during the selection of aminoacyl-tRNAs on the ribosome and their base pairing with mRNA . Recent studies have begun to reveal the molecular mechanisms underpinning quality control and go some way to explaining the phenomenal accuracy of translation first observed over three decades ago. Heredity, 1999 Oct, 83(# (Pt 4)), 469 - 75 Wolbachia infection in the terrestrial isopod oniscus asellus: sex ratio distortion and effect on fecundity Rigaud T, Moreau J, Juchault P. Maternally inherited Wolbachia bacteria are widespread in arthropods where they are responsible for various reproductive alterations . In terrestrial isopods (woodlice), Wolbachia may induce feminization or cytoplasmic incompatibility (CI), but their effect remains unknown in most host species . To increase our understanding of host/symbiont interactions in terrestrial isopods, the effect of Wolbachia was investigated in the oniscidean Oniscus asellus, mainly to discriminate between feminization and CI . The Wolbachia infection was not linked with a CI phenomenon, but females infected with Wolbachia produced female-biased broods compared with uninfected females . The fecundity of infected females was slightly lower than that of uninfected, but the number of young at the adult stage was similar between the two female categories . The experimental transfer of the symbiont into uninfected strains showed that Wolbachia was responsible for the feminization of a number of genetic males . In female-biased broods, Wolbachia were vertically transmitted to around 88% of the offspring, but the transmission rate was lower in the few male-biased progenies . The feminizing activity of these symbionts was not systematic, as many phenotypic males were infected . These results contrasted with what is known in another woodlouse species, and indicated that feminization has evolved in different ways in terrestrial isopods. Eur J Clin Invest, 1999 Nov, 29(11), 964 - 72 The growth and the control of human immunodeficiency virus in the lung: implications for highly active antiretroviral therapy; White NC et al.; In recent years, it has become apparent that the lung is an important niche for the proliferation of human immunodeficiency virus (HIV), which may have implications for highly active antiretroviral therapy (HAART) . The lung itself is a major site for the opportunistic infections associated with the progression to acquired immune deficiency syndrome (AIDS), specifically Pneumocystis carinii, Myobacterium tuberculosis and pyogenic bacteria . These cases of active pulmonary complications are direct indicators of enhanced progression to AIDS-defining illness and increased morbidity and mortality . It is therefore essential that the interaction between the lung and HIV is fully understood . Recent research indicates the lung may be a major sanctuary for the virus, with distinct evolution and replication in contrast to other target organs for HIV . In this review, we will discuss the recent findings of HIV infection, evolution, host factors involved in the control of HIV within the lung and the impact this may have on current therapy. Eur J Biochem, 1999 Dec, 266(3), 964 - 76 Implications of hemolin glycosylation and Ca2+-binding on homophilic and cellular interactions; Bettencourt R et al.; Insects are useful models for the study of innate immune mechanisms because of their lack of antibodies and receptors involved in adaptive immune response . Nevertheless, hemolin cloned from moths is a soluble and membrane associated Ig-related molecule that is up-regulated during immune response {Lanz-Mendoza, H . & Faye, I . (1999) Dev . Comp . Immunol . 23, 359-374} . The hemolin monomeric form has four, pair-wise, interacting Ig-domains, forming a strongly bent horseshoe structure {Su, X.-D., Gastinel, L.N., Vaughn, D.E., Faye, I., Poon, P . & Bjorkman, P . (1998) Science 281, 991-995} . To elucidate the nature of its homophilic and cellular interactions, the glycosylation and Ca2+-binding properties of hemolin were investigated . We used Hyalophora cecropia hemolin isolated from hemolymph of bacteria-injected pupae, or produced as a recombinant protein in a baculovirus/insect cell system . Both types of hemolin contain N-acetylglucosamine and probably sialic acid, as indicated by peptide:N-glycosidase F and neuraminidase digestion and glycosylation detection by Western-blotting analysis . The N-acetylglucosamine residues on hemolin were confirmed with the use of specific lectins . In addition, hemolin was shown to specifically bind calcium when spotted onto nitrocellulose and treated as for 45Ca2+ autoradiography . Earlier studies demonstrated that hemolin can bind to hemocytes and this was tested for its dependence on calcium and carbohydrates, using hemolin-coated fluorescent microspheres . A greater level of attachment of microspheres occurred in the presence of calcium than if calcium was absent . Furthermore, this binding was inhibited by EGTA and N-acetylglucosamine or N-acetylneuraminic acid, implying that carbohydrates and calcium are crucial factors in homophilic binding and cell-adhesion events mediated by this Ig-superfamily molecule. Hum Reprod Update, 1999 Sep-Oct, 5(5), 399 - 420 Infections in the male genital tract and reactive oxygen species; Ochsendorf FR; In the male genital tract, reactive oxygen species (ROS) are generated by spermatozoa and leukocytes including neutrophils and macrophages . ROS are involved in the regulation of sperm functions such as capacitation and the acrosome reaction . Infections lead to an excessive ROS production, resulting in an 'oxidative burst' from neutrophils/macrophages as a first-line defence mechanism . This is modulated by several cytokines and the pro-oxidant mechanisms of bacteria and viruses . At the site of an infection, the degree of activation of leukocytes, i.e . the amount of ROS produced, and the available antioxidative systems determine whether spermatozoa are damaged or not . During an infection, an imbalance of pro- and antioxidants favouring the former results in oxidative stress which impairs the sperm functions mentioned, as well as motility and fertilization . ROS produced during infections of the testis and epididymis are especially harmful to spermatozoa due to the longer contact time and the lack of antioxidant protection . In the final ejaculate, only very high numbers of ROS-producing leukocytes are detrimental to sperm functions . An infectious injury involving ROS in the prostate gland, seminal vesicles or epididymis could impair sperm functions indirectly . Pro- and antioxidative properties of therapeutics are currently receiving more attention as part of anti-infectious therapies . At present, there are many unresolved questions concerning the exact role of ROS during infections of the male genital tract because of the difficulty of specifically assessing the site of generation and the short-lived effects of ROS . New techniques may enable specific studies to fill this gap in the near future. J Hosp Infect, 1999 Nov, 43(3), 219 - 29 Properties of an enzyme-based low-level iodine disinfectant; Duan Y et al.; An enzyme-based iodine (EBI) disinfectant that continuously generates free molecular iodine in a controlled fashion was developed and evaluated for use in disinfecting flexible fibreoptic endoscopes (FFEs) . EBI is a powder concentrate that produces iodine from sodium iodide and calcium peroxide when catalyzed by horseradish peroxidase . After dissolution in water, it delivers relatively high concentrations of free molecular iodine (> 15 ppm) at relatively low concentrations of total iodine (30-40 ppm) . It demonstrates the ability to function as an effective low level iodine disinfectant by rapidly inactivating bacteria, fungi and viruses . A unique feature of the EBI system is the ability to reoxidize reduced iodine which results in a constant level of active (free molecular) iodine during use . EBI inactivates Mycobacterium bovis var BCG more rapidly than 2% glutaraldehyde (Cidex-7) . Its sporicidal activity, however, was found to be slower than the aldehyde formulation . The qualification of EBI for use as a practical disinfectant was shown by its negligible toxicity in dermal, ocular, oral and inhalation studies on animals, which is attributed to the low level of total iodine in the solution. Forensic Sci Int, 1999 Oct 11, 104(2-3), 179 - 87 Increase of pulmonary density of macrophages in sudden infant death syndrome; Lorin de la Grandmaison G et al.; A 1996 cytodensitometric study found increased cellular density in the pulmonary parenchyma of infants who died of sudden infant death syndrome (SIDS) . The present study clarifies these results in quantifying the density of immunohistochemical subtyped inflammatory cells . Histomorphometry was used to compare the density of macrophages, granulocytes and T and B lymphocytes in the lungs of two groups of infants . From the post-mortem records of infant deaths between 1983 and 1995, 29 (mean age = 5 months) were randomly selected including 16 cases of SIDS and 13 who died of other non-pulmonary causes . Densities of immunoreactive cells were measured under blind conditions in the parenchyma . The mean density of macrophages was significantly higher in cases of SIDS compared with the controls (P = 0.0318), but there were no differences for the lymphocytes and the granulocytes . These morphometrical results must be interpreted within the methodological limits of this study, especially the non-uniform level of lung inflation between selected subjects . However, the differences in level of inflation are not sufficient to explain the observed increase of macrophage density . Indeed, the mean values of alveolar surface area, which represent an indirect measure of lung inflation, are not significantly different between the two groups . Increase of pulmonary macrophage density in SIDS agrees with three non-exclusive hypotheses: (1) an abnormal inflammatory reaction by expression of Th1 helper cell phenotype activation; (2) consequence of passive smoking; and (3) post-agonal mechanisms . Bacterial superantigens produced by toxigenic bacteria in the respiratory tract could play a role as a trigger factor that initiates a fatal cascade with overproduction of cytokines leading to death . The significant increase of pulmonary macrophage density would be the morphological expression of this potential mechanism of death. Appl Biochem Biotechnol, 1999 Aug, 81(2), 107 - 17 Anaerobic toxicity and biodegradability of hydrolysis products of chemical warfare agents; Sklyar VI et al.; The toxicity and biodegradability of the main hydrolysis products of chemical warfare agents were investigated under methanogenic conditions . Among the tested substances, only MPhA does not have any toxic effect with regard to the aceticlastic methanogenic activity . The toxicity of other compounds varied between moderate (TDG, mercaptoethanol) to strong (ethanolamine, diisobutyl ester of MPhA) . Biodegradability tests showed that all the products of chemical detoxification of mustard gas (ethanolamine, ethylene glycol, TDG, mercaptoethanol) can be biomineralized under methanogenic conditions . On the contrary, phosphorus-containing compounds from the chemical detoxification of nerve warfare agents (Sarin, Soman, Vx-gases) are quite persistent under these conditions. Infect Control Hosp Epidemiol, 1999 Nov, 20(11), 736 - 40 Safety of peripheral intravenous catheters in children; Shimandle RB et al.; OBJECTIVES: To determine the overall and per-day risk of complications of short peripheral intravenous (PIV) catheters placed for indefinite periods . DESIGN: During 5 months, general pediatric patients receiving intravenous therapy through short PIV catheters were monitored . Patient and catheter characteristics were recorded, complications were noted, and rolled semiquantitative cultures of removed catheters were performed . Major endpoints were infection and phlebitis . Per-day risk of complications and catheter colonization (>15 colony-forming units) were calculated . SETTING: University children's hospital . PATIENTS: General pediatric ward inpatients with PIV . RESULTS: We studied 642 Teflon catheters in place >24 hours (mean, 3.7 days) in 525 patients . There were no cases of catheter sepsis (0%; 95% confidence interval {CI95}, 0%-0.6%), one possible insertion-site infection (0.2%; CI95, 0.004%-0.9%), and seven cases of phlebitis (1.1%; CI95, 0.4%-2.3%) . Catheter colonization occurred in 92 (26%) of 348 catheters cultured . Neither the per-day risk of phlebitis nor of catheter colonization increased significantly with placement >3 days . CONCLUSION: Current guidelines recommend replacement of PIV catheters in adults within 2 to 3 days; no recommendations are made for children . Our findings and those of others indicate that the overall risk of PIV catheter complications in children is extremely low and would not be reduced substantially by routine catheter replacement. J Morphol, 1999 Dec, 242(3), 283 - 94 Reexamination of hemocytes in brine shrimp (Crustacea, branchiopoda); Martin GG et al.; In 1941, a single type of hemocyte was described in the blood of the brine shrimp Artemia salina using light microscopy . This condition is unusual because most crustaceans examined using morphological, cytochemical, and functional methods have at least two types of hemoctyes . Upon examining A . franciscana, we found a single type of disk-shaped hemocyte, with a centrally located nucleus and about 15 large (6 microm diameter) granules . The granules stain for the presence of acid phosphatase and react with L-DOPA suggesting, respectively, that they are involved in degrading ingested material and possess the phenoloxidase system . Hemocytes require calcium for adhesion, bind together to mend small wounds in the body wall, and are able to phagocytose bacteria . Blood cells of A . franciscana are morphologically and functionally similar to those of the primitive chelicerate, Limulus polyphemus, and both forms have apparently given rise to more advanced taxa with multiple types of hemocytes . The major difference between the two species is the presence of the phenoloxidase system in the Crustacea and its apparent absence in the chelicerates . Vaccine, 1999 Dec 10, 18(9-10), 799 - 804 An aerosol challenge mouse model for Moraxella catarrhalis; Hu WG et al.; A simple, reproducible, and non-invasive mouse pulmonary clearance model for Moraxella catarrhalis via aerosol challenge was established . All of eight tested strains could be inoculated into mice at more than 10(5) colony-forming units (CFU)/lung with a challenge concentration of 1x10(9)-6x10(9) CFU/ml in a nebulizer . The number of bacteria retained at 6 h postchallenge was more than 10(4) CFU/lung while at 24 h postchallenge, approximate 10(3) CFU/ml or less remained in the lungs . A maximum of 100 mice could be challenged per aerosol exposure . The number of bacteria inoculated in the lungs could be adjusted by the bacterial challenge concentration, the exposure time, and the negative pressure . Lung tissue sections revealed that bacteria were evenly distributed in the lungs . Passive immunization significantly enhanced pulmonary clearance of the homologous strain in this model . These data indicate that this model will be useful for evaluating M . catarrhalis vaccine candidates and studying roles of immunity against M . catarrhalis. Biochim Biophys Acta, 1999 Dec 6, 1473(1), 35 - 53 Identification and characterization of large galactosyltransferase gene families: galactosyltransferases for all functions; Amado M et al.; Enzymatic glycosylation of proteins and lipids is an abundant and important biological process . A great diversity of oligosaccharide structures and types of glycoconjugates is found in nature, and these are synthesized by a large number of glycosyltransferases . Glycosyltransferases have high donor and acceptor substrate specificities and are in general limited to catalysis of one unique glycosidic linkage . Emerging evidence indicates that formation of many glycosidic linkages is covered by large homologous glycosyltransferase gene families, and that the existence of multiple enzyme isoforms provides a degree of redundancy as well as a higher level of regulation of the glycoforms synthesized . Here, we discuss recent cloning strategies enabling the identification of these large glycosyltransferase gene families and exemplify the implication this has for our understanding of regulation of glycosylation by discussing two galactosyltransferase gene families. FEBS Lett, 1999 Nov 26, 462(1-2), 94 - 100 The first determination of pseudouridine residues in 23S ribosomal RNA from hyperthermophilic Archaea Sulfolobus acidocaldarius; Massenet S et al.; We describe the first identification of pseudouridine (Psi) residues in ribosomal RNA (23S rRNA) of an hyperthermophilic Archaea Sulfolobus acidocaldarius . In contrast to Eucarya rRNA, only six Psi residues were detected, which is rather close to the situation in Bacteria . However, three modified positions (Psi(2479), Psi(2535) and Psi(2550)) are unique for S . acidocaldarius . Two Psi residues at positions 2060 and 2594 are universally conserved, while one other Psi (position 2066) is also common to Eucarya . Taken together the results argue against the conservation of Psi-synthases between Archaea and Bacteria and provide a basis for the search of snoRNA-like guides for Psi formation in Archaea. Gastroenterology, 1999 Dec, 117(6), 1308 - 15 Conservation of the cag pathogenicity island of Helicobacter pylori: associations with vacuolating cytotoxin allele and IS605 diversity; Slater E et al.; BACKGROUND & AIMS: Specific regions of the cag pathogenicity island (PAI) believed to enhance the virulence of Helicobacter pylori, as well as vacuolating cytotoxin gene alleles and IS605 inserts, were investigated to define diversity within infecting strain populations from patients with peptic ulcer disease and from healthy individuals . METHODS: The H . pylori studied comprised 67 isolates from 26 subjects and 14 reference strains . Specific polymerase chain reaction assays were used to test for cagA and picB in the cagI region, the virD4 homologue in the cagII region, IS605 in the genome and in the cag PAI, the "empty site" indicating absence of the cag PAI, and different vacA gene alleles . RESULTS: Most (89%) subjects were infected by H . pylori with a contiguous cag PAI . No intermediate forms were found . IS605 was not detected within the cag PAI of any strain but was present elsewhere in the genomes of strains from 62% of subjects . Twenty individuals were infected with genotypically conserved populations of H . pylori . Six subjects had mixed infections, and in 3 of these cag(+)/cag(-) variants were present . CONCLUSIONS: The cag PAI-positive H . pylori was a feature of most infected individuals, irrespective of severity of associated disease . Combined genotyping showed that 8 individuals (31%) had mixed infections, which suggests that strain population structure may be an additional contributing factor in disease development. Rev Enferm, 1999 Sep, 22(9), 633 - 5 {Enteral nutrition and the critically ill patient}; Planas M; Critically ill patients often suffer from malnutrition y loss of muscle weight throughout the whole time they are ill, even when they receive nutritional therapy, due to the tremendous amount of stress they undergo accompanied by a high degree of hypercatabolism . The most recent theories all coincide in the importance of the intestine as the preferred way for nutrients to enter the bodies of these patients because besides fulfilling its function to absorb and digest nutrients, the intestine plays an important role as a barrier to bacteria and their toxins . For these reasons, enteral nutrition should be the first option to consider whenever we must feed a critically ill patient by artificial means. Eur J Obstet Gynecol Reprod Biol, 1999 Nov, 87(1), 23 - 30 Tolerance of synthetic tissues in touch with vaginal scars: review to the point of 287 cases; Debodinance P et al.; With an experience of 287 vaginal way operations using synthetic material, the authors make a review about the tolerance of the tissues . Three tissues were used (polytetrafluoroethylene, Dacron and Lyodura) . The procedures are: Mouchel, big and small slings, Stamey and para vaginal refect procedures . At 30 months, the tolerance is 70% for Mouchel and 90% for sling procedures . The rejection rate with Dacron is globally 19.3% vs . 30.3% for Gore Tex . The authors describe materials' history, clinical symptoms and histopathologic signs of the intolerance . They think that the synthetic tissue tolerance is proportional to the exhibit surface and to the distance which separates it from the scar . The substratum of the intolerance process answers with two explanations: infection and foreign body reaction . Different theories are explained . Infection can be an ethiologic factor in early rejection . With rigid material, a small ulcer is formed and serves as a nidus for an ascending infection . Foreign material acts as an adjuvant by decreasing the number of bacteria necessary to produce an infection.The tissue reaction may be an immune response to Dacron, a delayed hypersensitivity reaction, or a graft vs . host antigen-antibody reaction . The ideal synthetic mesh material for pelvic surgery has yet to be developed. Inflamm Bowel Dis, 1999 Nov, 5(4), 285 - 94 Th1/Th2 cells; Romagnani S; A large body of evidence indicates the existence of functionally polarized CD4+ T-cell responses based on their profile of cytokine secretion . Type 1 T helper (Th1) cells produce interferon-gamma, interleukin (IL)-2, and tumour necrosis factor (TNF)-beta, which activate macrophages and are responsible for cell-mediated immunity and phagocyte-dependent protective responses . By contrast, type 2 Th (Th2) cells produce IL-4, IL-5, IL-10, and IL-13, which are responsible for strong antibody production, eosinophil activation, and inhibition of several macrophage functions, thus providing phagocyte-independent protective responses . Th1 cells mainly develop following infections by intracellular bacteria and some viruses, whereas Th2 cells predominate in response to infestations by gastrointestinal nematodes . Polarized Th1 and Th2 cells not only exhibit different functional properties, but also show the preferential expression of some activation markers and distinct transcription factors . Several mechanisms may influence the Th cell differentiation, which include the cytokine profile of "natural immunity" evoked by different offending agents, the nature of the peptide ligand, as well as the activity of some costimulatory molecules and microenvironmentally secreted hormones, in the context of the individual genetic background . In addition to playing different roles in protection, polarized Th1-type and Th2-type responses are also responsible for different types of immunopathological reactions . Th1 cells are involved in the pathogenesis of organ-specific autoimmune disorders, Crohn's disease, Helicobacter pylori-induced peptic ulcer, acute kidney allograft rejection, and unexplained recurrent abortions . In contrast, allergen-specific Th2 responses are responsible for atopic disorders in genetically susceptible individuals . Moreover, Th2 responses against still unknown antigens predominate in Omenn's syndrome, idiopathic pulmonary fibrosis, and progressive systemic sclerosis . Finally, the prevalence of Th2 responses may play some role in a more rapid evolution of human immunodeficiency virus infection to the full-blown disease . The Th1/Th2 paradigm also provides the rationale for the development of new types of vaccines against infectious agents and of novel strategies for the therapy of allergic and autoimmune disorders. Int J Pediatr Otorhinolaryngol, 1999 Oct 5, 49 Suppl 1, S275 - 8 Relationship between passive smoking, recurrent respiratory tract infections and otitis media in children; Gryczynska D et al.; The cause of upper respiratory tract infections (URTI) are multifactorial (enlarged adenoid, environmental conditions, staying at the care centers, smoking parents, allergy) . Directly, viral infection causes damage to the ciliary cells and mucociliary clearance in the nasopharynx and Eustachian tube, promotes tubal occlusion and provokes otitis media . Enlarged adenoids reduce ventilation to the nasopharynx, increase accumulation of the secretion and provide a good condition for bacteria . AIMS OF THE STUDY: Evaluation of the factors playing a role in recurrent URTI and otitis media in children . Clinical and histopathological examination of adenoid tissue of children who were passive smokers and children who were not exposed to cigarette smoke . Evaluation of the difference between ciliary-mucous transport among passive smokers and children not exposed to cigarette smoke . METHODS: The analysis of interview questionnaires in 1000 children aged 3-14 years . Histopathological examinations of adenoid tissue excised in the group of children of recurrent upper respiratory tract infections and serous otitis media exposed and not exposed to cigarette smoke . CONCLUSIONS: Among the risk factors for URTI, the most important are: (1) socio-economic conditions; (2) staying at day care centers; and (3) passive smoking . Allergy was confirmed in 35-38% of URTI children . Surgical treatment was undertaken in 11.4-32.5% of URTI children (tonsilloadenoidectomy) . Histopathological and ultrastructural evaluation of adenoid tissue in passive smoking children indicates significant differences to children not exposed to cigarette smoke. Int J Pediatr Otorhinolaryngol, 1999 Oct 5, 49 Suppl 1, S127 - 32 Waldeyer's ring and otitis media: the nasopharyngeal tonsil and otitis media; Bernstein JM; This overview of the relationship between the nasopharyngeal tonsil and otitis media will review three important concepts: (1) Adenoid inflammation leads to inflammatory obstruction of the Eustachian tube; (2) early colonization of the adenoid with the three major bacterial pathogens of otitis media is the most important factor in the early pathogenesis of otitis media; (3) the local immune system in the adenoid particularly specific secretory IgA directed against both viruses and bacterial pathogens are probably genetically controlled and represent the immunological factor in protecting the host against invasion of these agents in the Eustachian tube and middle ear . This overview of the relationship between the adenoid and the development of otitis media emphasizes that nasopharyngeal colonization with the three major middle ear pathogens is among the most important risk factors in the pathogenesis of otitis media . Inasmuch as these pathogens normally reside in the nasopharynges of most healthy children, the factors which trigger development of otitis media need to be carefully evaluated . Among these two triggers are viral infections and upper respiratory tract allergy. Dent Clin North Am, 1999 Oct, 43(4), 569 - 78, v Introduction to cariology; Clarkson BH; This article examines the discipline of cariology . A brief history is presented, and an overview of the origin and management of caries is discussed . This article frames the picture that the other authors in this issue paint. Schweiz Rundsch Med Prax, 1999 Sep 23, 88(39), 1573 - 80 {Pericardial effusion in the hospital--diagnosis and therapy}; Thummler F et al.; A pericardial effusion is a relatively common disease confronting the clinician . The most frequent causes are neoplasias (lung, breast and ovarial carcinoma, leukemia and lymphoma) uremia or idiopathic . Infections (frequently virus and seldom bacteria), myocardial infarction and rheumatic disease are also common . We present the clinical picture, the differential diagnosis and the various investigations of the pericardial effusion. Khirurgiia (Mosk), 1999, (10), 21 - 3 {Some aspects of intensive care for severe forms of anaerobic non-sporeforming infections of soft tissues}; Frantsuzov VN et al.; The results of treatment (in Hospital N.N . Burdenko) of 167 patients with anaerobic nonsporeforming ("nonclostridial") infection of soft tissues (ANIST) of various location are presented . Principal errors of intensive care in this category of patients, substantial difference in the course of infectious process in patients with ANIST depending the diseased area are shown . Programs of treatment for the patients with limited (up to 1600 cm2) and extended (over 1600 cm2) forms of ANIST are proposed . The effectiveness of such components of intensive care as ozonotherapy, correction of metabolic disturbances, HBO has been studied . Practical application of these programs allowed active influence on intoxication syndrome in ANIST which resulted in a decrease of lethality up to 10.2%. J Microbiol Methods, 2000 Jan, 39(2), 171 - 8 A simple, rapid and non-destructive procedure to extract cell wall-associated proteins from Frankia; Tavares F et al.; A simple cell fractionation procedure was developed to extract cell wall-associated proteins from the nitrogen-fixing actinomycete Frankia . The method was based on washing Frankia mycelia in 62.5 mM Tris-HCl (pH 6.8) buffer supplemented with 0.1% Triton X-100 as solubilizing agent . Cell wall-associated proteins were efficiently extracted in less than 10 min, recovering approximately 94.5+/-7.44 microg protein per extraction procedure from exponentially growing cells corresponding to 50 ml of culture . The amount of cell lysis occurring during the cell wall extraction was estimated to be 1.50+/-0.51% . Three peptidoglycan hydrolases with apparent molecular masses of 4.7, 12.1, and 17.8 kDa were detected by zymography in the cell wall-associated protein fraction . On the contrary, no cell wall lytic enzyme was detected in the cytoplasmic protein fraction . These results indicate that the present method enables a specific extraction of cell wall-associated proteins . Moreover, fluorescein isothiocyanate (FITC) labelling of the cell surface proteins showed an efficient removal of cell wall-associated proteins . Growth of the treated Frankia cells (i.e . cells from which the cell wall-associated proteins were removed) in semi-solid media suggested that these cells were still viable . This technique is of importance for functionality studies of cell wall-associated proteins, particularly for bacteria where traditional cell fractionation methods are difficult to be applied. FEMS Immunol Med Microbiol, 1999 Dec, 26(3-4), 203 - 7 Evidence for an intracellular niche for Bordetella pertussis in broncho-alveolar lavage cells of mice; Hellwig SM et al.; Bordetella pertussis can attach, invade and survive intracellularly in human macrophages in vitro . To study the significance of this bacterial feature in vivo, we analyzed the presence of viable bacteria in broncho-alveolar lavage (BAL) cells of mice infected with B . pertussis . We found B . pertussis to be present in a viable state in BAL fluid cells until at least 19 days after infection, suggesting B . pertussis to be able to survive in those cells . This intracellular niche may play an important role in the pathogenesis of pertussis . Pertussis toxin and the RGD sequence of the virulence factor filamentous hemagglutinin (FHA) both play a role in the attachment of B . pertussis to human and mouse macrophages in vitro and we hypothesized these virulence factors to be required for invasion and subsequent intracellular survival of B . pertussis in macrophages in vivo . A B . pertussis double mutant, in which the FHA RGD motif was changed to RAD and the ptx genes were deleted, was also found in a viable state in BAL fluid cells, albeit at lower levels than the wild-type strain . In our model, uptake of B . pertussis by alveolar phagocytes in vivo is thus, at least in part, determined by the bacterial virulence factors FHA and pertussis toxin. J Exp Biol, 1999 Dec, 202 Pt 24, 3541 - 53 Redox control in development and evolution: evidence from colonial hydroids Blackstone NW. Redox chemistry, involving the transfer of electrons and hydrogen atoms, is central to energy conversion in respiration, and the control of gene expression by redox state commonly occurs in bacteria, allowing rapid responses to environmental changes, for instance, in the food supply . Colonial metazoans often encrust surfaces over which the food supply varies in time or space; hence, in these organisms, redox control of the development of feeding structures and gastrovascular connections could be similarly adaptive, allowing colonies to adjust the timing and spacing of structures in response to a variable food supply . To investigate the possibility of redox control of colony development, the redox states of hydractiniid hydroid colonies were manipulated experimentally . As in many colonial animals, hydractiniid hydroids display a range of morphological variation from sheet-like forms (i.e . closely spaced polyps with high rates of stolon branching) to runner-like forms (i . e . widely spaced polyps with low rates of stolon branching) . In the runner-like Podocoryna carnea, azide, a blocker of the electron transport chain, and dinitrophenol, an uncoupler of oxidative phosphorylation, diminished the largely polyp-driven gastrovascular flow to a similar extent . Measures of the redox state of the polyp epitheliomuscular cells using the fluorescence of NAD(P)H suggest that azide shifts the redox state in the direction of reduction, while dinitrophenol shifts the redox state in the direction of oxidation . Colony development corresponds to redox state in that azide-treated colonies were more runner-like, while dinitrophenol-treated colonies were more sheet-like . Nevertheless, the functional role of polyps in feeding and generating gastrovascular flow probably contributed to a trade-off between polyp number and size such that azide-treated colonies had few large polyps, while dinitrophenol-treated colonies had many small polyps . Regardless of the treatment, P . carnea colonies developed to maturity and produced swimming medusae in the normal fashion . In the sheet-like Hydractinia symbiolongicarpus, treatment with azide resulted in complete suppression of the development of both the stolonal mat and the blastostyles, the reproductive polyps . Azide-treated H . symbiolongicarpus colonies therefore developed in a juvenilized, runner-like manner and much resembled colonies of P . carnea . Following cessation of azide treatment in H . symbiolongicarpus, normal colony development ensued, and both a stolonal mat and blastostyles formed . In both hydroid species, relative oxidization favors sheet-like growth, while relative reduction favors runner-like growth . Since feeding triggers strong contractions of polyp epitheliomuscular cells and results in relative oxidation, this experimental evidence supports the hypothesis of adaptive redox control of colony development and evolution. Biochimie, 1999 Aug-Sep, 81(8-9), 873 - 8 Cell reproduction cycle of mycoplasma; Miyata M et al.; The cell reproduction cycle of parasitic wall-free bacteria, mycoplasma, is reviewed . DNA replication of Mycoplasma capricolum starts at a fixed site neighboring the dnaA gene and proceeds to both directions after a short arrest in one direction . The initiation frequency fits to the slow speed of replication fork and DNA content is set constant . The replicated chromosomes migrate to one and three quarters of cell length before cell division to ensure delivery of the replicated DNA to daughter cells . The cell reproduction is based on binary fission but a branch is formed when DNA replication is inhibited . Mycoplasma pneumoniae has a terminal structure, designated as an attachment organelle, responsible for both host cell adhesion and gliding motility . Behavior of the organelle in a cell implies coupling of organelle formation to the cell reproduction cycle . Several proteins coded in three operons are delivered sequentially to a position neighboring the previous organelle and a nascent one is formed . One of the duplicated attachment organelles migrates to the opposite pole of the cell before cell division . It is becoming clear that mycoplasmas have specialized cell reproduction cycles adapted to the limited genome information and parasitic life. Curr Opin Pulm Med, 1999 Nov, 5(6), 365 - 70 Lung transplantation for cystic fibrosis: an update and practical considerations for referring candidates; Shapiro BJ et al.; In this article, the authors provide an update to Maurer and Chaparro's 1995 review in this journal of lung transplantation for cystic fibrosis . Bilateral (sequential) cadaver donor transplantation is the usual procedure of choice . The four-year survival rate for adult, all-disease, double-bilateral lung transplantation has improved to 53% . Because of lower {corrected} survival rate among adults, living-donor lobar transplantation should be performed only when cadaver lungs are unlikely to become available . The International Society for Heart and Lung Transplantation and the Cystic Fibrosis Foundation have promulgated uniform guidelines for transplantation candidate selection . Issues of diabetes mellitus, mechanical ventilation, osteoporosis, malnutrition, fungi and drug-resistant bacteria, pleural fibrosis, and sinusitis in relation to transplantation candidacy are discussed . Some practical points regarding transplantation center referral are presented, and a list of cystic fibrosis transplantation centers in the United States is supplied. Biochemistry, 1999 Oct 26, 38(43), 14387 - 95 Structural organization of the N-terminal domain of apolipoprotein A-I: studies of tryptophan mutants; Davidson WS et al.; Site-directed mutagenesis and detailed fluorescence studies were used to study the structure and dynamics of recombinant human proapolipoprotein (proapo) A-I in the lipid free state and in reconstituted high-density lipoprotein (rHDL) particles . Five different mutants of proapoA-I, each containing a single tryptophan residue, were produced in bacteria corresponding to each of the naturally occurring Trp residues (position -3 in the pro-segment, 8, 50, 72, and 108) in the N-terminal half of the protein . Structural analyses indicated that the conservative Phe-Trp substitutions did not perturb the conformation of the mutants with respect to the wild-type protein . Steady-state fluorescence studies indicated that all of the Trp residues exist in nonpolar environments that are highly protected from solvent in both the lipid-free and lipid-bound forms . Time-resolved lifetime and anisotropy studies indicated that the shape of the monomeric form of proapoA-I is a prolate ellipsoid with an axial ratio of about 6:1 . In addition, the region surrounding Trp 108 appears to be more mobile than the rest of the protein in the lipid-free state . However, in rHDL particles, no significant domain motion was detected for any of the Trp residues . The results presented in this work are consistent with a model for monomeric lipid-free proapoA-I in which the N-terminal half of the molecule is organized into a bundle of helices. BJU Int, 1999 Dec, 84(9), 976 - 81 Asymptomatic inflammation and/or infection in benign prostatic hyperplasia; Nickel JC et al.; OBJECTIVE: To determine the extent, pattern and clinical significance of asymptomatic histological inflammation and latent infection (National Institute of Health Category IV prostatitis) in benign prostatic hyperplasia (BPH) . PATIENTS AND METHODS: The study included 80 patients (from a cohort of 100 consecutive potentially eligible subjects) with a diagnosis of BPH, but no history or symptoms of prostatitis . Histological sections were obtained from specimens collected prospectively at transurethral resection of the prostate (TURP), immunostained for leukocyte common antigen and scanned using a computerized image-analysis system . Foci of inflammation were categorized as glandular, periglandular, stromal or peri-urethral, and the inflammatory cell density graded from 1 to 3 . Relationships and correlations were calculated between the volume, degree and type of inflammation, presence and type of bacteria (culture of deep prostatic biopsies), the use of catheters and prostate specific antigen (PSA) levels . RESULTS: Inflammation was identified in all patients but the mean tissue surface area involved was only 1.1% of the total specimen, with periglandular inflammation being the predominant pattern (0.5%) . Of the prostate specimens, 44% showed bacterial growth (in 67% of the catheterized patients and 28% of those uncatheterized; 42% of patients were catheterized before TURP) . There was no significant difference between any combination of inflammation pattern, volume or grade of inflammation in those catheterized or not (P=0.15) or culture-positive (pathogenic or not) and culture-negative cases (P=0.06) . Neither total PSA or PSA density was significantly correlated (P>0.05) with the amount, degree or distribution of inflammation . CONCLUSION: Prostatic inflammation is an extremely common histological finding in patients with symptoms of BPH who have no symptoms of prostatitis . There was no correlation between the degree and pattern of inflammation, catheterization, presence of bacteria, serum PSA or PSA density . The clinical significance of asymptomatic Category IV chronic prostatitis associated with BPH has yet to be determined. FEBS Lett, 1999 Oct 22, 460(1), 77 - 80 Mapping the cytochrome c553 interacting site using 1H and 15N NMR; Morelli X et al.; Cytochrome c553 is the electron transfer partner of formate dehydrogenase and of {Fel-hydrogenase, two metalloenzymes essential in the metabolism of sulfate reducing bacteria . These two enzymes contain a 'ferredoxin-like' domain which presents 30% identity with Desulfovibrio desulfuricans Norway ferredoxin 1 . This was chosen as a model for the 'ferredoxin-like' domain involved in the electron transfer reaction with cytochrome c553 . ID NMR titration of complex formation gave us the stoichiometry (1:1) and the dissociation constant of the complex (Kd approximately 3x10(-6) M) . 2D heteronuclear NMR experiments were performed to analyze the 1H and 15N chemical shift variations that are induced by the protein-protein recognition . This is the first mapping of the interaction site on a c-type cytochrome, using heteronuclear NMR. Biochim Biophys Acta, 1999 Oct 8, 1455(2-3), 105 - 38 Biochemical consequences of mutations causing the GM2 gangliosidoses; Mahuran DJ; The hydrolysis of GM2-ganglioside is unusual in its requirements for the correct synthesis, processing, and ultimate combination of three gene products . Whereas two of these proteins are the alpha- (HEXA gene) and beta- (HEXB) subunits of beta-hexosaminidase A, the third is a small glycolipid transport protein, the GM2 activator protein (GM2A), which acts as a substrate specific co-factor for the enzyme . A deficiency of any one of these proteins leads to storage of the ganglioside, primarily in the lysosomes of neuronal cells, and one of the three forms of GM2-gangliosidosis, Tay-Sachs disease, Sandhoff disease or the AB-variant form . Studies of the biochemical impact of naturally occurring mutations associated with the GM2 gangliosidoses on mRNA splicing and stability, and on the intracellular transport and stability of the affected protein have provided some general insights into these complex cellular mechanisms . However, such studies have revealed little in the way of structure-function information on the proteins . It appears that the detrimental effect of most mutations is not specifically on functional elements of the protein, but rather on the proteins' overall folding and/or intracellular transport . The few exceptions to this generalization are missense mutations at two codons in HEXA, causing the unique biochemical phenotype known as the B1-variant, and one codon in both the HEXB and GM2A genes . Biochemical characterization of these mutations has led to the localization of functional residues and/or domains within each of the encoded proteins. Gene, 1999 Sep 30, 238(1), 85 - 91 Centripetal modules and ancient introns; Roy SW et al.; We have created an algorithm which instantiates the centripetal definition of modules, compact regions of protein structure, as introduced by Go and Nosaka (M . Go and M . Nosaka, 1987 . Protein architecture and the origin of introns . Cold Spring Harbor Symp . Quant . Bio . 52, 915-924) . That definition seeks the minima of a function that sums the squares of C-alpha carbon distances over a window around each amino acid residue in a three-dimensional protein structure and identifies such minima with module boundari