Mikrobiologiia, 2000 Jul-Aug, 69(4), 553 - 8 {Potential activity of methane and ammonia oxidation by methanotropic communities from soda lakes of the southern Transbaikal}; Khmelenina VN et al.; Radioisotopic measurements of the methane consumption by mud samples taken from nine Southern Transbaikal soda lakes (pH 9.5-10.6) showed an intense oxidation of methane in the muds of lakes Khuzhirta, Bumalai Nur, Gorbunka, and Suduntuiskii Torom, with the maximum oxidation rate in the mud of lakes Khuzhirta (33.2 nmol/(ml day)) . The incorporation rate of the radioactive label from 14CH4 into 14CO2 was higher than into acid-stable metabolites . Optimum pH values for methane oxidation in water samples were 7-8, whereas mud samples exhibited two peaks of methane oxidation activity (at pH 8.15-9.4 and 5.8-7.0) . The majority of samples could oxidize ammonium to nitrites; the oxidation was inhibited by methane . The PCR amplification analysis of samples revealed the presence of genes encoding soluble and particulate methane monooxygenase and methanol dehydrogenase . Three alkaliphilic methanotrophic bacteria of morphotype I were isolated from mud samples in pure cultures, one of which, B5, was able to oxidize ammonium to nitrites at pH 7-11 . The data obtained suggest that methanotrophs are widely spread in the soda lakes of Southern Transbaikal, where they actively oxidize methane and ammonium. Microbes Infect, 2000 Aug, 2(10), 1285 - 92 The divergent role of tumor necrosis factor receptors in infectious diseases; Schluter D et al.; Tumor necrosis factor (TNF) receptor types 1 and 2 are broadly expressed by most cell types and are activated by binding of either TNF or lymphotoxin-beta . TNF receptor-mediated immune reactions are critically important in the pathogenesis and control of a variety of infections caused by bacteria, viruses, protozoa, and fungi . This review summarizes recent findings on the role of TNF receptors in infectious diseases and discusses the divergent functions of these receptors in immune responses. Microbes Infect, 2000 Aug, 2(10), 1207 - 14 Helicobacter pylori motility; O'Toole PW et al.; Motility is essential for Helicobacter pylori colonization . This review discusses the biochemistry, genetics and genomics of the H . pylori flagellum, and compares these features with well-characterized bacteria. Avian Dis, 2000 Jul-Sep, 44(3), 549 - 55 Ornithobacterium rhinotracheale infection in turkeys: immunoprophylaxis studies; Sprenger SJ et al.; Ornithobacterium rhinotracheale has been shown to cause serious clinical illness and is a significant concern to the turkey industry because of its potential economic impact . In this study, 6-wk-old turkeys were vaccinated intranasally with a live or subcutaneously with a killed O . rhinotracheale vaccine . At 14 or 21 wk of age, the birds were challenged intratracheally with live O . rhinotracheale . Airsacculitis and pneumonia occurred less frequently in vaccinated birds than in unvaccinated birds after challenge with O . rhinotracheale . Ornithobacterium rhinotracheale was recovered from unvaccinated, challenged birds but not from vaccinated, challenged or from unchallenged birds . Thus, turkeys inoculated with live or killed O . rhinotracheale vaccine were protected from pathologic changes. Proc R Soc Lond B Biol Sci, 2000 Aug 7, 267(1452), 1487 - 95 Evolutionary novelties in islands: Drosophila santomea, a new melanogaster sister species from São Tomé; Lachaise D et al.; The finding of new melanogaster sister species may help us in understanding more about how the emergence of genetic novelties, particularly in insular habitats, can result in speciation . Here we report on the discovery of Drosophila santomea, which is the first melanogaster sibling found off West-equatorial Africa, on Sao Tome, one of the Gulf of Guinea islands . Although the eight other melanogaster sister species are remarkably conservative in their morphology except for their terminalia, the new find has a morphological trait distinguishing it from all of these: a pure yellow body coloration of both sexes without the normal black abdominal banding . Evidence from the terminalia, polytene and mitotic chromosomes, period gene and allozymes are provided indicating that it is nonetheless the nearest relative of Drosophila yakuba with which it coexists on the island . The new find is a clear-cut taxon as shown by the production of sterile male hybrids, eventually with developmental defects, in both directions of cross with yakuba and by the existence of an altitudinal divide accompanied by a hybrid zone at mid-elevation on the island . Molecular and karyotypic data further support this conclusion . In contrast to the significant divergence of their nuclear DNAs, an intriguing similarity in their cytochrome b sequences was observed indicating a recent coalescence common to santomea, yakuba and also teissieri cytoplasms . These were shown to harbour the same Wolbachia endosymbiotic bacteria which could possibly be responsible for mitochondrial DNA hitchhiking across the species barrier. Proc Natl Acad Sci U S A, 2000 Oct 10, 97(21), 11391 - 6 Proliferation is necessary for both repair and mutation in transgenic mouse cells; Bielas JH et al.; Proliferating cells are often presumed to be more mutable than quiescent cells because they have less time to repair DNA damage before DNA replication . Direct tests of this hypothesis have been confounded by the need for cell division before a mutation can be detected . We have avoided this problem by showing that the Big Blue mouse cell line permits the dynamic quantification of both lesions and mutations in the complete absence of cell division . These cells carry the bacterial lacI gene in a lambda shuttle vector . Mutant plaques recovered by in vitro packaging of the mouse DNA can arise from mutations sustained either in mouse cells or in the bacteria . The proportion of mutant phage contained within a mutant plaque can distinguish these two types of mutation . Mutations formed in mouse cells yield >90% mutant phage because both DNA strands are mutant . On the other hand, mutations formed in the bacteria from adducted DNA yield </=50% mutant phage, because one of the DNA strands is wild type . Immediately after exposure to a test mutagen, ethylnitrosourea, all induced mutations were formed in the bacteria, but after approximately one cell division, the reverse was true and all mutations arose in the mouse cells . Only one-fifth as many mutations were recovered from quiescent cells and all arose in the bacteria, showing that the mouse cells made no mutations in the absence of proliferation . Surprisingly, the mouse cells did not repair any of the premutagenic damage during 4 days of quiescence . When these quiescent cells were induced to proliferate, however, both repair and mutation fixation ensued. J Periodontal Res, 2000 Oct, 35(5), 247 - 58 A novel closed-tube quantitative--PCR method for enumerating Porphyromonas gingivitis, Prevotella intermedia and Actinobacillus actinomycetemcomitans; Doung-udomdacha S et al.; Enumeration of specific periodontopathogens in subgingival plaque samples has been problematic because of either lack of sensitivity, specificity or the time taken to identify the organisms . These problems can be overcome using PCR, but quantification by this technique is more difficult . We report a simple quantitative PCR method developed for enumerating Porphyromonas gingivalis, Prevotella intermedia and Actinobacillus actinomycetemcomitans in clinical samples . The method relies upon inclusion of ethidium bromide in a closed-tube PCR reaction and measurement of resultant amplicons by quantifying the fluorescence generated when UV-light is passed through the walls of the amplification tube . Hence, both PCR and detection are performed in the same tube . The technique was compared with a quantitative competitive PCR and a commercial colorimetric quantitative PCR and proved to be at least as sensitive, specific and reliable for enumeration of the target bacteria . However, its speed and convenience make it particularly useful for large-scale analyses in both clinical laboratories and epidemiological studies. Verh K Acad Geneeskd Belg, 2000, 62(4), 245 - 67 {The immunological basis of the administration of DTP-polio vaccine}; Cohen H; The main subject of this essay is the analysis of the immunizing properties and possible side effects of DTP (whole cell)-polio vaccine, which since 1964 is the backbone of the immunization programme in the Netherlands . The concept, that the basis of the immunity conferred by diphtheria and tetanus toxoid and inactivated poliovirus is humoral, is now generally accepted . Assessment of the potency of these three components is based on their binding properties to specific immunoglobulines in standard-antisera . An example is the testing of neutralising antibodies against the three types of poliovirus in sera obtained from different groups of infants injected with increasing doses of this vaccine . Side-effects after injecting diphtheria- and tetanus toxoids and three types inactivated poliovaccine are negligible . For the pertussis component of the vaccine, this is not the case . The standardization of the potency of cellular pertussis vaccine is still based on the analysis of the protective capacity of a number of plain vaccine lots in the intercerebral mouse-protection test relative to the protection they confer in children . An international standard vaccine is available for this purpose, and the potency of individual lots can be expressed in International Units . At present acellular pertussis vaccines are also available for immunization against this disease . The components of such vaccines are at least for a number of components, arbitrarily chosen . Recently in a trial in Sweden, a five-component pertussis vaccine has given a similar protection of infants as a cellular vaccine of British origin . In addition this acellular vaccine induced less febrile reactions . However, acellular vaccines do not protect mice against intracerebral infection . because producers have decided to omit the "Outer Membrane Complex (OMC) as a component of the vaccine . In addition it has recently been shown that the presence of the cellular pertussis component in DPT vaccine will almost completely suppress tetanus antitoxins in the IgE fraction of the serum . This observation could indicate that this suppression is the result of a cellular immunity reaction by components in the bacteria. Biochim Biophys Acta, 2000 Jul 14, 1480(1-2), 342 - 52 The role of tryptophan residues in substrate binding to catalytic domains A and B of xylanase C from Fibrobacter succinogenes S85; McAllister KA et al.; Oxidation of the isolated catalytic domain B of xylanase C (XynC-B) from Fibrobacter succinogenes with N-bromosuccinimide (NBS) resulted in the modification of five of the seven Trp residues present in the enzyme . Hydrolytic activity of the enzyme was rapidly lost upon initiation of oxidation as a molar ratio of about two NBS molecules per molar equivalent of protein was sufficient to cause 50% inhibition of enzyme activity, and the addition of five molar equivalents of NBS resulted in less than 10% activity . Pre-incubation of XynC-B with the competitive inhibitor D-xylose resulted in the apparent protection of two Trp residues from oxidation . Xylose protection of the enzyme also resulted in a maintenance of activity, with 60% activity still evident after addition of 8-9 molar equivalents of NBS . This protection from inactivation was enhanced by the inclusion of xylohexaose in reaction mixtures . Under these conditions, however, a further Trp residue was protected from NBS oxidation . The three protected Trp residues were identified as Trp135, Trp161 and Trp202 by differential labelling and peptide mapping of NBS-oxidized preparations of the xylanase employing a combination of electrospray mass spectroscopic analysis and N-terminal sequencing . By analogy to the known structures of the family 11 xylanases, the fully conserved Trp202 residue is located on the only alpha-helix present in the enzymes, at the interface between it and the back of the beta-sheet which forms the active site cleft . Trp135 represents a highly conserved aromatic residue in family 11, but it is replaced with Thr in domain A of F . succinogenes xylanase C . To investigate the role of Trp135 in conferring the different activity profile of domain B relative to domain A, the Trp135Thr and Trp135Ala derivatives of domain B were prepared by site-directed mutagenesis . However, the kinetic parameters of the two domain B derivatives were not significantly different compared to the wild-type enzyme as reflected by K(M) and k(cat) values and product distribution profiles . Similar results were obtained with the Trp161Ala derivative of domain B, indicating that these two residues do not directly participate in the binding of substrate but likely form the foundation for binding subsite 2. Biochim Biophys Acta, 2000 Jul 24, 1492(2-3), 295 - 310 Differential developmental expression and cell type specificity of Dictyostelium catalases and their response to oxidative stress and UV-light; Garcia MX et al.; Cells of Dictyostelium discoideum are highly resistant to DNA damaging agents such as UV-light, gamma-radiation and chemicals . The genes encoding nucleotide excision repair (NER) and base excision repair (BER) enzymes are rapidly upregulated in response to UV-irradiation and DNA-damaging chemicals, suggesting that this is at least partially responsible for the resistance of this organism to these agents . Although Dictyostelium is also unusually resistant to high concentrations of H(2)O(2), little is known about the response of this organism to oxidative stress . To determine if transcriptional upregulation is a common mechanism for responding to DNA-damaging agents, we have studied the Dictyostelium catalase and Cu/Zn superoxide dismutase antioxidant enzymes . We show that there are two catalase genes and that each is differentially regulated both temporally and spatially during multicellular development . The catA gene is expressed throughout growth and development and its corresponding enzyme is maintained at a steady level . In contrast, the catB gene encodes a larger protein and is only expressed during the final stages of morphogenesis . Cell type fractionation showed that the CatB enzyme is exclusively localized to the prespore cells and the CatA enzyme is found exclusively in the prestalk cells . Each enzyme has a different subcellular localization . The unique developmental timing and cell type distribution suggest that the role for catB in cell differentiation is to protect the dormant spores from oxidative damage . We found that exposure to H(2)O(2) does not result in the induction of the catalase, superoxide dismutase, NER or BER mRNAs . A mutant with greatly reduced levels of catA mRNA and enzyme has greatly increased sensitivity to H(2)O(2) but normal sensitivity to UV . These results indicate that the natural resistance to oxidative stress is not due to an ability to rapidly raise the level of antioxidant or DNA repair enzymes and that the response to UV-light is independent from the response to reactive oxygen compounds. Biochim Biophys Acta, 2000 Aug 15, 1459(2-3), 390 - 6 Effects of nitric oxide and peroxynitrite on the cytochrome oxidase K(m) for oxygen: implications for mitochondrial pathology; Cooper CE et al.; This review summarises current knowledge about the effect of oxygen on cytochrome oxidase activity in vitro and in vivo . Cytochrome oxidase normally operates above its K(m) for oxygen in vivo . However, decreases in the intracellular oxygen concentration (hypoxia) under physiological extremes, or during pathophysiology, can cause mitochondrial respiration to become oxygen limited . Inhibitors that raise the enzyme's K(m) will induce oxygen limitation under apparently normoxic conditions . It is known that the concentrations of nitric oxide and peroxynitrite are raised in a number of pathophysiological conditions . These compounds are capable of reversibly and irreversibly raising the cytochrome oxidase K(m) for oxygen . Therefore, measurements of cell and mitochondrial respiration in vitro that fail to systematically vary oxygen through the range of physiological concentrations are likely to underestimate the effects of nitric oxide and peroxynitrite in vivo. Biochim Biophys Acta, 2000 Aug 15, 1459(2-3), 310 - 5 Catalysis in fumarate reductase; Reid GA et al.; In the absence of oxygen many bacteria are able to utilise fumarate as a terminal oxidant for respiration . In most known organisms the fumarate reductases are membrane-bound iron-sulfur flavoproteins but Shewanella species produce a soluble, periplasmic flavocytochrome c(3) that catalyses this reaction . The active sites of all fumarate reductases are clearly conserved at the structural level, indicating a common mechanism . The structures of fumarate reductases from two Shewanella species have been determined . Fumarate, succinate and a partially hydrated fumarate ligand are found in equivalent locations in different crystals, tightly bound in the active site and close to N5 of the FAD cofactor, allowing identification of amino acid residues that are involved in substrate binding and catalysis . Conversion of fumarate to succinate requires hydride transfer from FAD and protonation by an active site acid . The identity of the proton donor has been open to question but we have used structural considerations to suggest that this function is provided by an arginine side chain . We have confirmed this experimentally by analysing the effects of site-directed mutations on enzyme activity . Substitutions of Arg402 lead to a dramatic loss of activity whereas neither of the two active site histidine residues is required for catalysis. Biochim Biophys Acta, 2000 Aug 15, 1459(2-3), 274 - 83 Diversity and origin of alternative NADH:ubiquinone oxidoreductases; Kerscher SJ; Mitochondria from various organisms, especially plants, fungi and many bacteria contain so-called alternative NADH:ubiquinone oxidoreductases that catalyse the same redox reaction as respiratory chain complex I, but do not contribute to the generation of transmembrane proton gradients . In eucaryotes, these enzymes are associated with the mitochondrial inner membrane, with their NADH reaction site facing either the mitochondrial matrix (internal alternative NADH:ubiquinone oxidoreductases) or the cytoplasm (external alternative NADH:ubiquinone oxidoreductases) . Some of these enzymes also accept NADPH as substrate, some require calcium for activity . In the past few years, the characterisation of several alternative NADH:ubiquinone oxidoreductases on the DNA and on the protein level, of substrate specificities, mitochondrial import and targeting to the mitochondrial inner membrane has greatly improved our understanding of these enzymes . The present review will, with an emphasis on yeast model systems, illuminate various aspects of the biochemistry of alternative NADH:ubiquinone oxidoreductases, address recent developments and discuss some of the questions still open in the field. Biochim Biophys Acta, 2000 Aug 15, 1459(2-3), 248 - 57 Solution structure of the NADP(H)-binding component (dIII) of proton-translocating transhydrogenase from Rhodospirillum rubrum; Jeeves M et al.; Transhydrogenase is a proton pump found in the membranes of bacteria and animal mitochondria . The solution structure of the expressed, 21.5 kDa, NADP(H)-binding component (dIII) of transhydrogenase from Rhodospirillum rubrum has been solved by NMR methods . This is the first description of the structure of dIII from a bacterial source . The protein adopts a Rossmann fold: an open, twisted, parallel beta-sheet, flanked by helices . However, the binding of NADP(+) to dIII is profoundly different to that seen in other Rossmann structures, in that its orientation is reversed: the adenosine moiety interacts with the first betaalphabetaalphabeta motif, and the nicotinamide with the second . Features in the structure that might be responsible for changes in nucleotide-binding affinity during catalysis, and for interaction with other components of the enzyme, are identified . The results are compared with the recently determined, high-resolution crystal structures of human and bovine dIII which also show the reversed nucleotide orientation. An Esp Pediatr, 2000 Mar, 52(3), 245 - 50 {Experience with ventriculitis at a neonatology department}; Nieto Del Rincon N et al.; OBJECTIVE: To study the characteristics, treatment and follow-up of patients with ventriculitis in our neonatal unit . PATIENTS AND METHODS: Retrospective study of patients diagnosed with ventriculitis from January 1990 to December 1997 . Diagnostic criteria were the identification of any bacteria in the ventricular fluid and pleocytosis (> or = 100 leukocytes per microl) . Personal history, clinical and analytical findings and evolution after diagnosis were studied . RESULTS: We recorded ten cases of ventriculitis in nine patients . Six of them occurred as a complication of previous meningitis and four occurred after neurosurgical treatment . The mean age at diagnosis was of 38.8 days (range 8-130), and mean gestational age was 29.4 weeks (range 25-38) . Clinical and ventricular fluid anomalies were seen in six cases and in four the diagnosis was made at autopsy . Treatment was systemic antibiotics . In two cases intraventricular antibiotics were added . Six patients died, ventriculitis being the direct cause of death in five . Of the three survivors, one had mental retardation and cerebral palsy and the other two had minor disabilities . CONCLUSIONS: During the neonatal period, a high degree of clinical suspicion and techniques for an early diagnosis and treatment are needed for ventriculitis. Mol Cell Biol, 2000 Oct, 20(20), 7784 - 97 Mmf1p, a novel yeast mitochondrial protein conserved throughout evolution and involved in maintenance of the mitochondrial genome; Oxelmark E et al.; A novel protein family (p14.5, or YERO57c/YJGFc) highly conserved throughout evolution has recently been identified . The biological role of these proteins is not yet well characterized . Two members of the p14.5 family are present in the yeast Saccharomyces cerevisiae . In this study, we have characterized some of the biological functions of the two yeast proteins . Mmf1p is a mitochondrial matrix factor, and homologous Mmf1p factor (Hmf1p) copurifies with the soluble cytoplasmic fraction . Deltammf1 cells lose mitochondrial DNA (mtDNA) and have a decreased growth rate, while Deltahmf1 cells do not display any visible phenotype . Furthermore, we demonstrate by genetic analysis that Mmf1p does not play a direct role in replication and segregation of the mtDNA . rho(+) Deltammf1 haploid cells can be obtained when tetrads are directly dissected on medium containing a nonfermentable carbon source . Our data also indicate that Mmf1p and Hmf1p have similar biological functions in different subcellular compartments . Hmf1p, when fused with the Mmf1p leader peptide, is transported into mitochondria and is able to functionally replace Mmf1p . Moreover, we show that homologous mammalian proteins are functionally related to Mmf1p . Human p14.5 localizes in yeast mitochondria and rescues the Deltammf1-associated phenotypes . In addition, fractionation of rat liver mitochondria showed that rat p14.5, like Mmf1p, is a soluble protein of the matrix . Our study identifies a biological function for Mmf1p and furthermore indicates that this function is conserved between members of the p14.5 family. J Dairy Sci, 2000 Sep, 83(9), 2139 - 48 Whole-herd optimization with the Cornell Net Carbohydrate and Protein System . I . Predicting feed biological values for diet optimization with linear programming; Tedeschi LO et al.; We developed a diet optimizer for least-cost diet formulation with the Cornell Net Carbohydrate and Protein System (CNCPS) using linear programming . The CNCPS model is intrinsically nonlinear, and feed biological values vary with animal and feed characteristics . To allow linear diet optimization, we first used the CNCPS model to generate biological values to characterize the energy and protein content of each feed for the specific group for which the diet was being formulated . The biological values used were metabolizable energy (Mcal/kg), metabolizable protein {(% dry matter (DM)}, passage rate (%/h), bacteria yield efficiencies (g/g), and degradation rate of the carbohydrate B2 fraction (%/h) . In addition, the ruminal balances for nitrogen and peptides were included in the optimizer to optimize ruminal degradation of fiber . The objective function was to minimize diet cost subject to animal requirement and feed availability constraints . The animal constraints were set by requirements for DM intake (kg/d), metabolizable energy (Mcal/kg), metabolizable protein (%DM), and effective neutral detergent fiber (%DM) for a given level of production . Data from a dairy farm were used to evaluate this linear diet optimizer . Across all classes of dairy cattle, the CNCPS 4.0 model typically obtained a solution in less than six iterations that met the requirements with nearly 100% accuracy . We conclude this linear optimizer can be used to accurately formulate least-cost diets with the CNCPS model. Parasitol Res, 2000 Sep, 86(9), 724 - 8 Direct amplification of Entamoeba histolytica DNA from amoebic liver abscess pus using polymerase chain reaction; Zaman S et al.; An important and serious complication of intestinal infection with Entamoeba histolytica is the involvement of the liver (hepatic amoebiasis) . Hepatic amoebiasis is usually diagnosed by the clinical picture (pain in the right upper quadrant and fever), ultrasound examination and positive serology . However, none of these tests are definitive and the picture overlaps with pyogenic liver abscess caused by bacteria . It is for this reason that the feasibility of using polymerase chain reaction (PCR) for the detection of E . histolytica DNA in liver abscess pus was investigated . A comparative study was done to verify the sensitivity of ten pairs of primers specific for detecting E . histolytica in stools . Samples of liver abscess pus from 22 serology-positive patients were collected under ultrasound guidance; and these were used directly in PCR assays without any prior pre-treatment of the samples . Of the ten pairs of previously published primers tested, two pairs of primers (PI + P2 and P11 + P12) were found to give 100% sensitivity . Based on these results, we recommend that PCR assay can be successfully used to confirm the diagnosis of amoebic liver abscess with the primers identified. J Med Virol, 2000 Oct, 62(2), 140 - 3 Cytomegalovirus infection is not necessarily a poor prognostic factor in adult T-cell leukemia/lymphoma; Fujiwara H et al.; The relationship between cytomegalovirus (CMV) antigenemia and the clinical course was examined in 57 patients with adult T-cell leukemia/lymphoma (ATLL) . All patients included had the acute/lymphoma type of ATL according to the criteria of the Japan Lymphoma Study Group (LSG) . CMV antigenemia was assessed on admission and at the time when the patients had fever higher than 37 . 5 degrees C, which did not respond to antibiotics for longer than 3 days . The incidence of CMV antigenemia was 44% . Approximately 90% of patients with CMV antigenemia died of infections with viruses, bacteria, and/or fungi, while approximately 40% of patients without CMV antigenemia died of deterioration of ATLL (P<0.0001) . In this study, the patients with CMV antigenemia tended to survive longer than those negative for it (321.4 days vs . 266.2 days), although there was no statistical significance (P=0.09) . Kaplan-Meier analysis revealed that CMV antigenemia was not a poor prognostic factor . When the disease status of ATLL was evaluated by thymidine kinase (TK) and soluble interleukin 2 receptor (sIL-2R), both had lower titers during CMV antigenemia (TK: P=0.01, sIL-2R: P=0.03, respectively) . Therefore, CMV infections in ATLL patients seemed to have bimodal meanings; CMV infection at the end of clinical course were life-threatening, but infection during the first half of clinical course seemed to suppress ATLL activity and to contribute to the longer survival of the patients . Biotechnol Appl Biochem, 2000 Oct, 32 ( Pt 2), 95 - 107 Design and production of recombinant subunit vaccines; Hansson M et al.; The development of subunit vaccines is presently the main strategy being evaluated for prevention of infectious diseases . The use of recombinant-DNA techniques has facilitated the development of new principles for design and production of subunit vaccines . First of all, the properties of a target protein immunogen can be improved by the use of gene-fusion technology or by the creation of specific changes, to generate 'second-generation protein vaccines' . Properties that can be modified include protein solubility, protein stability, in vivo half-lives, etc . In addition, for subunit protein vaccine candidates, the immunogenic properties can be significantly augmented by the addition of immunopotentiating tags or by means of targeting to immunoreactive sites . The recombinant subunit vaccine can furthermore be adapted by gene-fusion technology, to be efficiently incorporated into immunopotentiating adjuvant systems . Also in passive vaccination strategies, i.e . the use of antibodies or antibody fragments for prevention of infectious diseases, the recombinant strategies have become increasingly important . Humanized antibodies and antibody fusion proteins represent common present anti-infectious-disease agents . The selected examples will indicate that recombinant strategies will indeed have an impact on the design, selection and production of recombinant proteins to be used in the prevention of infectious diseases. J Nucl Med Technol, 2000 Sep, 28(3), 162 - 4 The impact of brushing teeth on carbon-14 urea breath test results; Higazy E et al.; OBJECTIVE: The 14C urea breath test noninvasively detects the presence of the urease-producing bacteria Helicobacter pylori in the stomach . Several sources of errors have been identified to cause false or indeterminate results on the test . The objective of this study was to identify whether brushing teeth affects the test results . METHODS: We performed the 14C urea breath test on 168 patients, with breath samples counted at 10 and 20 min after oral administration of 2 microCi (74 kBq) 14C urea . Ninety-four patients brushed their teeth before the test while 74 did not . RESULTS: Thirty-six of the 74 patients (49%) who did not brush their teeth had positive results at 10 min, which became negative at 20 min . None of the 94 patients who brushed their teeth before testing showed this pattern with agreement of results at 10 and 20 min . CONCLUSION: We recommend brushing teeth before the 14C urea breath test since it significantly decreased the ambiguous results of the test in our laboratory. J Biotechnol, 2000 Sep 29, 83(1-2), 77 - 84 How CD4(+) T cells may eliminate extracellular gastric Helicobacter? Aebischer T, Lucas B, Koesling J, Bumann D, Meyer TF. Helicobacter pylori is recognised as a causal agent in the pathogenesis of gastritis, gastric and duodenal ulcer disease as well as gastric cancers . Eradication of the bacteria with antibiotics is currently used to treat symptomatic, infected individuals . Theoretically the infection could also be controlled by vaccination . Several immunisation protocols were developed in small animal models and primates in order to validate this approach . Recently making use of mice with defined genetic defects, H . pylori-specific CD4(+) T cells were found to be crucial for protective vaccination . This was unexpected and poses the question of how activation of CD4(+) T cells leads to the elimination of bacteria that reside primarily in the mucin layer behind a barrier of epithelial cells . CD4(+) T cells fulfil their effector function by secreting lymphokines and by engaging specific surface ligands on interacting cells . Here we propose that phagocytes and epithelial cells stimulated either by direct interaction with CD4(+) T cells or by soluble mediators such as cytokines or neuropeptides are the ultimate effector populations in protective immunity induced by vaccination. J Biotechnol, 2000 Sep 29, 83(1-2), 51 - 5 The porcine gastrointestinal lamina propria: an appropriate target for mucosal immunisation? Stokes CR, Bailey M. During the course of a lifetime, it has been calculated that we may consume between 100 and 700 tons of food . For the average British citizen, this is likely to include some 550 poultry, 36 pigs, 36 sheep, eight oxen, 10000 eggs and dairy products (milk, butter cheese, etc.) equivalent to 18 tonnes of milk . As if that were not sufficient enough a challenge, the homeostasis within the intestine is further complicated by the presence of 10(5)-10(11) bacteria (pathogenic and non-pathogenic) per gram of mucus and the constant turnover of gut epithelial cells . Given such a magnitude of challenge, which, at least in health is heavily biased in favour of harmless antigens, it can be reasonably hypothesised that the default response of the intestinal mucosal immune system would appear to be set heavily in favour of non-responsiveness and oral tolerance . The purpose of this review is to briefly describe recent progress from studies of the pig that support this hypothesis and to discuss the implications for future mucosal vaccine design. Ann Genet, 2000 Apr-Jun, 43(2), 69 - 74 Human TIP49b/RUVBL2 gene: genomic structure, expression pattern, physical link to the human CGB/LHB gene cluster on chromosome 19q13.3; Parfait B et al.; Bacterial DNA helicase RuvB protein is an essential component in homologous recombination and DNA double-strand break repair . Here, we report the gene structure of TIP49b/RUVBL2, a second putative human homologue of the bacterial RuvB gene . This gene contains 15 exons and 14 introns . The TIP49b/RUVBL2 open reading frame encodes a protein of 463 amino acids, showing 43% identity with the RUVBL1 protein . The TIP49b/RUVBL2 gene is physically linked to the human CGB/LHB gene cluster on chromosome 19q13.3 . Genomic sequence analysis revealed that the TIP49b/RUVBL2 gene is very close (55 nucleotides in length) to the LHB gene, in the opposite orientation . The very close co-location of the mouse homologues of the human TIP49b/RUVBL2 and LHB genes was also conserved on mouse chromosome 7 . Co-ordinated transcriptional regulation between the TIP49b/RUVBL2 and LHB genes was not observed . TIP49b/RUVBL2, like RUVBL1, was expressed ubiquitously in all human tissues examined and more strongly in testis . As TIP49b/RUVBL2 is expected to be involved in recombination repair and is located in a chromosome region frequently amplified in breast cancer, we quantified TIP49b/RUVBL2 gene expression by using real-time quantitative RT-PCR in a series of breast tumour samples . None of the tumour samples showed an altered TIP49b/RUVBL2 transcription level relative to normal breast tissue. Emerg Infect Dis, 2000 Sep-Oct, 6(5), 449 - 57 Antigenic variation in vector-borne pathogens; Barbour AG et al.; Several pathogens of humans and domestic animals depend on hematophagous arthropods to transmit them from one vertebrate reservoir host to another and maintain them in an environment . These pathogens use antigenic variation to prolong their circulation in the blood and thus increase the likelihood of transmission . By convergent evolution, bacterial and protozoal vector-borne pathogens have acquired similar genetic mechanisms for successful antigenic variation . Borrelia spp . and Anaplasma marginale (among bacteria) and African trypanosomes, Plasmodium falciparum, and Babesia bovis (among parasites) are examples of pathogens using these mechanisms . Antigenic variation poses a challenge in the development of vaccines against vector-borne pathogens. Biochem J, 2000 Oct 1, 351(Pt 1), 281 - 8 Cloning and functional expression of a gene encoding a vacuolar-type proton-translocating pyrophosphatase from Trypanosoma cruzi; Hill JE et al.; Acidocalcisomes are acidic Ca(2+)-storage organelles found in trypanosomatids that are similar to organelles known historically as volutin granules . Acidification of these organelles is driven in part by a vacuolar H(+)-pyrophosphatase (V-H(+)-PPase), an enzyme that is also present in plant vacuoles and in some bacteria . Here, we report the cloning and sequencing of a gene encoding the acidocalcisomal V-H(+)-PPase of Trypanosoma cruzi . The protein (T . cruzi pyrophosphatase, TcPPase) predicted from the nucleotide sequence of the gene has 816 amino acids and a molecular mass of 85 kDa . Several sequence motifs found in plant V-H(+)-PPases were present in TcPPase, explaining its sensitivity to N-ethylmaleimide and N,N'-dicyclohexylcarbodi-imide . Heterologous expression of the cDNA encoding TcPPase in the yeast Saccharomyces cerevisiae produced a functional enzyme . Phylogenetic analysis of the available V-H(+)-PPase sequences indicates that TcPPase is nearer to the vascular plant cluster and the branch containing Chara, a precursor to land plants, than to any of the other pyrophosphatase sequences included in the analysis . The apparent lack of such a V-H(+)-PPase in mammalian cells may provide a target for the development of new drugs. Anal Biochem, 2000 Oct 1, 285(1), 113 - 20 Selective extraction and purification of a mycobacterial outer membrane protein; Heinz C et al.; MspA forms water-filled channels in the mycolic acid layer of Mycobacterium smegmatis thereby allowing the diffusion of hydrophilic solutes through this permeability barrier into the periplasm . MspA is the first member of a new family of porins and is extremely stable against chemical and thermal denaturation . We developed a purification procedure based on selective extraction of MspA with detergents from whole cells of M . smegmatis at high temperatures . Anion-exchange and size-exclusion chromatography yielded about 230 microg apparently pure and highly active MspA per liter of culture . This was a 20-fold increased yield compared to previous purification protocols . Similar amounts of pure MspA were obtained with the detergents isotridecylpolyethyleneglycolether, lauryldimethylamine oxide, and octylpolyethylene oxide indicating that this purification procedure is not restricted to a specific detergent . This study will promote the structural and functional analysis of MspA and might be valuable for the isolation of porins from other mycolic acid-containing bacteria . Mol Microbiol, 2000 Sep, 37(6), 1470 - 9 Interdependence of environmental factors influencing reciprocal patterns of gene expression in virulent Borrelia burgdorferi; Yang X et al.; The paradigm for differential antigen expression in Borrelia burgdorferi, the agent of Lyme disease, is the reciprocal expression of its outer surface (lipo)proteins (Osp) A and C; as B . burgdorferi transitions from its arthropod vector into mammalian tissue, ospC is upregulated, and ospA is downregulated . In the current study, using B . burgdorferi cultivated under varying conditions in BSK-H medium, we found that a decrease in pH, in conjunction with increases in temperature (e.g . 34 degrees C or 37 degrees C) and cell density, acted interdependently for the reciprocal expression of ospC and ospA . The lower pH (6.8), which induced the reciprocal expression of ospC and ospA in BSK-H medium, correlated with a drop in pH from 7.4 to 6.8 of tick midgut contents during tick feeding . In addition to ospC and ospA, other genes were found to be regulated in reciprocal fashion . Such genes were either ospC-like (e.g . ospF, mlp-8 and rpoS) (group I) or ospA-like (lp6.6 and p22) (group II); changes in expression occurred at the mRNA level . That the expression of rpoS, encoding a putative stress-related alternative sigma factor (sigma(s)), was ospC-like suggested that the expression of some of the group I genes may be controlled through sigma(s) . The combined results prompt a model that allows for predicting the regulation of other B . burgdorferi genes that may be involved in spirochaete transmission, virulence or mammalian host immune responses. Structure Fold Des, 2000 Aug 15, 8(8), 897 - 904 Crystal structure of archaeal RNase HII: a homologue of human major RNase H; Lai L et al.; BACKGROUND: RNases H are present in all organisms and cleave RNAs in RNA/DNA hybrids . There are two major types of RNases H that have little similarity in sequence, size and specificity . The structure of RNase HI, the smaller enzyme and most abundant in bacteria, has been extensively studied . However, no structural information is available for the larger RNase H, which is most abundant in eukaryotes and archaea . Mammalian RNase H participates in DNA replication, removal of the Okazaki fragments and possibly DNA repair . RESULTS: The crystal structure of RNase HII from the hypothermophile Methanococcus jannaschii, which is homologous to mammalian RNase H, was solved using a multiwavelength anomalous dispersion (MAD) phasing method at 2 A resolution . The structure contains two compact domains . Despite the absence of sequence similarity, the large N-terminal domain shares a similar fold with the RNase HI of bacteria . The active site of RNase HII contains three aspartates: Asp7, Asp112 and Asp149 . The nucleotide-binding site is located in the cleft between the N-terminal and C-terminal domains . CONCLUSIONS: Despite a lack of any detectable similarity in primary structure, RNase HII shares a similar structural domain with RNase HI, suggesting that the two classes of RNases H have a common catalytic mechanism and possibly a common evolutionary origin . The involvement of the unique C-terminal domain in substrate recognition explains the different reaction specificity observed between the two classes of RNase H. EXS, 2000, 89, 227 - 37 Delivery systems and adjuvants for vaccination against HIV; Velin D et al.; Epidemiological studies have revealed that HIV-1 infections occur through contact with contaminated blood or during unprotected vaginal or anal intercourse . Hence, to protect against HIV infection, vaccines should ideally induce both mucosal and systemic immune responses . We present a brief review of the different delivery systems and adjuvants which can be used to elicit mucosal immune responses . Oral or nasal administration of recombinant attenuated bacteria or viruses can induce both mucosal and systemic immune responses against the carried antigen . The oral delivery of mucosal adjuvants (such as cholera toxin) in association with antigens has been shown to enhance mucosal and systemic immune responses against them . Recently developed vaccination strategies using naked DNA or other antigen delivery systems are also discussed. Biotechniques, 2000 Sep, 29(3), 602 - 4, 606-8, 609 Assessing the binding and endocytosis activity of cellular receptors using GFP-ligand fusions; Medina-Kauwe LK et al.; We have developed a simple scheme for characterizing ligand-receptor binding and post-binding activity on living cells . Our approach makes use of green fluorescent protein (GFP) as an auto-fluorescent tag to label protein ligands . We have constructed GFP-tagged ligands that can be expressed in bacteria as soluble fusion proteins . A cell-binding assay using fluorescence-activated cell sorting (FACS) demonstrates that GFP-tagged proteins retain their wild-type receptor-binding specificity . Using this assay, we measure ligand binding on unfixed cells and demonstrate receptor specificity using specific competitors . To determine the ability of receptor targets to internalize, we developed a second FACS-based assay to detect the rate and percentage of internalized ligand in living cells . Noninternalizing control ligands and fluorescence microscopy of treated cells confirm that our assay is reliable for determining receptor internalization activity. Med Clin (Barc), 2000 Jul 1, 115(5), 176 - 7 {Analysis using ELISA test of antibody response to Fusobacterium nucleatum and Eikenella corrodens in subjects with periodontal disease}; Gutierrez J et al.; BACKGROUND: To know the synthesis of IgG, IgA, and IgM to Fusobacterium nucleatum and Eikenella corrodens in serum, crevicular liquid and saliva in subjects with periodontal disease, using ELISA test . PATIENTS AND METHODS: We studied 26 patients with high bias to the disease aged less than 35 years and 30 individuals with comparable age and with scarce bias to the disease . RESULTS: No differences were found for IgG and IgM titers between the groups, except for IgM to E . corrodens . For IgA, differences were found when the saliva and crevicular liquid were studied for both bacteria, as occurred in serum IgA compared to E . corrodens . In all the cases antibody levels were lower in the patients . CONCLUSIONS: The lower synthesis of IgM and IgA, fundamentally the latter, to F . nucleatum and E . corrodens in patients with periodontal disease would contribute to the pathogenesis of this illness. Comp Biochem Physiol A Mol Integr Physiol, 2000 Sep, 127(1), 1 - 20 Silk produced by hornets: thermophotovoltaic properties-a review; Kirshboim S et al.; This article deals with the silk weave produced by pupating larvae of the Oriental hornet and its electric properties . Larvae of this hornet commence pupation at approximately 2 weeks of age . Creation of the cocoonal silk weave requires a number of hours and the encased pupa remains in the cocoon for approximately 2 more weeks before ecloding as an adult . The silk weave is initially of a creamish white color, but gradually becomes brown-gray owing to the activity of certain bacteria secreted in the silk . The silk weave is composed of fibers arranged in multiple layers with interposed surfaces occupying a considerable part of the area and containing pockets of bacteria . The spun silk contains both metallic and non-metallic elements, mostly K and Cl but also Mg, P, S, Ca, Ti and V . Shaped as a dome, the silk projects considerably beyond the cell proper, contributing importantly to its total volume and providing a shield for the contained pupa against predators, parasites, or extreme changes in temperature, as well as affording a 'sterile and clean room' in which the pupa can form its new cuticle without the interference of contaminating dust particles or the turbulence of air currents . The silk is endowed with electric properties . Inter alia, a thermoelectric phenomenon was observed in the dark, namely, upon increase in temperature the current rose to several hundred nano Amperes (nA); in light, a photovoltaic effect was observed involving voltages of several dozen millivolts (mV), with a sharp transition between the current and voltage during transition from darkness to light . Also recorded was a very high electric capacitance, amounting to scores of milli farads (mF) . In all, the pupal silk behaves like an organic semiconductor, in that its electric properties are temperature-dependent, and it also displays ferroelectric properties . Additionally, a luminescence phenomenon was recorded on the silk, wherein excitation at wavelengths within the UV(i.e . 249, 290 and 312 nm) range yielded an emission spectrum at a wavelength of 450 and of 530 nm . The silk caps are anisotropic in that the emission from the outside is lower than that from the inside . By way of recap, the various mentioned properties of the pupal silk are discussed from their biological and physical aspects. Immunopharmacology, 2000 Sep, 49(3), 419 - 23 Interleukin 2 maintains biologic stability and sterility over prolonged time; Safar M et al.; The FDA approved interleukin 2 (IL2) for clinical use in 1992 in a high-dose bolus intravenous infusion schedule . IL2 administered by continuous low- and intermediate-dose infusion can result in a variety of immunologic effects including the expansion of the Natural Killer (NK) cell pool and immune reconstitution in immune-deficient hosts . These immune modifications are essential for augmentation of both currently available and evolving immunotherapies . The manufacturer's data indicate stability of the IL2 for a period of 6 days . This time frame is not practical for prolonged infusional schemes necessitating frequent changes of drug depots . We tested the biologic stability and sterility of the commercially available recombinant IL2 preparation (aldesleukin; Proleukin, Chiron) under clinical conditions for up to 30 days . Our results confirm that IL2 retains its biologic activity and sterility under these conditions for prolonged periods . This information will simplify IL2 outpatient regimens, allowing for convenient intervals for drug depot renewal, leading to improved patient compliance and conserved health care expenditures. Genomics, 2000 Sep 15, 68(3), 343 - 7 Tissue-specific splicing of Omi stress-regulated endoprotease leads to an inactive protease with a modified PDZ motif; Faccio L et al.; Omi is a human serine protease whose catalytic domain is homologous to a bacterial heat shock endoprotease (HtrA), a protein indispensable to the survival of bacteria at elevated temperatures . Omi is expressed ubiquitously, and its protein product is predominantly localized in the endoplasmic reticulum of mammalian cells . Here we present the genomic structure of Omi, consisting of eight exons located on human chromosome 2p12-p13 . Furthermore, we describe an alternatively splice form of Omi (D-Omi) that is expressed predominantly in the kidney, colon, and thyroid . D-Omi lacks peptide sequence encoded by two exons (exons III and VII) . The absence of exon VII leads to a protein with a modified PDZ domain unable to interact with a known partner, the Mxi2 protein . The absence of exon III affects the catalytic domain and leads to a protein with no detectable protease activity . Our studies suggest that D-Omi may have a unique role in the normal function of kidney, colon, and thyroid . Proc Natl Acad Sci U S A, 2000 Sep 26, 97(20), 10899 - 904 Borrelia burgdorferi periplasmic flagella have both skeletal and motility functions; Motaleb MA et al.; Bacterial shape usually is dictated by the peptidoglycan layer of the cell wall . In this paper, we show that the morphology of the Lyme disease spirochete Borrelia burgdorferi is the result of a complex interaction between the cell cylinder and the internal periplasmic flagella . B . burgdorferi has a bundle of 7-11 helically shaped periplasmic flagella attached at each end of the cell cylinder and has a flat-wave cell morphology . Backward moving, propagating waves enable these bacteria to swim in both low viscosity media and highly viscous gel-like media . Using targeted mutagenesis, we inactivated the gene encoding the major periplasmic flagellar filament protein FlaB . The resulting flaB mutants not only were nonmotile, but were rod-shaped . Western blot analysis indicated that FlaB was no longer synthesized, and electron microscopy revealed that the mutants were completely deficient in periplasmic flagella . Wild-type cells poisoned with the protonophore carbonyl cyanide-m-chlorophenylhydrazone retained their flat-wave morphology, indicating that the periplasmic flagella do not need to be energized for the cell to maintain this shape . Our results indicate that the periplasmic flagella of B . burgdorferi have a skeletal function . These organelles dynamically interact with the rod-shaped cell cylinder to enable the cell to swim, and to confer in part its flat-wave morphology. Am J Reprod Immunol, 2000 Aug, 44(2), 73 - 9 Infection of polarized primary epithelial cells from rat uterus with Chlamydia trachomatis: cell-cell interaction and cytokine secretion; Kaushic C et al.; PROBLEM: The objective of this study was to examine the susceptibility of rat uterine epithelial cells (UEC) to infection with Chlamydia trachomatis and to study the epithelial-stromal interactions following infection . METHOD OF STUDY: UEC were isolated from adult rats and grown in culture . Polarized, confluent monolayers of UEC were infected with 10(6) IFU/well C . trachomatis (MoPn) . In order to confirm infection, MoPn was labeled with a fluorescent tracking dye, PKH-26, and then used in epithelial cell infections . Transepithelial resistances were measured prior to and following infection to test the effect of Chlamydia on the integrity of the epithelial monolayers . In other experiments, polarized epithelial cultures were infected in the presence and absence of stromal cells . Media was collected from the apical and basolateral compartments of the cultures before and after infection and analyzed for cytokines IL-1alpha and TNF-alpha . RESULTS: Epithelial cell cultures infected with PKH-26 labeled MoPn were examined 4-5 days later . Bacterial inclusions were detected inside epithelial cells indicating infection had occurred . Co-localization of PKH-26 labeled bacteria with FITC-labelled anti-Chlamydia antibody on the epithelial cells confirmed infection . No changes were found in resistance across the monolayers of epithelial cells in the presence or absence of infection . ELISA results indicate that UEC secrete IL-1alpha constitutively in citro . Stromal cells secrete very little IL-1alpha . When stromal cells were co-incubated with epithelial cells there was a decrease in the amount of IL-1alpha secreted by epithelial cells 48 hr post-infection . On the other hand, maximum TNF-alpha was found in stromal cells . both with and without infection . Epithelial cells, in these studies made very little TNF-alpha . CONCLUSIONS: These results show that primary rat epithelial cells can be infected with Chlamydia in vitro . Epithelial and stromal cells from uteri of adult rats make IL-1alpha and TNF-alpha in vitro both prior to and following infection with Chlamydia . This system can be used to analyze the role played by epithelial-stromal interactions in providing protection on this mucosal surface. Prikl Biokhim Mikrobiol, 2000 Jul-Aug, 36(4), 484 - 7 {Granulated preparations of azotobacter in a clay-mineral base}; Kirdish IK et al.; Interaction of Azotobacter chroococcum 20 cells with clay minerals increased their viability at supraoptimal temperatures . Therefore, clay minerals were used to develop granular bacterial preparations with high viable cell counts and stable compositions during long-term storage . The titers of viable bacteria in the preparations remained 60-70% of the initial level after 12-month storage. J Pharm Pharm Sci, 2000 May-Aug, 3(2), 228 - 33 Pharmaceutical interference with the {14C} carbon urea breath test for the detection of Helicobacter pylori infection; Abrams DN et al.; Helicobacter pylori bacteria reside in the mucosal lining of the stomach where, due to a variety of factors, the infection predisposes patients to peptic ulcer disease . Detection of H . pylori is important in the treatment and follow-up of patients with peptic ulcer disease and the urea breath test is the method of choice . This article will briefly review the methods for diagnosing H . pylori, emphasizing the {(14)C}urea breath test . The agents which can interfere with the results of the breath test will be reviewed and the role of the consulting pharmacist will be discussed. J Mol Biol, 2000 Sep 29, 302(4), 761 - 75 Analysis of a cell-cycle promoter bound by a response regulator; Ouimet MC et al.; Caulobacter crescentus CtrA protein, an OmpR-type response regulator, receives cell-cycle signals and binds the proposed consensus TTAA-N7-TTAA present inside the chromosome replication origin and cell-cycle transcription promoters . We synthesized a 42 bp cell-cycle promoter based on this consensus and elements of the fliL promoter . Over 100 promoter mutations were assayed for transcription directed by CtrA . Although both CtrA binding half-sites cooperate and the N7 spacing is critical for transcription, the upstream half-site is relatively flexible . The downstream CtrA half-site is less flexible and more important for cell-cycle regulation . A CtrA binding site and a -10 promoter element are sufficient for cell-cycle transcription, and both sequences cooperate and compensate for respective defects . Mutations in the CtrA binding site, but not in the -10 promoter sequence, perturb cell-cycle transcription . A single base-pair change switches a cell-cycle promoter into a strong conventional promoter . We propose rules for CtrA binding and promoter interactions implying how CtrA evolved into a versatile regulator of cell-cycle functions including flagellar biogenesis, cell division, DNA methylation, and chromosome replication . Biosci Biotechnol Biochem, 2000 Aug, 64(8), 1771 - 3 Preliminary communication . MRNA expression of MT1-MMP, MMP-9, cathepsin K, and TRAP in highly enriched osteoclasts cultured on several matrix proteins and ivory surfaces; Uemura T et al.; We demonstrated that the transcriptional expression of MT1-MMP, MMP-9, cathepsin K, and TRAP in highly enriched osteoclasts were regulated by different matrix proteins that bind to integrin on osteoclast, such as collagen type I (CoI), fibronectin (FN), vitronectin (VN), osteopontin (OPN), and ivory . Results suggested that the OPN-integrin alphavbeta3 binding plays a more important role than CoI-alpha2beta1 binding in the regulation of osteoclast activity. Nature, 2000 Sep 7, 407(6800), 78 - 80 Uptake of dissolved organic carbon and trace elements by zebra mussels; Roditi HA et al.; Zebra mussels (Dreissena polymorpha) are widespread and abundant in major freshwater ecosystems in North America, even though the phytoplankton food resources in some of these systems seem to be too low to sustain them . Because phytoplankton biomass is greatly depleted in ecosystems with large D . polymorpha populations and bacteria do not seem to be an important food source for this species, exploitation of alternative carbon sources may explain the unexpected success of D . polymorpha in such environments . Here we examine the possibility that absorption of dissolved organic carbon (DOC) from water could provide a nutritional supplement to zebra mussels . We find that mussels absorb 14C-labelled DOC produced by cultured diatoms with an efficiency of 0.23%; this indicates that DOC in natural waters could contribute up to 50% of the carbon demand of zebra mussels . We also find that zebra mussels absorb some dissolved metals that have been complexed by the DOM; although absorption of dissolved selenium was unaffected by DOC, absorption of dissolved cadmium, silver and mercury by the mussels increased 32-, 8.7- and 3.6-fold, respectively, in the presence of high-molecular-weight DOC. Rev Med Liege, 2000 Jun, 55(6), 527 - 30 {Bites by terrestrial vertebrates}; Henry F et al.; Bites by terrestrial vertebrates, reptiles or mammals, represent a special risk in tropical regions . Envenomation is possible by a few lizards and many snakes . For mammals, tissular destructions due to the bite can be severe . Whatever is the offending animal, bites can further become infected by transmitted viruses or bacteria. Gac Med Mex, 2000 Jul-Aug, 136(4), 329 - 34 {Characteristics of peritoneal fluid in children with severe diarrhea}; Gomez-Alcala AV; BACKGROUND: A significant number of acute diarrheal diseases produce severe damage to the bowel wall that can lead the patient to death . Peritoneal fluid (PF) analysis has been proposed as a tool to establish an early diagnosis of these injuries . MATERIAL AND METHOD: Thirty patients with a diagnosis of acute diarrhea and suspected enteric perforation or gangrene were submitted to paracentesis . LP was classified according to its appearance immediately after obtention by the surgeon, and determinations of leukocytes, proteins, glucose, and chloride were done . Clinical diagnosis was established during laparotomy or necropsy, or after an uneventual recovery . Data were analyzed by means of diagnostic test statistics . RESULTS: Clear and yellow PF was considered a sign of the absence of enteric perforation or gangrene, with a sensitivity (S) of 87%, specificity (SP) and positive predictive value (PPV) of 100%, and a negative predictive value (NPV) of 91.6%, with a predictive accuracy (PA) of 95% . A bloody PF was considered a sign of enteric gangrene, with an S of 54%, SP and PPV of 100%, and NPV of 72% (PA 80%) . More than 1000 leukocytes per mm3 of PF were considered an sign of some kind of macroscopic injury of the bowel wall, with an S of 68%, a SP of 88%, a PPV of 91% and a NPV of 61% (PA 76%) . PF glucose level less than 40 mg/dl was considered a sign of enteric gangrene (S 42%, SP 78%, PPV 71%, NPV 50%, with a PA of 57%), and also of bacteria viability in PF cultures (S and NPV 100%, SP 80%, PPV 67% and PA of 86%) . Chloride levels greater than 90 meq/L were considered a sign of enteric perforation or gangrene, with an S and NPV of 100%, a SP of 60%, and a PPV de 82% (PA 86%) . CONCLUSION: PF analysis is of great value in the evaluation of the child with acute severe diarrhea in which enteric perforation or gangrene is suspected . A clear yellow fluid with less than 1000 leukocytes per mm3, more than 40 mg/dL of glucose, and less than 90 meq/L of chloride suggest that enteritis has not caused irreversible injury to the bowel wall. Infect Immun, 2000 Oct, 68(10), 6069 - 72 Differential effects of virulent versus avirulent Legionella pneumophila on chemokine gene expression in murine alveolar macrophages determined by cDNA expression array technique; Nakachi N et al.; The cDNA expression array technique is a powerful tool to determine, at one time from many genes, specific gene messages modulated by infection . In the present study, we identified genes modulated in response to virulent versus avirulent Legionella pneumophila infection of the alveolar macrophage cell line MH-S by the cDNA expression array technique . Many macrophage genes were found to be modulated after 5 h of in vitro infection with L . pneumophila . In particular, it was found that the monocyte chemotactic protein 3 (MCP-3) gene expression was significantly induced by infection with virulent L . pneumophila but not with avirulent L . pneumophila . In contrast, other chemokine genes, such as macrophage inflammatory protein (MIP) 1alpha, were induced by both virulent and avirulent L . pneumophila . Reverse transcription (RT)-PCR assay of total RNA isolated from macrophages infected with the bacteria for 5 or 24 h confirmed the differential induction of the chemokine genes by virulent versus avirulent L . pneumophila . Thus, the cDNA expression array technique readily revealed differential induction by L . pneumophila infection of select chemokine genes of macrophages from more than 1,100 genes . These results also indicate that certain chemokine genes may be selectively induced by virulent bacteria. Infect Immun, 2000 Oct, 68(10), 5960 - 9 Infection of human endothelial cells with Bartonella bacilliformis is dependent on Rho and results in activation of Rho; Verma A et al.; Bartonella bacilliformis was continuously internalized into human endothelial cells beginning shortly after addition of the bacteria and continuing for at least 24 h after infection in vitro, with a major increase in uptake occurring between 16 and 24 h . Preincubation of endothelial cells with C3 exoenzyme, which inactivated intracellular Rho-GTPase, blocked internalization of the bacteria . Addition of C3 exoenzyme at any time after addition of the bacteria blocked further internalization of bacteria, including the major uptake of bacteria internalized at 16 to 24 h . Rho, a key signaling protein in pathways involving actin organization, was directly shown to be activated in endothelial cells undergoing infection with B . bacilliformis, with maximal activation and translocation to the plasma membrane at 12 to 16 h . At late times of infection, most of the bacteria were found in a perinuclear location . Staining of the Golgi complex with specific markers, anti-human Golgin-97, anti-KDEL receptor, and BODIPY-TR ceramide, showed colocalization of bacteria in the Golgi complex region . Disruption of the Golgi complex with brefeldin A scattered the bacteria from this perinuclear location and resulted in inhibition of internalization of the bacteria in endothelial cells. Infect Immun, 2000 Oct, 68(10), 5575 - 80 Construction and characterization of a Mycobacterium tuberculosis mutant lacking the alternate sigma factor gene, sigF; Chen P et al.; The alternate RNA polymerase sigma factor gene, sigF, which is expressed in stationary phase and under stress conditions in vitro, has been deleted in the virulent CDC1551 strain of Mycobacterium tuberculosis . The growth rate of the DeltasigF mutant was identical to that of the isogenic wild-type strain in exponential phase, although in stationary phase the mutant achieved a higher density than the wild type . The mutant showed increased susceptibility to rifampin and rifapentine . Additionally, the DeltasigF mutant displayed diminished uptake of chenodeoxycholate, and this effect was reversed by complementation with a wild-type sigF gene . No differences in short-term intracellular growth between mutant and wild-type organisms within human monocytes were observed . Similarly, the organisms did not differ in their susceptibilities to lymphocyte-mediated inhibition of intracellular growth . However, mice infected with the DeltasigF mutant showed a median time to death of 246 days compared with 161 days for wild-type strain-infected animals (P < 0.001) . These data indicate that M . tuberculosis sigF is a nonessential alternate sigma factor both in axenic culture and for survival in macrophages in vitro . While the DeltasigF mutant produces a lethal infection of mice, it is less virulent than its wild-type counterpart by time-to-death analysis. J Biol Chem, 2000 Dec 8, 275(49), 38254 - 60 H2O2-sensitive fur-like repressor CatR regulating the major catalase gene in Streptomyces coelicolor; Hahn JS et al.; Streptomyces coelicolor produces three distinct catalases to cope with oxidative and osmotic stresses and allow proper growth and differentiation . The major vegetative catalase A (CatA) is induced by H(2)O(2) and is required for efficient aerobic growth . In order to investigate the H(2)O(2)-dependent regulatory mechanism, an H(2)O(2)-resistant mutant (HR40) overproducing CatA was isolated from S . coelicolor A3(2) . Based on the genetic map location of the mutated locus in HR40, the wild type catR gene was isolated from the ordered cosmid library of S . coelicolor by screening for its ability to suppress the HR40 phenotype . catR encodes a protein of 138 amino acids (15319 Da), with sequence homology to ferric uptake regulator (Fur)-like proteins . Disruption of catR caused CatA overproduction as observed in the HR40 mutant, confirming the role of CatR as a negative regulator of catA expression . The levels of catA and catR transcripts were higher in HR40 than in the wild type, implying that CatR represses transcription of these genes . Transcripts from the catA and catR genes were induced within 10 min of H(2)O(2) treatment, suggesting that the repressor activity of CatR may be directly modulated by H(2)O(2) . A putative CatR-binding site containing an inverted repeat of 23 base pairs was localized upstream of the catA and catR gene, on the basis of sequence comparison and deletion analysis . CatR protein purified in the presence of dithiothreitol bound to this region, whereas oxidized CatR, treated with H(2)O(2) or diamide, did not . The redox shift of CatR involved thiol-disulfide exchange as judged by modification of free thiols with 4-acetamido-4'-maleimidylstilbene-2,2'-disulfonate . From these results we propose that CatR regulates its downstream target genes as a repressor whose DNA binding ability is directly modulated by redox changes in the cell. Bull Mem Acad R Med Belg, 1999, 154(6 Pt 2), 371 - 5 {Effect of ammonia on the pig respiratory system . Study of the dose-response relation}; Gustin P; The effects of ammonia on the respiratory system and somatic growth have been measured in piglets taking into account the interactions with endotoxins from gram-bacteria and dust . Dose-response curves were constructed for the inflammatory reactions induced in nasal cavities, the smooth muscle hyperresponsiveness in the trachea, the permeability of alveolo-capillary barrier, the pulmonary vasomotricity, the cough reflex and the systemic effects . An acceptable level of pollution was proposed. Phys Rev Lett, 2000 Jun 5, 84(23), 5435 - 8 Long-lived amide I vibrational modes in myoglobin; Xie A et al.; Pump-probe experiments in the infrared measure vibrational relaxation rates . Myoglobin, which is almost entirely alpha helix in secondary structure, has an unusually long, nonexponential excited state relaxation generated by optically pumping at the blue side ( 5 . 85 microm) of the amide I band . The amino acid alanine and the predominantly beta sheet protein photoactive yellow protein do not have such a long-lived state, suggesting that the alpha helix in proteins can support nonlinear states of 15 ps characteristic times. Carbohydr Res, 2000 Aug 7, 327(4), 447 - 53 An improved model for calculating the optical rotation of simple saccharides; Stroyan EP et al.; A calculational model for the optical rotation (OR) at the sodium D-line of simple saccharides has been developed that eliminates deficiencies of a previous model . Conformational conclusions based on the earlier model are not affected, as established by a recalculation of the OR phi,psi-map of methyl 3-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranoside . The model relocates the strong long-wavelength sigma-sigma* circular dichroism (CD) component, which is mainly responsible for the NaD OR from 160 to below 130 nm, where it is now known to occur . That correction allows future modeling of CD bands of different origins that appear in the 150-190 nm region . In order to demonstrate the utility of the revised model, it was applied to calculating the OR of methyl 3-O-(alpha-L-rhamnopyranosyl)-alpha-L-rhamnopyranoside . The results provide experimental support for conformational conclusions derived from a molecular mechanics study of that molecule. Transfusion, 2000 Sep, 40(9), 1041 - 7 Survival of Ehrlichia chaffeensis in refrigerated, ADSOL-treated RBCs; McKechnie DB et al.; BACKGROUND: The purpose of this study was to investigate the persistence of viable Ehrlichia chaffeensis in ADSOL-treated RBCs stored at 4 to 6 degrees C . STUDY DESIGN AND METHODS: The continuous monocytic cell lines THP-1 and DH82 were infected with E . chaffeensis (St . Vincent isolate) . Packed RBC units were inoculated in separate experiments with E . chaffeensis-infected cells as final concentrations of 8.02 x 10(4) (DH82) and 1.43 x 10(4) (THP-1) infected cells per mL . Aliquots were stored at 4 to 6 degrees C for 1 to 42 days . At selected intervals, nucleated cells from the RBC aliquots were obtained by using a ficoll-isopaque separation procedure . Uninfected DH82 cell cultures were inoculated with the harvested nucleated cells or supernatant . The cell cultures were evaluated for infection by weekly examination of Wright's (Diff-Quik) stained cytocentrifuged slides . PCR amplification was also used to test the harvested nucleated cells or supernatant for the presence of E . chaffeensis DNA . RESULTS: In both types of infected cell lines, E . chaffeensis was reisolated in DH82 cells for as long as 11 days from the cellular fraction and for up to 5 days from the supernatant fraction . PCR results were positive throughout the 42-day testing period . CONCLUSION: Cell-associated E . chaffeensis remains viable in ADSOL-treated RBCs stored at 4 to 6 degrees C for at least 11 days . These data suggest that transfusion-acquired infection is possible . Successful reisolation was achieved from the supernatant fraction, which suggests that RBC products treated with a WBC-reduction procedure may still present a risk for transfusion transmission . No correlation between PCR positivity and viability of bacteria was noted. MMWR Morb Mortal Wkly Rep, 2000 Sep 1, 49(34), 777 - 8 Legionnaires' Disease associated with potting soil--California, Oregon, and Washington, May-June 2000. {How does one treat the osteitis and osteoarthritis of the extremities in older leprosy patients using granulated table sugar?} Grauwin MY, Cartel JL, Lepers JP. Institut de Leprologie Appliquee, Dakar, SenegalA common problem of osteitis and septic arthritis is the recurrent bone infection after surgical debridement, a problem frequently encountered in patients with sequela leprosy . In these cases the authors propose the use of an ancient method of post surgical wound care based on the treatment with ordinary granulated sugar . The hyperosmolar climate created this way in the wounds inhibits the bacterial growth, enhances bacterial death and therefore permits the growth of granulation tissue in order to recover the debrided nude bones . At ILAD (Leprosy Institute of Dakar), 36 osteitis and septic arthritis were treated and healed during the last 2 years from March 1995 to March 1997 using this technic . All the wounds healed in the mean-time of 44 days . Only two of them needed a second debridement and healed afterwards . Up to now the method using ordinary sugar was applied in the treatment of infected wounds, eschars and postsurgical infections . Our experience shows that it also can be indicated to treat bone infections . This method is easy to apply also under often difficult field conditions and is very cheap. Pediatr Int, 2000 Aug, 42(4), 428 - 39 Rotavirus infection in children in Japan; Zhou Y et al.; Currently, a high morbidity of rotavirus diarrhea has been seen in children in developed and developing countries . Improvement of the vaccines is necessary in order to reduce the burden of diarrhea caused by rotavirus . A survey of rotavirus infection from diarrheal stool specimens in children of seven regions in Japan was conducted from 1984 to 1999 . The present study discusses the survey results and reviews the national and international data of more than 23 papers and congress proceedings about rotavirus infection in Japan . We analyze the prevalence of rotavirus infection in acute diarrheal in- and outpatients, the distribution of rotavirus G-serotypes and surveillance data for seasonality and age groups in Japan . The data indicated that rotavirus is the most important cause of diarrhea in Japan among young children, with the prevalence ranging from approximately 9.7 to 88% . The most common rotavirus strains belonged to serotype G1, specifically since 1993 . Serotypes G2, G3 and G4 had also been documented to be predominantly based in the area and year before 1992 . However, untypeable rotavirus strains had been found each year, with a prevalence up to 56.7% which suggests that rare serotypes (except G1-4) or new serotypes might exist . Unexpectedly, in Tokyo and Sapporo from 1998 to 1999, G9 was found to be the first most prevailing serotype with a high prevalence of 52.9 and 71.4%, respectively . Despite these data from different geographic areas, the year under investigation was relatively clear in respect to seasonality, with a peak of rotavirus activity in late winter (February) through early spring (March) . Age distribution had also characterized that the infection was predominant among children aged 1-2 years of age, although it was also common in children of 2-3 years . In addition, mixed infection with bacteria was documented. Mutat Res, 2000 Oct 10, 470(1), 85 - 92 Determination of the vaporization of solutions of mutagenic antineoplastic agents at 23 and 37 degrees C using a desiccator technique; Connor TH et al.; This study evaluated the ability of mutagenic antineoplastic agents to vaporize at room temperature (23 degrees C) and 37 degrees C . A bacterial mutagenicity assay was used to determine the mutagenicity of these agents in the vapor phase . Open plates of bacteria were exposed to varying amounts of drug solutions in sealed glass containers for 24h . The drug solutions were prepared as they would be for patient treatment and were tested at 0.25, 0.5 and 1.0 ml of each drug solution per 10 l of air . Following exposure, the plates exposed at 23 degrees C were incubated an additional 48 h at 37 degrees C to allow for expression of mutations . Those exposed at 37 degrees C were incubated for an additional 24h at 37 degrees C . Carmustine, cyclophosphamide, ifosfamide, thiotepa, and mustargen demonstrated vaporization at 37 degrees C . Carmustine and mustargen also demonstrated significant vaporization at 23 degrees C, while cyclophosphamide demonstrated a 50% increase in revertants at this temperature . In addition, sodium azide, a known mutagen used as a control was also mutagenic as a vapor at both temperatures . Doxorubicin, cisplatin, etoposide, 5-fluorouracil and mitomycin were not detected as vaporizing in this assay . The study found that vaporization of standard solutions of some antineoplastic agents is possible at room temperature and increases as the temperature increases . Therefore, vaporization of spilled antineoplastic agents may present an additional route of exposure to healthcare workers through inhalation. J Bacteriol, 2000 Oct, 182(19), 5615 - 9 The Synechococcus strain PCC 7942 glnN product (glutamine synthetase III) helps recovery from prolonged nitrogen chlorosis; Sauer J et al.; We report the cloning and sequencing of the glnN gene encoding a class III glutamine synthetase from the cyanobacterium Synechococcus strain PCC 7942 . Mapping of the transcriptional start site revealed a DNA sequence in the promoter region that resembles an imperfect NtcA binding motif . Expression of glnN is impaired in NtcA- and P(II)-deficient mutants . The only parameter which was negatively affected in the glnN mutant compared to the wild type was the recovery rate of prolonged nitrogen-starved cells with low concentrations of combined nitrogen. J Bacteriol, 2000 Oct, 182(19), 5433 - 9 DNA polymerase I is essential for growth of Methylobacterium dichloromethanicum DM4 with dichloromethane; Kayser MF et al.; Methylobacterium dichloromethanicum DM4 grows with dichloromethane as the unique carbon and energy source by virtue of a single enzyme, dichloromethane dehalogenase-glutathione S-transferase . A mutant of the dichloromethane-degrading strain M . dichloromethanicum DM4, strain DM4-1445, was obtained by mini-Tn5 transposon mutagenesis that was no longer able to grow with dichloromethane . Dichloromethane dehalogenase activity in this mutant was comparable to that of the wild-type strain . The site of mini-Tn5 insertion in this mutant was located in the polA gene encoding DNA polymerase I, an enzyme with a well-known role in DNA repair . DNA polymerase activity was not detected in cell extracts of the polA mutant . Conjugation of a plasmid containing the intact DNA polymerase I gene into the polA mutant restored growth with dichloromethane, indicating that the polA gene defect was responsible for the observed lack of growth of this mutant with dichloromethane . Viability of the DM4-1445 mutant was strongly reduced upon exposure to both UV light and dichloromethane . The polA'-lacZ transcriptional fusion resulting from mini-Tn5 insertion was constitutively expressed at high levels and induced about twofold after addition of 10 mM dichloromethane . Taken together, these data indicate that DNA polymerase I is essential for growth of M . dichloromethanicum DM4 with dichloromethane and further suggest an important role of the DNA repair machinery in the degradation of halogenated, DNA-alkylating compounds by bacteria. Med Hypotheses, 2000 Sep, 55(3), 185 - 6 Monoselenolipoic acid may be an outstanding pharmaceutical antioxidant with direct thioredoxin-like activity; McCarty MF; Owing to the low pKa of its selenohydryl group, reduced monoselenolipoic acid (MSL) can be expected to be a very versatile antioxidant with direct thioredoxin-like activity . Since MSL supports the growth of lipoate-dependent bacteria, it can be anticipated that MSL will be susceptible to reversible reduction by one or more of the enzymes that reduces lipoic acid - thus greatly potentiating MSL's utility as an antioxidant . If it is not metabolized to release toxic free selenide, MSL may have interesting pharmaceutical potential . Arch Microbiol, 2000 Jul-Aug, 174(1-2), 35 - 41 Transformation of ortho-substituted biphenyls by Methylosinus trichosporium OB3b: substituent effects on oxidation kinetics and product formation; Lindner AS et al.; The ability of Methylosinus trichosporium OB3b, expressing soluble methane monooxygenase, to oxidize a range of ortho-substituted biphenyls was examined to better understand how substituents affect both the rate and products of oxidation in comparison to biphenyl . Inhibition of oxidation was observed over the tested substrate range for both biphenyl and ortho-halogenated biphenyls (2-chloro-, 2-bromo-, and 2-iodobiphenyl) . No inhibition was observed during the oxidation of 2-hydroxybiphenyl and 2-methylbiphenyl . Analysis of the products of oxidation showed that, depending on the substituent, ring hydroxylation, substituent oxidation, and elimination pathways could occur . The type and abundance of products formed along with the relatively high kinetic isotope effect observed for deuterated vs . nondeuterated biphenyl (k(h)/k(d) = 3.4+/-0.02) are consistent with mechanisms that include both hydrogen abstraction and NIH-shift pathways . Knowledge of these substituent-dependent reaction rates and mechanisms enhances our understanding of the methanotrophic aryl transformation potential and allows for better prediction of the formation of oxidized intermediates by methanotrophic bacteria. Int J Tuberc Lung Dis, 2000 Sep, 4(9), 807 - 26 Case management of childhood pneumonia in developing countries: recent relevant research and current initiatives; Rasmussen Z et al.; Acute respiratory infections (ARI), mostly pneumonia, are one of the leading causes of death in young children in developing countries, accounting for 28% of childhood mortality . This paper provides a summary of the research and technical development efforts made in the last 15 years which contributed to improving the effectiveness of the case management strategy to reduce mortality from pneumonia in children in developing countries . Community intervention studies provided strong evidence that the strategy was feasible and effective in producing a substantial impact on pneumonia mortality . Clinical studies provided the rationale for improving the sensitivity and specificity of key signs of pneumonia, and for enhancing the therapeutic efficacy of standard home treatment . Research also provided data to deal with the problem of the clinical overlap of pneumonia and malaria in children . Technological initiatives succeeded in making appropriate diagnostic and therapeutic devices available . An important body of socio-cultural knowledge about family practices regarding pneumonia and ARI in children was built up and provided orientation on effective communication between health workers and families about home care of children with ARI . Health systems research focused on methods for surveillance of bacterial drug resistance and methodologies for evaluating the control programmes . Despite advances in the development of vaccines against respiratory bacteria and in the prevention of risk factors for pneumonia in children, case management will continue to be a central strategy for preventing mortality . Current international research initiatives are looking into measures that can improve the referral of severe pneumonia and effective management of severe pneumonia at first level hospitals. Bioessays, 2000 Oct, 22(10), 947 - 53 Redox control and the evolution of multicellularity; Blackstone NW; Redox chemistry, involving the transfer of electrons and hydrogen atoms, is central to energy conversion in respiration; in addition, control of gene expression by redox state commonly occurs in bacteria, allowing a rapid response to environmental changes, such as altered 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 of development and spacing of structures in response to a variable food supply and other environmental factors . Experimental perturbations of redox state in colonial hydroids support this notion of adaptive redox control, and redox signaling in metazoans may have evolved in this ecological context . At the same time, redox signaling has important consequences for the evolutionary transition from unicellular to multicellular organisms . Unlike protein or peptide signaling, redox signaling acting in concert with programmed cell death may automatically inflict a cost on those cells that "defect," that is, selfishly favor their own replication rate over that of the multicellular group . In this way, redox signaling may have allowed multicellular individuality to evolve and more easily be maintained. Proc Natl Acad Sci U S A, 2000 Sep 26, 97(20), 10855 - 60 Postsymbiotic plasmid acquisition and evolution of the repA1-replicon in Buchnera aphidicola; Van Ham RC et al.; Buchnera aphidicola is an obligate, strictly vertically transmitted, bacterial symbiont of aphids . It supplies its host with essential amino acids, nutrients required by aphids but deficient in their diet of plant phloem sap . Several lineages of Buchnera show adaptation to their nutritional role in the form of plasmid-mediated amplification of key-genes involved in the biosynthesis of tryptophan (trpEG) and leucine (leuABCD) . Phylogenetic analyses of these plasmid-encoded functions have thus far suggested the absence of horizontal plasmid exchange among lineages of Buchnera . Here, we describe three new Buchnera plasmids, obtained from species of the aphid host families Lachnidae and Pemphigidae . All three plasmids belong to the repA1 family of Buchnera plasmids, which is characterized by the presence of a repA1-replicon responsible for replication initiation . A comprehensive analysis of this family of plasmids unexpectedly revealed significantly incongruent phylogenies for different plasmid and chromosomally encoded loci . We infer from these incongruencies a case of horizontal plasmid transfer in Buchnera . This process may have been mediated by secondary endosymbionts, which occasionally undergo horizontal transmission in aphids. J Allergy Clin Immunol, 2000 Sep, 106(3), 409 - 18 Recombinant allergens for diagnosis and therapy of allergic disease; Chapman MD et al.; Many of the problems associated with using natural allergenic products for allergy diagnosis and treatment can be overcome with use of genetically engineered recombinant allergens . Over the past 10 years, the most important allergens from mites, pollens, animal dander, insects, and foods have been cloned, sequenced, and expressed . In many cases the three-dimensional allergen structure has been determined and B-cell and T-cell epitopes have been mapped . These studies show that allergens have diverse biologic functions (they may be enzymes, enzyme inhibitors, lipocalins, or structural proteins) and that as a rule the allergen function is unrelated to its ability to cause IgE antibody responses . High-level expression systems have been developed to produce recombinant allergens in bacteria, yeast, or insect cells . Recombinant allergens show comparable IgE antibody binding to their natural counterparts (where available) and show excellent reactivity on skin testing and in in vitro diagnostic tests . Cocktails of recombinant allergens can be formulated with predetermined and uniform allergen levels, which could replace natural allergens and result in the development of innovative, patient-based tests for allergy diagnosis . Recombinant allergens also offer the exciting possibility of developing new forms of allergen immunotherapy, including the use of hypoallergens, allergens coupled to IgE suppressive adjuvants, and peptide-based therapies . The production of recombinant allergens as defined molecular entities makes it feasible to consider the possibility of developing prophylactic allergen vaccines . The introduction of recombinant allergens in research and in clinical trials should lead to significant improvements in allergy diagnosis and treatment. Nature, 2000 Aug 31, 406(6799), 998 - 1001 A role for the C3a anaphylatoxin receptor in the effector phase of asthma; Humbles AA et al.; Asthma is a chronic inflammatory disease of the airways and lung mucosa with a strong correlation to atopy and acquired (IgE) immunity . However, many features of bronchial asthma, such as smooth muscle contraction, mucus secretion and recruitment of inflammatory cells, are consistent with the actions of complement anaphylatoxins, in particular C3a and C5a . Complement activation forms a central core of innate immune defence against mucosal bacteria, viruses, fungi, helminths and other pathogens . As a system of 'pattern-recognition molecules', foreign surface antigens and immune complexes lead to a proteolytic cascade culminating in a lytic membrane attack . The anaphylatoxins C3a and C5a are liberated as activation byproducts and are potent pro-inflammatory mediators that bind to specific cell surface receptors and cause leukocyte activation, smooth muscle contraction and vascular permeability . Here we show that in a murine model of allergic airway disease, genetic deletion of the C3a receptor protects against the changes in lung physiology seen after allergen challenge . Furthermore, human asthmatics develop significant levels of ligand C3a following intra-pulmonary deposition of allergen, but not saline . We propose that, in addition to acquired immune responses, the innate immune system and complement (C3a in particular) are involved in the pathogenesis of asthma. Proc R Soc Lond B Biol Sci, 2000 Jul 22, 267(1451), 1469 - 73 Male-killing Wolbachia in a flour beetle; Fialho RF et al.; The bacteria in the genus Wolbachia are cytoplasmically inherited symbionts of arthropods . Infection often causes profound changes in host reproduction, enhancing bacterial transmission and spread in a population . The reproductive alterations known to result from Wolbachia infection include cytoplasmic incompatibility (CI), parthenogenesis, feminization of genetic males, fecundity enhancement, male killing and, perhaps, lethality Here, we report male killing in a third insect, the black flour beetle Tribolium madens, based on highly female-biased sex ratios of progeny from females infected with Wolbachia . The bias is cytoplasmic in nature as shown by repeated backcrossing of infected females with males of a naturally uninfected strain . Infection also lowers the egg hatch rates significantly to approximately half of those observed for uninfected females . Treatment of the host with antibiotics eliminated infection, reverted the sex ratio to unbiased levels and increased the percentage hatch . Typically Wolbachia infection is transmitted from mother to progeny, regardless of the sex of the progeny; however, infected T . madens males are never found . Virgin females are sterile, suggesting that the sex-ratio distortion in T . madens results from embryonic male killing rather than parthenogenesis . Based on DNA sequence data, the male-killing strain of Wolbachia in T . madens was indistinguishable from the CI-inducing Wolbachia in Tribolium confusum, a closely related beetle . Our findings suggest that host symbiont interaction effects may play an important role in the induction of Wolbachia reproductive phenotypes. Nurs Times, 1999 Nov 3-9, 95(44), 62, 65 - 6 Infection control . Unhappy nappy-changing; Chudleigh J et al.; This study was conducted in a special care baby unit (SCBU) and concentrated on hand decontamination and glove-wearing during nappy-changing . The aims were to determine whether or not nurses were adhering to existing infection control policies and guidelines as well as determining the most appropriate product to use for hand decontamination . The study demonstrated that not all nurses were adhering to existing infection control policies and guidelines . On the majority of occasions, alcohol was significantly better than soap at removing bacteria from the hands, but gloves provided the best protection overall for both babies and nurses by preventing the acquisition of bacteria. J Zoo Wildl Med, 2000 Jun, 31(2), 255 - 8 Zinc-responsive dermatosis in a red wolf (canis rufus); Kearns K et al.; An 18-mo-old male red wolf (Canis rufus) presented with footpad hyperkeratosis, suppurative paronychia, distal limb pyoderma, and peripheral lymphadenopathy . Diet for the previous 11 mo consisted of a mixture of two commercially prepared dog foods with a mineral supplement containing primarily calcium . Culture of the draining tracts on the distal limbs yielded a mixed population of opportunistic bacteria . Histopathologic findings were consistent with a diagnosis of zinc deficiency . Medical therapy consisted of 15 mg/kg amoxicillin p.o . b.i.d . and 10 mg/kg zinc sulfate p.o . s.i.d . Calcium supplementation was discontinued . Clinical signs resolved by 10 wk after the initiation of treatment. FEMS Microbiol Lett, 2000 Sep 1, 190(1), 103 - 8 Evaluation of strain-specific primer sequences from an abortifacient strain of ovine Chlamydophila abortus (Chalmydia psittaci) for the detection of EAE by PCR; Creelan JL et al.; Strain-specific primer sequences derived from the helicase gene of an ovine abortifacient strain (S26/3) of Chlamydophila abortus (Chlamydia psittaci) were evaluated for the diagnosis of enzootic abortion in ewes (EAE) by polymerase chain reaction (PCR) . C abortus DNA was amplified from tissues submitted from ovine abortion cases using genus-specific and strain-specific primers in a standard thermal cycler . Amplification was followed by Southern blotting and hybridisation with a strain-specific probe . Real-time PCR was also evaluated using strain-specific primers in a microvolume fluorimeter-based thermal cycler (LightCycler) . Detection using both PCR methods was compared with other diagnostic methods against the standard of McCoy cell culture isolation . In this paper we report the application of strain-specific PCR as a fast, sensitive, specific method for the detection of EAE. Curr Opin Struct Biol, 2000 Aug, 10(4), 443 - 7 beta-Barrel membrane proteins; Schulz GE; beta-Barrel proteins are found in the outer membranes of bacteria, mitochondria and chloroplasts . The presently known sizes range from small eight-stranded to large twenty-two-stranded beta barrels existing as monomers and oligomers . Their functions are as diverse as active ion transport, passive nutrient intake, membrane anchors, membrane-bound enzymes and defense against attack proteins . Of further interest are the folding process, the crystallization, the observed limited structural diversity and the manifold channel engineering options of these beta-barrel proteins. Int J Cardiol, 2000 Aug 31, 75 Suppl 1, S37 - 45; discussion S47-52 Animal models of infection-mediated vasculitis: implications for human disease; Dal Canto AJ et al.; The human vasculitides including Takayasu's arteritis are idiopathic syndromes for which both autoimmune and infectious etiologies have been proposed . Although proof of a correlation between infection and human vasculitis would aid in patient management, it is difficult to confirm causality . To study infection-mediated vascular disease, different animal models have been developed . Infections with the bacteria C . pneumoniae, an RNA virus, and herpesviruses all cause vascular pathology and will be reviewed here . Many aspects of the human diseases are recapitulated in these models, so further animal studies may help elucidate mechanisms of infection-mediated vasculitis . Such results may improve management, and potentially, prevention of these important human diseases . Importantly, the animal models provide an opportunity to define how the immune and inflammatory processes function in the great vessels and the molecular basis for the selectivity of certain viral infections for the great elastic arteries. Int J Parasitol, 2000 Aug, 30(9), 995 - 1000 Anhydrobiotic potential and long-term storage of entomopathogenic nematodes (Rhabditida: Steinernematidae); Grewal PS; Anhydrobiosis is considered to be an important means of achieving storage stability of entomopathogenic nematodes that are used in biological control . This study explored the effects of anhydrobiosis on longevity and infectivity of infective juveniles (IJs) of three species of entomopathogenic nematodes Steinernema carpocapsae, Steinernema feltiae, and Steinernema riobrave at 5 and 25 degrees C . Anhydrobiosis was induced in water-dispersible granules (WG) at 0.966-0.971 water activity and 25 degrees C following a 7-day preconditioning of IJs at 5 degrees C in tap water . Survival and infectivity of the desiccated (anhydrobiotic) IJs was compared with non-desiccated IJs stored in water for different periods . Anhydrobiosis increased longevity of S . carpocapsae IJs by 3 months and of S . riobrave by 1 month in WG at 25 degrees C as compared with IJs stored in water . However, desiccation decreased S . feltiae longevity at 25 degrees C and of all three species at 5 degrees C . These results demonstrate a shelf-life of 5 months for S . carpocapsae at 25 degrees C and 9 months at 5 degrees C in WG with over 90% IJ survival . For S . feltiae, over 90% survival occurred only for 2 months at 25 degrees C and 5 months at 5 degrees C in WG . Steinernema riobrave had over 90% survival only for 1 month at 25 degrees C and the survival dropped below 85% within 1 month at 5 degrees C . Induction of anhydrobiosis in WG resulted in 85, 79 and 76% reduction in oxygen consumption by S . carpocapsae, S . feltiae, and S . riobrave IJs, respectively . Differences in IJ longevity among three species in water at 25 degrees C were related both to the initial lipid content and the rate of lipid utilisation, but not at 5 degrees C . The one-on-one infection bioassays indicated that desiccation had no negative effect on the infectivity of any of the nematode species suggesting no harmful effect on the IJs and/or their symbiotic bacteria . The species differences in IJ longevity and desiccation survival at different temperatures are discussed in relation to their foraging strategy and temperature adaptation. J Am Soc Echocardiogr, 2000 Sep, 13(9), 876 - 81 Echocardiography in nonbacterial thrombotic endocarditis: from autopsy to clinical entity; Reisner SA et al.; Bacteria-free verrucae, frequently termed "non-bacterial thrombotic endocarditis," have been recognized in autoimmune disorders as well as in neo-plastic diseases . The antemortem diagnosis of non-bacterial thrombotic endocarditis is rare, and most existing data result from postmortem examinations . In 3 prospective echocardiographic studies we found typical cardiac valvular lesions in patients with primary antiphospholipid syndrome, myelo-proliferative disorders, and solid malignant tumors . Cardiac lesions associated with these 3 different entities had common echocardiographic appearance and correlated positively with thromboembolic events . The possibility of common pathogenesis is suggested, and clinical significance is discussed. J Infect Dis, 2000 Oct, 182(4), 1264 - 7 Epub 2000 Sep 05. Reduced ex vivo chemokine production by polymorphonuclear cells after in vivo exposure of normal humans to endotoxin; Schultz MJ et al.; Monocytes from patients with sepsis have a reduced capacity to produce cytokines, a state referred to as immunoparalysis . To determine whether polymorphonuclear leukocytes (PMNL) can be rendered hyporesponsive, PMNL from 6 healthy volunteers intravenously challenged with lipopolysaccharide (LPS; 4 ng/kg) were stimulated ex vivo with heat-killed bacteria or LPS, and the release of the CXC chemokines interleukin-8, epithelial-derived neutrophil attractant-78, and growth-related oncogen-alpha was measured . At 1 and 2 h after LPS administration in vivo, PMNL produced fewer CXC chemokines after stimulation with bacteria or LPS (all P<.05) . Serum obtained 2 h after in vivo administration of LPS did not influence chemokine production by PMNL from 6 healthy volunteers not previously exposed to LPS . Thus, intravenous injection of LPS induces a refractory state of PMNL that is not caused by soluble factors produced in response to in vivo exposure to LPS. Biocell, 2000 Aug, 24(2), 123 - 32 The laticifer system of Chamaesyce thymifolia: a closed host environment for plant trypanosomatids; da Cunha M et al.; Specimens of Chamaesyce thymifolia (Euphorbiaceae) infected and uninfected by Phytomonas sp., a parasite of the Trypanosomatidae family, were anatomically and ultrastructurally analyzed with special emphasis on the laticifer system . C . thymifolia presents branched non-articulated laticifers and was heavily infected by Phytomonas sp . in all collection sites . Infection was often observed in the initial stages inside the vacuole, when the latex particles could be seen . In intermediary stages of laticifer differentiation, Phytomonas sp . were found free in the cytoplasm, inside small vacuoles or in the central vacuole . In differentiated laticifers that had only the plasma membrane, Phytomonas sp . were free in the latex and close to the cell membrane . Infected and uninfected plants showed identical anatomy and ultrastructure and the starch grain numbers in the latex were not reduced in the presence of this flagellate . Biochemical analysis of the latex of infected and uninfected plants presented similar levels of protein, carbohydrate and beta-1,3-glucanase, suggesting that this species is not pathogenic for the host . Besides, all infected plants complete its life cycle . Plants infected with Phytomonas presented occasionally virus like particles and bacteria inside the laticifer tubes. Isr Med Assoc J, 2000 Aug, 2(8), 580 - 2 Association of Ureaplasma urealyticum colonization in male urethra and Condyloma acuminatum; Zvulunov A et al.; BACKGROUND: The frequent coexistence of two or more sexually transmitted diseases in one patient has been reported in non-dermatological literature, mostly in languages other than English . Identification of Ureaplasma urealyticum, Chlamydia trachomatis and Mycoplasma hominis in men with other STDs is important, since these bacteria have been implicated in a variety of diseases such as non-gonococcal urethritis, premature rupture of fetal membranes, and infertility in female sexual partners of these patients . OBJECTIVE: To assess the frequency of concomitant STD, particularly urethral colonization of U . urealyticum, C . trachomatis and M . hominis, in men consulting for suspected STD-related symptoms . METHODS: All patients attending our dermatology clinic for STD-related symptoms during a 12 month period in 1996-97 underwent systematic clinical and laboratory screening for syphilis, gonorrhea, NGU, prostatitis, genital herpes simplex infection, Condyloma acuminatum, urethral carriage of U . urealyticum, C . trachomatis and M . hominis, as well as serological screening for HIV, and hepatitis B and C infections . RESULTS: A total of 169 men with STD-related symptoms were enrolled in the study . The following clinical diagnoses were established: NGU in 109 men, C . acuminatum in 40, genital herpes simplex in 10, prostatitis in 7, latent syphilis in 6, primary syphilis in 1, and Behcet's disease in 1 . No clinical evidence of STD was found in 13 patients . Of the 169 patients, 39 (23%) had two or more concomitant STDs, of whom 27 (69%) had C . acuminatum associated with one or more of the urethral pathogens . A positive U . urealyticum culture was found in 67.5% (27/40) of the men with C . acuminatum as compared to 42% (40/96) among the patients with NGU who did not have C . acuminatum (P = 0.004, chi 2 test) . Conversely, the prevalence of C . acuminatum among patients positive for U . urealyticum was significantly higher than the prevalence among those who were negative--27/75 (36%) vs . 13/94 (14%), P < 0.0009, chi 2 test . About half of the U . urealyticum-positive patients with C . acuminatum had no clinical signs or symptoms of urethritis . CONCLUSION: Our findings suggest that patients with C . acuminatum should be assessed for U . urealyticum carriage and, when identified, their sexual contacts should be actively sought and treated. Nihon Kokyuki Gakkai Zasshi, 2000 Jun, 38(6), 490 - 3 {Pulmonary dirofilariasis with cavity formation and pleural effusion}; Sano A et al.; A 61-year-old man visited a community hospital because of hemosputum . A solitary nodule in the left lower lung field was pointed out on a chest roentgenogram . The patient was treated with antibiotics, but the solitary nodule increased in size . He was referred to our hospital because of high fever and observations of cavity formation and pleural effusion on a chest roentgenogram . The pleural effusion showed no cytologic evidence of malignancy, and cultures were also negative for bacteria . An increased percentage of lymphocytes was detected in the pleural effusion, but slight eosinophilia was found in blood samples . Dot enzyme-linked immunosorbent assay and Ouchterlony's double-diffusion test yielded a diagnosis of pulmonary dirofilariasis . After drainage of the pleural effusion by thoracentesis, spontaneous regression was observed . Cavity formation, pleural effusion, and spontaneous regression are in general rare in patients with pulmonary dirofilariasis. Biochemistry, 2000 Sep 5, 39(35), 10684 - 94 Molecular architecture of the mutagenic active site of human immunodeficiency virus type 1 reverse transcriptase: roles of the beta 8-alpha E loop in fidelity, processivity, and substrate interactions; Weiss KK et al.; Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) is a putative source of the genomic hypermutation that promotes rapid evolution of HIV-1 . To understand the molecular strategies that create a highly mutagenic DNA polymerase active site in HIV-1 RT, we investigated the roles of four residues in the beta 8-alpha E loop . Gln151, which interacts with the sugar of the incoming dNTP, and Lys154, which interacts with the template, yielded site-directed mutants with increased fidelity, suggesting that these residues are directly involved in the mutagenic architecture of the active site . Mutations at Gln151 and Lys154 also reduced processivity . Q151N RT showed enhanced ability to discriminate between TTP and AZT triphosphate, consistent with the observation that the Q151M mutation confers AZT resistance in vivo . Mutations at Gly152 greatly decreased RT activity; molecular modeling suggests that Gly152 is critical for the proper geometric alignment that permits base-pairing of the incoming dNTP with the template . Mutations at Trp153 reduced the expression level, and presumably the stability, of RT proteins in bacteria . These observations support the conclusion that interactions of active site residues in the beta 8-alpha E loop with incoming dNTPs and the template are determinants of the accuracy, processivity, and substrate selectivity of HIV-1 RT. Cell Mol Biol (Noisy-le-grand), 2000 Jul, 46(5), 895 - 913 New techniques in fast time-resolved structure determination; Perman B et al.; New techniques in fast time-resolved X-ray crystallography provide a different approach to understanding the structural basis of protein function . Two biological systems have been studied as part of the refinement of these techniques, and have actually spurred new ideas in time-resolved structural studies . The dissociation of carbon monoxide from carbon-monoxy myoglobin has earlier been investigated over a time range spanning 18 orders of magnitude (femtoseconds to hours) using spectroscopic methods . Rapid time-resolved determination of the entire myoglobin structure made it possible to determine both the position of the CO after photodissociation and the entire globin structure, over a time range from nanoseconds to milliseconds, during which the heme and globin relax and the carbon monoxide rebinds . Photoactive yellow protein, a relative newcomer to biophysical research, has a fully-reversible photocycle containing several spectrally distinct intermediates . Identifying and solving the structures of each intermediate is the initial goal in time-resolved studies on this protein and will contribute to a greater understanding of the biological process of light driven signal transduction. Am J Vet Res, 2000 Sep, 61(9), 1145 - 9 Detection of cell detachment activity induced by Moraxella bovis; Marrion RM et al.; OBJECTIVE: To characterize the effect that filtrate obtained from cultures of Moraxella bovis has on cultured corneal epithelial cells and other types of cultured mammalian cells . SAMPLE POPULATION: Cultured hamster corneal epithelial cells, bovine epithelial cells, and several transformed cell lines exposed to culture filtrate from a pathogenic isolate of M bovis . PROCEDURE: Moraxella bovis was cultured, and bacteria were removed by filtration . The resulting bacterial culture filtrate was incubated with various types of cultured cells, and effects of the filtrate on detachment of various mammalian cell types was quantified by the use of neutral red dye . Additionally, bacterial culture filtrate was treated with protease inhibitors as well as trypsin and heat prior to incubation with cultured mammalian cells . RESULTS: Bacterial culture filtrate of M bovis caused all types of cultured cells to detach from each other and from the substrate, with the maximal effect evident 2 hours after initiating incubation . Detached cells were alive, and detachment was reversible . Serine protease inhibitors (phenylmethylsulfonylfluoride and alpha2-macroglobulin) inhibited cell detachment attributable to bacterial culture filtrate . Heating and treatment with trypsin also inhibited cell detachment . CONCLUSIONS AND CLINICAL RELEVANCE: Moraxella bovis produces a soluble factor that causes reversible detachment of cultured cells . This activity may play a role in the pathogenesis of infectious bovine keratoconjunctivitis. J Biol Chem, 2000 Dec 8, 275(49), 38302 - 10 Redox switch of hsp33 has a novel zinc-binding motif; Jakob U et al.; The chaperone activity of the heat shock protein Hsp33 is regulated by reversible disulfide bond formation . Oxidized Hsp33 is active, and reduced Hsp33 is inactive . We show that zinc binding is essential for the function of this redox switch . Our results reveal that Hps33 contains a new, high affinity (K(a) > 10(17) m(-)(1)), zinc-binding motif in the form Cys-X-Cys-X(27-32)-Cys-X-X-Cys . All four conserved cysteines within this motif act to coordinate a single zinc atom . Experiments where reduced wild type Hsp33 is reconstituted with cobalt or cadmium demonstrate that the metal-coordinating cysteines are present as highly reactive thiolate anions . This ionization may allow for the fast and successful activation of the chaperone function of Hsp33 upon incubation in oxidizing agents. Protein Sci, 2000 Aug, 9(8), 1503 - 18 Chemical cross-linking with thiol-cleavable reagents combined with differential mass spectrometric peptide mapping--a novel approach to assess intermolecular protein contacts; Bennett KL et al.; The intermolecular contact regions between monomers of the homodimeric DNA binding protein ParR and the interaction between the glycoproteins CD28 and CD80 were investigated using a strategy that combined chemical cross-linking with differential MALDI-MS analyses . ParR dimers were modified in vitro with the thiol-cleavable cross-linker 3,3'-dithio-bis(succinimidylproprionate) (DTSSP), proteolytically digested with trypsin and analyzed by MALDI-MS peptide mapping . Comparison of the peptide maps obtained from digested cross-linked ParR dimers in the presence and absence of a thiol reagent strongly supported a "head-to-tail" arrangement of the monomers in the dimeric complex . Glycoprotein fusion constructs CD28-IgG and CD80-Fab were cross-linked in vitro by DTSSP, characterized by nonreducing SDS-PAGE, digested in situ with trypsin and analyzed by MALDI-MS peptide mapping (+/- thiol reagent) . The data revealed the presence of an intermolecular cross-link between the receptor regions of the glycoprotein constructs, as well as a number of unexpected but nonetheless specific interactions between the fusion domains of CD28-IgG a