Proc Natl Acad Sci U S A, 2000 Apr 25, 97(9), 4873 - 8 A self-recombining bacterial artificial chromosome and its application for analysis of herpesvirus pathogenesis; Smith GA et al.; A self-recombining bacterial artificial chromosome (BAC) containing the 142-kb pseudorabies virus genome was constructed such that the viral genome is rapidly excised from the BAC vector backbone on delivery into mammalian cells . The recombination is mediated by loxP sites in the plasmid and Cre recombinase encoded within the BAC vector . A synthetic intron inserted in the middle of the cre ORF completely inhibits recombination in Escherichia coli, but is spliced out after delivery of the plasmid into mammalian cells . Recombination is efficient, and pure virus lacking the BAC vector backbone is immediately isolated from transfected mammalian cells without the need of serial passage or plaque purification. Proc Natl Acad Sci U S A, 2000 Apr 25, 97(9), 4730 - 5 A mutant hunt for defects in membrane protein assembly yields mutations affecting the bacterial signal recognition particle and Sec machinery; Tian H et al.; We describe an Escherichia coli genetic screen that yields mutations affecting two different cellular processes: disulfide bond formation and membrane protein assembly . The mutants defective in disulfide bond formation include additional classes of dsbA and dsbB mutations . The membrane protein assembly defective mutants contain a mutation in the secA operon and three mutations in the ffs gene, which encodes 4.5S RNA . These latter mutations are the only ones to be isolated in a gene encoding a component of the bacterial signal recognition particle by screening in vivo for defects in membrane protein insertion . A sensitive method for examining membrane protein localization shows that the ffs and secA locus mutations affect membrane assembly of the polytopic membrane protein, MalF . The ffs mutations also affect the membrane insertion of the FtsQ and the AcrB proteins . Although both the ffs and the secA locus mutations interfere with membrane protein assembly, only the latter also reduces export of a protein containing a cleavable signal sequence. Proc Natl Acad Sci U S A, 2000 Apr 25, 97(9), 4649 - 53 Robust perfect adaptation in bacterial chemotaxis through integral feedback control; Yi TM et al.; Integral feedback control is a basic engineering strategy for ensuring that the output of a system robustly tracks its desired value independent of noise or variations in system parameters . In biological systems, it is common for the response to an extracellular stimulus to return to its prestimulus value even in the continued presence of the signal-a process termed adaptation or desensitization . Barkai, Alon, Surette, and Leibler have provided both theoretical and experimental evidence that the precision of adaptation in bacterial chemotaxis is robust to dramatic changes in the levels and kinetic rate constants of the constituent proteins in this signaling network {Alon, U., Surette, M . G., Barkai, N . & Leibler, S . (1998) Nature (London) 397, 168-171} . Here we propose that the robustness of perfect adaptation is the result of this system possessing the property of integral feedback control . Using techniques from control and dynamical systems theory, we demonstrate that integral control is structurally inherent in the Barkai-Leibler model and identify and characterize the key assumptions of the model . Most importantly, we argue that integral control in some form is necessary for a robust implementation of perfect adaptation . More generally, integral control may underlie the robustness of many homeostatic mechanisms. Lab Anim, 1999 Apr, 33(2), 143 - 8 Bacterial counts in canine duodenal fluid after exposure to saline, sodium bicarbonate and hypertonic dextrose solutions used to maintain patency of chronically implanted catheters; Lynch TM et al.; Flushing of intestinal vascular access ports (VAPs) is commonly performed to prevent the problems of blockage and infection, and in this study four different flushing solutions were compared . The growth of bacteria from canine duodenal contents was compared in: 0.9% saline, 50% dextrose, 8.4% sodium bicarbonate (NaHCO3) and 0.01 M phosphate buffered saline (PBS) . Duodenal contents from three laboratory beagles were serially diluted in these four solutions, spread plated onto agar at 24 h periods for 7 days and bacterial counts were performed . Immediately after the duodenal juices were added, no significant differences could be seen in bacterial counts with any of the solutions . Over the 7 day period, bacterial numbers greatly increased in saline and phosphate buffered saline, but greatly decreased in dextrose and sodium bicarbonate solutions . Dextrose and sodium bicarbonate appeared to be the most promising flushing solutions tested to minimize infections of associated intestinal VAPs. FEMS Microbiol Ecol, 2000 Apr 1, 32(1), 69 - 75 Characterization of bacterial strains able to grow on high molecular mass residues from crude oil processing; Yuste L et al.; Oil residues containing high molecular mass hydrocarbons, rich in polyaromatic compounds, are frequent end-products of crude oil processing and are poorly biodegradable . Their disposal poses an environmental problem . Through batch-enrichments from contaminated soils we have isolated and characterized seven bacterial strains that can use a residue from crude oil processing as a source of carbon and energy . The residue was a complex mixture of high molecular mass compounds, including saturated, aromatic and polycyclic aromatic hydrocarbons (PAHs) . Analysis of the metabolic profiles of the strains isolated showed that they could all metabolize long-chain-length alkanes efficiently, but not PAHs . Strains degrading naphthalene, a simple PAH, did exist in the soil inocula used, but could be isolated only when enrichments were performed using pure naphthalene as the sole carbon source . All strains tested emulsified the oil residue and their ability to produce surfactants was studied. Biophys J, 2000 May, 78(5), 2590 - 6 B800-->B850 energy transfer mechanism in bacterial LH2 complexes investigated by B800 pigment exchange; Herek JL et al.; Femtosecond transient absorption measurements were performed on native and a series of reconstituted LH2 complexes from Rhodopseudomonas acidophila 10050 at room temperature . The reconstituted complexes contain chemically modified tetrapyrrole pigments in place of the native bacteriochlorophyll a-B800 molecules . The spectral characteristics of the modified pigments vary significantly, such that within the B800 binding sites the B800 Q(y) absorption maximum can be shifted incrementally from 800 to 670 nm . As the spectral overlap between the B800 and B850 Q(y) bands decreases, the rate of energy transfer (as determined by the time-dependent bleaching of the B850 absorption band) also decreases; the measured time constants range from 0.9 ps (bacteriochlorophyll a in the B800 sites, Q(y) absorption maximum at 800 nm) to 8.3 ps (chlorophyll a in the B800 sites, Q(y) absorption maximum at 670 nm) . This correlation between energy transfer rate and spectral blue-shift of the B800 absorption band is in qualitative agreement with the trend predicted from Forster spectral overlap calculations, although the experimentally determined rates are approximately 5 times faster than those predicted by simulations . This discrepancy is attributed to an underestimation of the electronic coupling between the B800 and B850 molecules. Presse Med, 2000 Mar 25, 29(11), 584 - 8 {Rapid diagnosis of the type of meningitis (bacterial or viral) by the assay of serum procalcitonin}; Viallon A et al.; OBJECTIVE: It has been shown that serum procalcitonin (PCT) can be used to differentiate bacterial from viral meningitis in children in all cases . The aim of this study was to demonstrate the interest of PCT in the management of suspected meningitis in adults . PATIENTS AND METHODS: We conducted a prospective study including 179 consecutive patients admitted to the emergency department for suspected meningitis . All samples were taken at patient admission . The discriminant potential between bacterial and viral meningitis was studied for cerebrospinal fluid parameters (cytology, protein, glucose, lactate) and serum parameters (C reactive protein, PCT) . RESULTS: Thirty-two patients had bacterial meningitis, 90 had viral meningitis and meningitis was ruled out in 57 . Among all studied parameters, the most discriminant for distinguishing between bacterial and viral meningitis in 100% of the cases proved to be serum procalcitonin with a threshold value of 0.93 ng/ml . CONCLUSION: Serum procalcitonin is an interesting parameter in the emergency department for management of meningitis suspicion in adults. Indian J Pediatr, 1998 Nov-Dec, 65(6), 841 - 8 Perinatal bacterial infection: an update; Yeagley TJ et al.; This article reviews the current trends in the evaluation and management of bacterial infection involving the uterus, placenta, membranes, amniotic fluid, and fetus occurring near the time of birth . The discussion includes information regarding risk, incidence, pathophysiology, bedside diagnosis, interventional options including antibiotics, corticosteroids, fetal monitoring, and delivery, and possible preventive measures which affect the outcome . The adequate evaluation and management of perinatal infection requires a team approach with obstetricians and pediatricians . Clinical screening is useful in developing the diagnosis, but amniotic fluid evaluation remains the proposed gold standard . The role of cytokines is becoming increasingly important, as is seen in the association of IL-6 with positive amniotic fluid cultures and periventricular leukomalacia . Prompt recognition and management of the pregnancy affected by infection can improve perinatal outcomes . A management protocol is presented to help structure the approach to suspected infection . Premature delivery due to perinatal infection may be preventable. Digestion, 2000, 61(3), 165 - 71 Comparison of the 1-gram (14)C-D-xylose breath test and the 50-gram hydrogen glucose breath test for diagnosis of small intestinal bacterial overgrowth; Stotzer PO et al.; BACKGROUND/AIMS: Culture of small bowel aspirate is the most direct method and the gold standard for diagnosing small intestinal bacterial overgrowth . However, cultures are cumbersome and fluoroscopy is required for obtaining aspirate . Therefore, different breath tests such as the xylose breath test and the hydrogen breath test have been developed . There is no general agreement as to which test is to be preferred . In the only previous direct comparison between these two tests an advantage for the 1-gram-(14)C-D-xylose breath test was found . The aim of the study was to compare the 50-gram glucose hydrogen breath test and the 1-gram (14)C-D-xylose breath test in relation to results of cultures of small bowel aspirate . METHODS: Forty-six consecutive patients, mean age 57 (range 27-87) years, 12 men and 34 women, were included because of suspicion of small intestinal bacterial overgrowth . After small bowel aspiration, all patients received a solution of 1 g xylose, labelled with 50 microg (14)C-D-xylose, and 50 g glucose dissolved in 250 ml water . The concentration of breath hydrogen was analyzed every 15 min for 2 h and (14)CO(2) was analyzed every 30 min for 4 h . A positive hydrogen breath test was defined as a rise in hydrogen concentration of 15 ppm . A positive xylose test was defined as an accumulated dose 4.5% after 4 h . Two definitions for a positive culture were used, either growth of 10(5 )colonic-type bacteria/ml or growth of 10(5) bacteria/ml of any type . RESULTS: Twenty-four patients had growth of 10(5) bacteria, of whom 10 had growth of 10(5) colonic-type bacteria in small bowel aspirate . Twenty-two patients had no significant growth . The hydrogen breath test and the xylose breath test had a sensitivity for growth of 10(5) bacteria of 58 and 42%, respectively . For growth of 10(5 )colonic-type bacteria the sensitivity was 90% for the hydrogen breath test and 70% for the xylose breath test . The specificity was similar for the two tests . CONCLUSION: Although no significant difference between the two tests was found, there was a tendency in favor of the 50-gram glucose hydrogen breath test . The simplicity in combination with high sensitivity makes the hydrogen breath test suitable as a screening method to select patients for further investigation . Neurosci Lett, 2000 Apr 28, 284(3), 159 - 62 Effects of bacterial lipopolysaccharide on central monoamines and fever in the rat: involvement of the vagus; MohanKumar SM et al.; Lipopolysaccharide (LPS) is known to produce a number of central and neuroendocrine effects but the mechanisms involved are still unclear . This study was done to investigate the possibility that LPS-induced fever and activation of central monoamines are mediated through the vagus . Adult male rats were subjected to sub-diaphragmatic vagotomy (SDV), or sham operation and treated with saline or LPS in saline (10 microg/kg bw) 2 h later . Rectal temperature was monitored at half-hourly intervals for 5 h after which the animals were sacrificed and monoamine concentrations in hypothalamic nuclei were measured using HPLC-EC . SDV delayed the rise in rectal temperature induced by LPS by 1 h when compared to Sham animals . It also increased the concentrations of monoamines in the paraventricular nucleus of both Sham and SDV rats . This indicates that routes other than the vagus probably mediate LPS' actions on the central nervous system. Transfus Sci, 2000 Feb-Apr, 22(1-2), 139 - 43 Does bacterial contamination of platelet concentrates influence the leucocyte content and the rate of platelet storage lesion? Seghatchian J, Krailadsiri P, Beard M. An observational study was carried out to assess the effect of bacterial growth on changes in platelet cellular indices and leucocyte content . Platelet derived from pooled buffy coats, with and without an additional leucocyte removal step by filtration and platelet derived from Cobe LRS system were used . These were spiked with two doses of several types of bacteria (1 and 50 CFU) in a paired control and test experiment . The changes in cellular parameters were monitored by an automated cell counter (Sysmex SE-9000) and the concentration of the residual leucocyte were evaluated by two automated techniques, based on DNA staining principles (flow cytometry-EPICS-XL Coutter and Imagn 2000) . Our results indicate that initially bacterial growth is associated with a decrease in leucocyte count, followed by a concomitant increase in platelet large cell ratio, possibly due to aggregate formation . During the prolonged storage a dramatic increase in pseudoleucocytes was observed by both Imagn 2000 and flow cytometry techniques with an abnormal dot plot around FL3 regions in the latter counting method, making the true identification of native leucocytes rather difficult . It is concluded that bacterial growth is associated with both changes in platelet cellular indices and development of cellular aggregates and/or partially fragmented cells with DNA binding properties appearing as pseudoleucocytes . Further work on the true nature of so called 'pseudoleucocyte' is in progress. Indian J Pediatr, 1997 Jul-Aug, 64(4), 517 - 22 Dexamethasone as an adjunctive treatment of bacterial meningitis; Shembesh NM et al.; This study was conducted on 77 Libyan infants and children aged month to 10 years with acute bacterial meningitis . Upon admission, the patients were divided randomly in two groups . Group I (38 patients) received ceftriaxone plus dexamethasone i.v . Group II (39 patients) received ceftriaxone alone . Both groups were compared for mean changes in CSF sugar, CSF protein and CSF polymorph count at 4th day of treatment . There was a significant difference between the two groups in CSF sugar and protein changes (P < 0.05) but not in CSF polymorph (P > 0.05) . Both groups showed prompt clinical response and similar occurrence of acute complications, fatality rate and permanent neurological sequelae . However, group I manifested shorter duration of fever (P < 0.05) . Dexamethasone improved the inflammatory reaction in acute bacterial meningitis and shortened the duration of fever but it did not have any significant effect on the fatality and the occurrence of neurological sequelae of this disease. Anticancer Res, 2000 Jan-Feb, 20(1A), 357 - 62 Anti-tumor effect of N-beta-alanyl-5-S-glutathionyldihydroxyphenylalanine (5-S-GAD), a novel anti-bacterial substance from an insect; Akiyama N et al.; PURPOSE: We examined the anti-tumor effect of 5-S-GAD, a novel potent inhibitor of protein tyrosine kinases, isolated from the flesh fly in order to investigate the potential use of this compound as an anti-tumor agent . METHODS: In vitro growth inhibition was evaluated using the alamarBlue assay kit . In vivo anti-tumor activity was evaluated by i.p . treatment of 5-S-GAD against xenografted melanoma (LOX-IMV1) and breast carcinoma (MDA-MB-435S) in nude mice . RESULTS: Of 38 human cancer cell lines examined, this compound showed significant cytotoxicity toward two estrogen-negative breast carcinomas (MDA-MB-231 and MDA-MB-435S) and one malignant melanoma (LOX-IMV1) in vitro, indicating that it exhibits selective cytotoxicity to certain tumor cell lines . In accordance with its in vitro anti-tumor effect, 5-S-GAD was shown to significantly repress the growth of sensitive tumor cells in nude mice . CONCLUSION: These results indicate that 5-S-GAD is potentially useful to treat certain human cancer. Zentralbl Chir, 2000, 125(3), 275 - 9 {Primary bacterial aortitis}; Tiesenhausen K et al.; Primary bacterial aortitis represents a rare disease with a high lethality . From June 1997 to April 1999 5 patients with an abdominal aortic infection were treated by resection of the infected aorta and in-situ reconstruction or by extra-anatomic bypass . There was no treatment in one case because of the infaust prognosis . 3 patients survived, one with a paraparesis as a result of spinal ischemia . On the basis of our patients the pathogenesis, clinical symptoms with diagnosis and the therapeutic options are discussed. Minerva Stomatol, 1999 Nov, 48(11), 509 - 23 Bacterial adhesion to dental alloys . The role of the surface and composition; Capopreso S et al.; BACKGROUND: In the oral cavity, many bacteria can only survive by adhering to hard surfaces . The roughness and free energy of these surfaces play an important part in this process . Precision dental alloys may undergo corrosion, but findings show that this does not seem to cause problems of biocompatibility . The release of metallic ions into the oral cavity may both inhibit bacterial growth and influence bacterial adhesion . The object of the present study was to bring to light any possible correlation between corrosion and/or ionic release and bacterial adhesion with regard to 18 different types of dental alloy, both before and after polishing . METHODS: Electrochemical analyses were carried out (cyclic potentiodynamic and potentiostatic polarisation tests) . Corrodible elements were analysed through Atomic Absorption Spectroscopy of specimens of each alloy . The inhibition of bacterial adhesion and growth was determined using bacteria specific to the oral cavity . RESULTS: All the alloys examined show a tendency towards spontaneous passivation with low values of anodic current . The evaluation of ionic release confirmed the biocompatibility of the tested materials and the solutions conditioned with the alloys did not inhibit bacterial growth . There was no significant bacterial adhesion after polishing . Bacteria adhere to unpolished alloys in a specific manner and are inhibited from doing so in the presence of alloys for gold-resin restorations containing silver and copper . CONCLUSIONS: When polished, all the alloys are resistant to in vitro electrochemical corrosion and bacterial adhesion . The possibility cannot be excluded that bacterial adhesion occurs after the materials have been in place in the oral cavity for some time . The alloys which were found to inhibit bacterial attack may be more suitable, while not representing a biological risk for the surrounding tissues. FEBS Lett, 2000 Apr 14, 471(2-3), 165 - 8 Scorpine, an anti-malaria and anti-bacterial agent purified from scorpion venom; Conde R et al.; A novel peptide, scorpine, was isolated from the venom of the scorpion Pandinus imperator, with anti-bacterial activity and a potent inhibitory effect on the ookinete (ED(50) 0.7 microM) and gamete (ED(50) 10 microM) stages of Plasmodium berghei development . It has 75 amino acids, three disulfide bridges with a molecular mass of 8350 Da . Scorpine has a unique amino acid sequence, similar only to some cecropins in its N-terminal segment and to some defensins in its C-terminal region . Its gene was cloned from a cDNA library. Oncol Rep, 2000 May-Jun, 7(3), 645 - 9 Cloning, characterisation and bacterial expression of full length cDNA for the mouse liver microsomal glutathione S-transferase; Raza H et al.; We have isolated a cDNA encoding full length microsomal glutathione S-transferase (MGST) from mouse liver . The cDNA was isolated by RT-PCR using primers designed from published cDNA sequence of rat MGST with the addition of 5' Nde-1 and 3' HindIII sites, and cloned into bacterial expression vector pSP19T7LT . Deduced amino acid sequence (155 amino acids, calculated mol.mass 17512 Dalton) confirmed the identity of microsomal GST from mouse liver which has sequence homology with that of rat and human liver MGST1 . Recombinant GST cDNA (Genbank accession # 159050) was expressed in BL21(DE3) in the presence of 1 mM IPTG at 30 degrees C . The expressed GST protein was found to be localised in the bacterial membrane as determined by measuring catalytic activity using CDNB and cumene hydroperoxide substrates, SDS-PAGE and Western blot analysis . We have demonstrated the cloning and expression of full length cDNA for MGST from mouse liver and have characterised the functionally active product as MGST protein . These results should facilitate studies on the role of MGST in the regulation of chemical carcinogenesis and in the prevention of oxidative stress caused by endogenous and exogenous chemicals. Hum Mol Genet, 2000 Apr 12, 9(6), 937 - 43 Analysis of mammalian central nervous system gene expression and function using bacterial artificial chromosome-mediated transgenesis; Heintz N; The anatomical complexity of the mammalian central nervous system (CNS) presents special problems for the analysis of CNS gene expression and function . The most difficult challenge is presented by the simple fact that there are hundreds of functionally and morphologically defined cell types in the CNS . Given this complexity, the interpretation of CNS phenotypes is often problematic . The preparation of transgenic mice carrying marked bacterial artificial chromosomes (BACs) provides an important avenue for improving our understanding of CNS-expressed genes and phenotypes . This approach can allow efficient analysis of patterns of gene expression, subcellular localization of their encoded products and neuronal projection patterns . BAC transgenic mice can also provide access to information relevant to gene function based on phenotypes arising from increased gene dosage or expression of activating and dominant-negative alleles . This review will concentrate on these issues and their relevance to the analysis of CNS-expressed genes. J Biol Chem, 2000 Apr 21, 275(16), 11740 - 9 Magnesium-induced linear self-association of the FtsZ bacterial cell division protein monomer . The primary steps for FtsZ assembly; Rivas G et al.; The bacterial cell division protein FtsZ from Escherichia coli has been purified with a new calcium precipitation method . The protein contains one GDP and one Mg(2+) bound, it shows GTPase activity, and requires GTP and Mg(2+) to polymerize into long thin filaments at pH 6.5 . FtsZ, with moderate ionic strength and low Mg(2+) concentrations, at pH 7.5, is a compact and globular monomer . Mg(2+) induces FtsZ self-association into oligomers, which has been studied by sedimentation equilibrium over a wide range of Mg(2+) and FtsZ concentrations . The oligomer formation mechanism is best described as an indefinite self-association, with binding of an additional Mg(2+) for each FtsZ monomer added to the growing oligomer, and a slight gradual decrease of the affinity of addition of a protomer with increasing oligomer size . The sedimentation velocity of FtsZ oligomer populations is compatible with a linear single-stranded arrangement of FtsZ monomers and a spacing of 4 nm . It is proposed that these FtsZ oligomers and the polymers formed under assembly conditions share a similar axial interaction between monomers (like in the case of tubulin, the eukaryotic homolog of FtsZ) . Similar mechanisms may apply to FtsZ assembly in vivo, but additional factors, such as macromolecular crowding, nucleoid occlusion, or specific interactions with other cellular components active in septation have to be invoked to explain FtsZ assembly into a division ring. Cancer Gene Ther, 2000 Mar, 7(3), 438 - 45 Gene therapy of metastatic colon carcinoma: regression of multiple hepatic metastases by adenoviral expression of bacterial cytosine deaminase; Block A et al.; Colon carcinoma accounts for 20% of deaths due to malignancies in the Western world . Once metastases occur, therapeutic options are limited, with an approximate 5-year survival of only 5% . To investigate the potential of new gene therapeutic approaches, a hepatic micrometastasis model of colon carcinoma in BALB/c mice was established . Inoculation of syngeneic MCA26 colon carcinoma cells into the spleens of 18- to 20-week-old mice resulted in the formation of multiple hepatic metastases . Selective transduction of developing hepatic metastases was demonstrated using a beta-galactosidase-expressing recombinant adenovirus . Cytosine deaminase (CD) can metabolize 5-fluorocytosine into the chemotherapeutic reagent 5-fluorouracil (5FU) . The antitumoral potential of this suicide gene therapy approach was explored by systemic application of a recombinant replication-deficient adenovirus encoding for the bacterial CD gene under the control of the cytomegalovirus promoter (Ad.CMV-CD) . Injection into the tail vein of tumor-bearing mice resulted in delayed tumor growth with significant reduction in hepatic metastases . The potential of this experimental approach for possible future clinical applications was evaluated by investigating adenoviral transduction efficiency, 5FU sensitivity, and 5-fluorocytosine-dependent Ad.CMV-CD toxicity in a variety of human colon cancer cell lines . Although the murine cell lines MCA26 and CC36 were highly sensitive to 5FU, the human colon cancer cell lines showed a 1-100 times higher resistance to 5FU . Specific Ad.CMV-CD toxicity correlates with 5FU toxicity . Transduction efficiency in human colon carcinoma cell lines was shown to be 10-1700 times higher compared with murine cell lines, thus compensating for 5FU resistance . In conclusion, suicide gene therapy using CD may be promising as an adjuvant treatment regimen for hepatic micrometastases of human colon carcinoma. Scott Med J, 2000 Feb, 45(1), 12 - 3 Bacterial contamination of children's toys used in a general practitioner's surgery; McKay I et al.; General practitioners--and other healthcare professionals are encouraged to make their premises child friendly . One way of doing this is to provide toys for children to use . We looked at the appearance and bacterial colonisation of 50 toys after a busy morning surgery in an inner city general practice . The toys appeared generally unclean and 10% were contaminated by potential pathogens . Bacteria were cultured more frequently from soft toys than from hard toys (odds ratio 8.14; 95% confidence range 0.74-107.49) . Although toys may appear to be physically dirty after use, the bacteria isolated from their surfaces are generally non-pathogenic to children with normal immune function and probably no worse than other objects in the environment . However, there does exist an appreciable (1 in 10) risk of cross-infection with the use of toys in a clinic . Toys with a hard surface are preferred as these are less likely to be contaminated and are more easily disinfected. FEBS Lett, 2000 Apr 7, 471(1), 7 - 11 Fine architecture of bacterial inclusion bodies; Carrio MM et al.; The molecular organisation of protein aggregates, formed under physiological conditions, has been explored by in vitro trypsin treatment and electron microscopy analysis of bacterially produced inclusion bodies (IBs) . The kinetic modelling of protein digestion has revealed variable proteolysis rates during protease exposure that are not compatible with a surface-restricted erosion of body particles but with a hyper-surfaced disintegration by selective enzymatic attack . In addition, differently resistant species of the IB proteins coexist within the particles, with half-lives that differ among them up to 50-fold . During in vivo protein incorporation throughout IB growth, a progressive increase of proteolytic resistance in all these species is observed, indicative of folding transitions and dynamic reorganisations of the body structure . Both the heterogeneity of the folding state and the time-dependent folding transitions undergone by the aggregated polypeptides indicate that IBs are not mere deposits of collapsed, inert molecules but plastic reservoirs of misfolded proteins that would allow, at least up to a certain extent, their in vivo recovery and transference to the soluble cell fraction. Mol Microbiol, 2000 Mar, 35(6), 1540 - 9 Death effector domain of a mammalian apoptosis mediator, FADD, induces bacterial cell death; Lee SW et al.; FADD is a mammalian pro-apoptotic mediator consisting of the N-terminal death effector domain (DED) and the C-terminal death domain (DD) . The N-terminal 88-residue fragment of murine FADD was defined as the stable structural unit of DED, as determined by proteolytic digestion and conformational analysis . This domain induced bacterial as well as mammalian cell death, whereas the full-length or DD of FADD did not . The Escherichia coli cells expressing FADD-DED showed elongated cell morphology and an increased level of nicked chromosomal DNA and mutation . The lethality of FADD-DED was abolished by co-expression of thioredoxin and superoxide dismutase or relieved by the addition of vitamin E as a reducing agent and under anaerobic growth conditions . The toxicity of FADD-DED was genetically suppressed by various oxidoreductases of E . coli . All these results suggest that the death effector domain of mammalian FADD induced bacterial cell death by enhancing cellular levels of reactive oxygen species (ROS). Mol Microbiol, 2000 Mar, 35(6), 1405 - 12 Homing of a bacterial group II intron with an intron-encoded protein lacking a recognizable endonuclease domain; Martinez-Abarca F et al.; RmInt1 is a functional group II intron found in Sinorhizobium meliloti where it interrupts a group of IS elements of the IS630-Tc1 family . In contrast to many other group II introns, the intron-encoded protein (IEP) of RmInt1 lacks the characteristic conserved part of the Zn domain associated with the IEP endonuclease activity . Nevertheless, in this study, we show that RmInt1 is capable of inserting into a vector containing the DNA spanning the RmInt1 target site from the genome of S . meliloti . Efficient homing was also observed in the absence of homologous recombination (RecA- strains) . In addition, it is shown that RmInt1 is able to move to its target in a heterologous host (S . medicae) . Homing of RmInt1 occurs very efficiently upon DNA target uptake (conjugation/electroporation) by the host cell resulting in a proportion of invaded target of 11-30% . Afterwards, the remaining intronless target DNA is protected from intron invasion. Genes Cells, 2000 Mar, 5(3), 155 - 67 Bacterial cell death induced by human pro-apoptotic Bax is blocked by an RNase E mutant that functions in an anti-oxidant pathway; Nanbu-Wakao R et al.; BACKGROUND: Bax is a member of the Bcl-2 family and induces apoptosis of mammalian cells . We have shown that a trace amount of human Bax induces the cell death of Escherichia coli, accompanied by damage to DNA, and that the region of Bax which is lethal to E . coli is also responsible for apoptosis-inducing activity in the mammalian cells . RESULTS: We isolated a Bax-resistant mutant from E . coli cells that survive in the presence of paraquat, a generator of superoxide, by screening a library constructed from the random insertion of a transposon . Psb1 (paraquat-resistant, suppressor of Bax-1) mutant had a Tn 10 transposon inserted in the rne gene of E . coli, splitting the RNase E gene (rne) into N- and C-terminal halves . The introduction of the truncated 5' end of rne specifically enhanced resistance to paraquat, prevented cell death induced by Bax and decreased the intracellular H2O2 concentration . The region responsible for the paraquat- and Bax-resistance was not the catalytic site for the endoribonuclease activity of RNase E . CONCLUSIONS: The N-terminal region of the RNase E protein inhibits bacterial death induced by human Bax as well as paraquat through a unique mechanism that is distinct from RNA digestion . This study implies that the protection of bacterial death induced by Bax is associated with an anti-oxidant pathway and that a mutant RNase E has a novel function as an anti-oxidant. Br J Surg, 2000 Apr, 87(4), 439 - 42 Bacterial translocation in patients undergoing abdominal aortic aneurysm repair; Woodcock NP et al.; BACKGROUND: Bacterial translocation occurs in humans and is associated with an increased incidence of septic morbidity . The aims of this study were to determine the prevalence of bacterial translocation in patients undergoing open abdominal aortic aneurysm (AAA) repair and to identify any association with postoperative septic complications . METHODS: This was a prospective observational study in which patients undergoing aneurysm repair were assessed for evidence of bacterial translocation by culture of a mesenteric lymph node (MLN), small bowel serosal exudate and thrombus within the aneurysm . All postoperative septic complications were recorded . RESULTS: A total of 51 patients was studied (40 men, 11 women; median age 72 years) . Enteric bacteria were isolated from the MLNs of five patients (prevalence of bacterial translocation 10 per cent), one of whom also yielded growth from the serosal exudate . Septic morbidity occurred in four of five patients in whom bacterial translocation was identified, compared with nine of 46 in those without translocation (P = 0.013, Fisher's exact test, mid P) . One patient in whom Escherichia coli was grown from the MLN developed an aortoenteric fistula, with a coliform species isolated from the graft . CONCLUSION: This study suggests that bacterial translocation occurs in patients undergoing AAA repair . It is associated with an increased incidence of postoperative septic morbidity and provides a possible mechanism for infection of prosthetic aortic grafts. Plant J, 2000 Feb, 21(3), 239 - 48 Functional rescue of a bacterial dapA auxotroph with a plant cDNA library selects for mutant clones encoding a feedback-insensitive dihydrodipicolinate synthase; Vauterin M et al.; Dihydrodipicolinate synthase (DHDPS; EC4.2.1.52) catalyses the first reaction of lysine biosynthesis in plants and bacteria . Plant DHDPS enzymes are strongly inhibited by lysine (I0.5 approximately 10 microM), whereas the bacterial enzymes are less (50-fold) or insensitive to lysine inhibition . We found that plant dhdps sequences expressing lysine-sensitive DHDPS enzymes are unable to complement a bacterial auxotroph, although a functional plant DHDPS enzyme is formed . As a consequence of this, plant dhdps cDNA clones which have been isolated through functional complementation using the DHDPS-deficient Escherichia coli strain encode mutated DHDPS enzymes impaired in lysine inhibition . The experiments outlined in this article emphasize that heterologous complementation can select for mutant clones when altered protein properties are requisite for functional rescue . In addition, the mutants rescued by heterologous complementation revealed a new critical amino acid substitution which renders lysine insensitivity to the plant DHDPS enzyme . An interpretation is given for the impaired inhibition mechanism of the mutant DHDPS enzyme by integrating the identified amino acid substitution in the DHDPS protein structure. J Am Coll Surg, 2000 Apr, 190(4), 423 - 31 Beneficial effects of growth hormone and insulin-like growth factor I on intestinal bacterial translocation, endotoxemia, and apoptosis in experimentally jaundiced rats; Scopa CD et al.; BACKGROUND: This study was undertaken to investigate the effect of growth hormone (GH) and insulin-like growth factor I (IGF-I), two well-known growth factors, on bacterial translocation, endotoxemia, enterocyte apoptosis, and intestinal and liver histology in a model of experimental obstructive jaundice in rats . STUDY DESIGN: One hundred six male Wistar rats were divided into five groups: I (n = 21), controls; II (n = 22), sham operated; III (n = 22), bile duct ligation (BDL); IV (n = 21), BDL and GH treatment; and V (n = 20), BDL and IGF-I administration . By the end of the experiment, on day 10, blood bilirubin was determined, and mesenteric lymph nodes, liver specimens, and bile from the bile duct stump were cultured . Endotoxin was measured in portal and aortic blood . Tissue samples from the terminal ileum and liver were examined histologically and apoptotic body count (ABC) in intestinal mucosa was evaluated . Mucosal DNA and protein content were also determined . RESULTS: Bilirubin increased significantly after BDL (p < 0.001) . Bile from the bile duct was sterile . In group III, MLN and liver specimens were contaminated by gut origin bacteria (significant versus group I and II, p < 0.001, respectively) . GH reduced significantly positive cultures (p < 0.01), and IGF-I had no effect . BDL resulted in significant increase in portal and aortic endotoxemia (p < 0.001); treatment with GH and IGF-I reduced it (p < 0.001) . Mucosal DNA and protein content were reduced in animals with BDL and after treatment with GH or IGF-I; an increase to almost normal levels was noted in DNA, but not in protein . Overall the ileal architecture remained intact in all animal groups . The ABC increased after BDL . After GH and IGF-I administration, the ABC decreased significantly, and there was no difference between GH and IGF-I treated animals . After BDL, liver biopsies displayed typical changes of biliary obstruction, which were significantly improved after administration of GH and IGF-I . CONCLUSIONS: Treatment with GH and IGF-I in rats with experimental obstructive jaundice reduces endotoxemia, and it improves liver histology . Apoptosis, in the intestinal epithelium, may serve as a morphologic marker of the ileal mucosal integrity, demonstrating the proliferative potential of GH and IGF-I in cases of obstructive jaundice, and this might be of potential value in patients with such conditions. Rev Med Chir Soc Med Nat Iasi, 1999 Jul-Dec, 103(3-4), 210 - 5 {A comparative study of the bacterial dental plaque formation on fixed dentures with different surface characteristics}; Forna N et al.; The accumulation of dental plaque on fixed prostheses made of various biomaterials is influenced by the roughness of the surface . For the investigation of this aspects it was necessary of a method rapid, facile and cheaper in comparison with bacterial culturing and identification which is laborious and expansive . In this study we followed the accumulation of dental plaque on biomaterials like chrome-cobalt and gaudent polished and unpolished, at different periods of time (3 and 14 days) . We observed that after 3 days the aspect is predominant coccoid and after 14 days is predominant bacillary . On unpolished crowns the width of dental plaque is greater and on gaudent crowns the plaque is structural organised around of filamentous organisms. Acta Microbiol Pol, 1999, 48(3), 243 - 59 Technical aspects of random amplified polymorphic DNA (RAPD) technique in genotyping of bacterial strains; Gzyl A et al.; Random Amplified Polymorphic DNA (RAPD) method became widely applied for sensitive, efficient and fast distinguishing of different isolates of a given species, if pure culture is available . Problems with reproducibility and discriminatory power, frequently cited in the literature, can be overcome by precise optimization procedure allowing to achieve reliable conditions for each species analysed . Basing on two examples of different species, H . pylori and E . faecium, particular parameters of RAPD fingerprinting were evaluated with respect to selection of best working primers generating medium-complex profiles, using only high quality DNA samples and evaluation optimum for every reaction reagent . Stable and informative amplification patterns were obtained with different best working primers which could discriminate between all H . pylori and E . faecium strains tested . For both analysed species different optima were found, suggesting species-specific need of precise RAPD conditions evaluation . This study proved high sensitivity and efficiency of optimized RAPD profiling applicable for searching the epidemiology traces for both species. J Biotechnol, 2000 Mar 31, 78(3), 209 - 19 Implication of gene distribution in the bacterial chromosome for the bacterial cell factory; Rocha EP et al.; As bacterial genome sequences accumulate, more and more pieces of data suggest that there is a significant correlation between the distribution of genes along the chromosome and the physical architecture of the cell, suggesting that the map of the cell is in the chromosome . Considering sequences and experimental data indicative of cell compartmentalisation, mRNA folding and turnover, as well as known structural features of protein and membrane complexes, we show that preliminary in silico analysis of whole genome sequences strongly substantiates this hypothesis . If there is a correlation between the genome sequence and the cell architecture, it must derive from some selection pressure in the organisms growing in the wild . As a consequence, the underlying constraints should be optimised in genetically modified organisms if one is to expect high product yields . Consequences in terms of gene expression for biotechnology are straightforward: knocking genes out and in genomes should not be randomly performed, but should follow the rules of chromosome organisation. J Pediatr Gastroenterol Nutr, 2000, 30 Suppl 2, S2 - 7 Role of nutrients and bacterial colonization in the development of intestinal host defense; Walker WA; In this introduction to the supplement on the use of pre- and probiotics in the health and disease of pediatric patients, I have summarized factors affecting the initial colonization of the neonatal intestine . The term bacterial-epithelial cross-talk was defined, and examples of the enterocyte response to both pathologic and indigenous flora stimulation illustrated . Immaturities in the human neonatal intestinal response to bacteria and their toxins were reviewed in the context of the pathogenesis of age-specific, bacterial gastrointestinal infectious diseases . Finally, the importance of pre- and probiotics as measures to strengthen the neonate's intestinal host defenses in the prevention and treatment of specific age-related disease were considered. J Biol Chem, 2000 Jun 30, 275(26), 19752 - 8 Correlated switch binding and signaling in bacterial chemotaxis; Schuster M et al.; In Escherichia coli, swimming behavior is mediated by the phosphorylation state of the response regulator CheY . In its active, phosphorylated form, CheY exhibits enhanced binding to a switch component, FliM, at the flagellar motor, which induces a change from counterclockwise to clockwise flagellar rotation . When Ile(95) of CheY is replaced by a valine, increased clockwise rotation correlates with enhanced binding to FliM . A possible explanation for the hyperactivity of this mutant is that residue 95 affects the conformation of nearby residues that potentially interact with FliM . In order to assess this possibility directly, the crystal structure of CheY95IV was determined . We found that CheY95IV is structurally almost indistinguishable from wild-type CheY . Several other mutants with substitutions at position 95 were characterized to establish the structural requirements for switch binding and clockwise signaling at this position and to investigate a general relationship between the two properties . The various rotational phenotypes of these mutants can be explained solely by the amount of phosphorylated CheY bound to the switch, which was inferred from the phosphorylation properties of the mutant CheY proteins and their binding affinities to FliM . Combined genetic, biochemical, and crystallographic results suggest that residue 95 itself is critical in mediating the surface complementarity between CheY and FliM. J Biol Chem, 2000 Jun 9, 275(23), 17510 - 6 A dimer as a building block in assembling RNA . A hexamer that gears bacterial virus phi29 DNA-translocating machinery; Chen C et al.; Six RNA (pRNA) molecules form a hexamer, via hand-in-hand interaction, to gear bacterial virus phi29 DNA translocation machinery . Here we report the pathway and the conditions for the hexamer formation . Stable pRNA dimers and trimers were assembled in solution, isolated from native gels, and separated by sedimentation, providing a model system for the study of RNA dimers and trimers in a protein-free environment . Cryo-atomic force microscopy revealed that monomers displayed a check mark outline, dimers exhibited an elongated shape, and trimers formed a triangle . Dimerization of pRNA was promoted by a variety of cations including spermidine, whereas procapsid binding and DNA packaging required specific divalent cations, including Mg(2+), Ca(2+), and Mn(2+) . Both the tandem and fused pRNA dimers with complementary loops designed to form even-numbered rings were active in DNA packaging, whereas those without complementary loops were inactive . We conclude that dimers are the building blocks of the hexamer, and the pathway of building a hexamer is: dimer --> tetramer --> hexamer . The Hill coefficient of 2.5 suggests that there are three binding sites with cooperative binding on the surface of the procapsid . The two interacting loops played a key role in recruiting the incoming dimer, whereas the procapsid served as the foundation for hexamer assembly. Curr Opin Microbiol, 2000 Apr, 3(2), 203 - 9 The biogenesis and assembly of bacterial membrane proteins; Bernstein HD; Bacterial proteins in the inner and outer membranes differ dramatically in their architecture . Although both types of proteins are transported across the inner membrane through a common pore, recent studies have identified distinct factors that target them to transport sites and catalyze proper folding. Microbes Infect, 2000 Feb, 2(2), 199 - 211 Bacterial modulation of antigen processing and presentation; Maksymowych WP et al.; This review examines the mechanisms by which bacteria influence the antigenic processing of endogenous and exogenous antigens presented by class I, class II, and nonclassical MHC molecules . Consequent effects on presentation of bacterial antigens, the ability of bacteria to evade host defences, and the potential induction of autoimmunity are discussed. Microbes Infect, 2000 Feb, 2(2), 157 - 66 Temperature-regulated expression of bacterial virulence genes; Konkel ME et al.; Virulence gene expression in most bacteria is a highly regulated phenomenon, affected by a variety of parameters including osmolarity, pH, ion concentration, iron levels, growth phase, and population density . Virulence genes are also regulated by temperature, which acts as an 'on-off' switch in a manner distinct from the more general heat-shock response . Here, we review temperature-responsive expression of virulence genes in four diverse pathogens. Appl Environ Microbiol, 2000 Apr, 66(4), 1685 - 91 Bacterial symbiont transmission in the wood-boring shipworm Bankia setacea (Bivalvia: Teredinidae); Sipe AR et al.; The Teredinidae (shipworms) are a morphologically diverse group of marine wood-boring bivalves that are responsible each year for millions of dollars of damage to wooden structures in estuarine and marine habitats worldwide . They exist in a symbiosis with cellulolytic nitrogen-fixing bacteria that provide the host with the necessary enzymes for survival on a diet of wood cellulose . These symbiotic bacteria reside in distinct structures lining the interlamellar junctions of the gill . This study investigated the mode by which these nutritionally essential bacterial symbionts are acquired in the teredinid Bankia setacea . Through 16S ribosomal DNA (rDNA) sequencing, the symbiont residing within the B . setacea gill was phylogenetically characterized and shown to be distinct from previously described shipworm symbionts . In situ hybridization using symbiont-specific 16S rRNA-directed probes bound to bacterial ribosome targets located within the host gill coincident with the known location of the gill symbionts . These specific probes were then used as primers in a PCR-based assay which consistently detected bacterial rDNA in host gill (symbiont containing), gonad tissue, and recently spawned eggs, demonstrating the presence of symbiont cells in host ovary and offspring . These results suggest that B . setacea ensures successful inoculation of offspring through a vertical mode of symbiont transmission and thereby enables a broad distribution of larval settlement. Appl Environ Microbiol, 2000 Apr, 66(4), 1444 - 52 Imaging the enzymatic digestion of bacterial cellulose ribbons reveals the endo character of the cellobiohydrolase Cel6A from Humicola insolens and its mode of synergy with cellobiohydrolase Cel7A; Boisset C et al.; Dispersed cellulose ribbons from bacterial cellulose were subjected to digestion with cloned Cel7A (cellobiohydrolase {CBH} I) and Cel6A (CBH II) from Humicola insolens either alone or in a mixture and in the presence of an excess of beta-glucosidase . Both Cel7A and Cel6A were effective in partially converting the ribbons into soluble sugars, Cel7A being more active than Cel6A . In combination, these enzymes showed substantial synergy culminating with a molar ratio of approximately two-thirds Cel6A and one-third Cel7A . Ultrastructural transmission electron microscopy (TEM) observations indicated that Cel7A induced a thinning of the cellulose ribbons, whereas Cel6A cut the ribbons into shorter elements, indicating an endo type of action . These observations, together with the examination of the digestion kinetics, indicate that Cel6A can be classified as an endo-processive enzyme, whereas Cel7A is essentially a processive enzyme . Thus, the synergy resulting from the mixing of Cel6A and Cel7A can be explained by the partial endo character of Cel6A . A preparation of bacterial cellulose ribbons appears to be an appropriate substrate, superior to Valonia or bacterial cellulose microcrystals, to visualize directly by TEM the endo-processivity of an enzyme such as Cel6A. J Biochem (Tokyo), 2000 Apr, 127(4), 673 - 9 Functional construction of the anti-mucin core protein (MUC1) antibody MUSE11 variable regions in a bacterial expression system; Asano R et al.; A bacterial expression system for the variable region fragments (Fvs) of the anti-MUC1 tumor antigen antibody MUSE11 has been constructed . The Fv fragment showed binding specificity toward TFK-1 cells, with slightly reduced affinity compared to its parent IgG . The single-chain Fv fragment was arranged in two orders, VH-linker-VL and VL-linker-VH . However, linking the regions with a flexible peptide linker (GGGGS)(3) or with a shorter linker (GGGGS) led to a dramatic decrease in the biological activity toward the target antigen in both arrangements, suggesting that the MUSE11 antibody loses its activity when the domains are linked with polypeptide linkers . These results indicate that the variable region domains of the anti-MUC1 antibody MUSE11 have specificity only in the Fv form, and that linking the domains strongly reduces the association with its target antigen . Gel filtration analysis indicates that the scFv has a dimeric structure, suggesting that the inactivation of MUSE11 scFv is due to unfavorable intermolecular associations of the scFv chains . To our knowledge, this is the first report of a significant reduction in affinity caused by linking the variable domains in both arrangements, i.e., VH-VL and VL-VH. J Biochem (Tokyo), 2000 Apr, 127(4), 525 - 30 Expression of cytokines in bacterial and viral infections and their biochemical aspects; Imanishi J; Cytokines are very important in the host defense system, and play a critical role in protection against bacterial and viral infections . Cytokines are also involved in the pathogenesis and development of symptoms in infections . In this article, Helicobacter pylori (H . pylori) infection as bacterial infection, and influenza virus infection, encephalomyocarditis virus (EMCV) infection, and herpes simplex virus (HSV) infection as viral infection are mentioned . In H . pylori infection, various chemokines, especially interleukin (IL)-8, induce inflammatory responses in the gastroduodenal mucosa . Furthermore, IL-6, IL-7, tumor necrosis factor (TNF)-alpha, and interferon (IFN)-gamma are involved in both protection and pathogenesis . In influenza virus infection, IFN-alpha/beta, IFN-gamma, and IL-6 play protective roles . In EMCV infection, IL-6 and TNF-alpha play important roles as a protective and exacerbative factor in acute myocarditis, respectively . Furthermore, in HSV infection, the production of inflammatory cytokines is closely correlated with the pathogenesis of herpetic keratitis, and IFN-gamma plays an important role in enhancing viral clearance from the cornea and trigeminal ganglions. J Microbiol Methods, 2000 Mar, 40(1), 89 - 97 Quantification of bacterial adhesion forces using atomic force microscopy (AFM); Fang HH et al.; This study demonstrated that atomic force microscopy (AFM) can be used to obtain high-resolution topographical images of bacteria, and to quantify the tip-cell interaction force and the surface elasticity . Results show that the adhesion force between the Si3N4 tip and the bacteria surface was in the range from -3.9 to -4.3 nN . On the other hand, the adhesion forces at the periphery of the cell-substratum contact surface ranged from -5.1 to -5.9 nN and those at the cell-cell interface ranged from -6.5 to -6.8 nN . The two latter forces were considerably greater than the former one, most likely due to the accumulation of extracellular polymer substance (EPS) . Results also show that the elasticity varied on the cell surface. Proc Natl Acad Sci U S A, 2000 Apr 11, 97(8), 3965 - 70 Predicting ligand-binding function in families of bacterial receptors; Johnson JM et al.; The three-dimensional fold of a new protein sequence can often be inferred directly from sequence homology to a protein of known structure . The function of a new protein sequence is more difficult to predict, however, since homologues can have different molecular and cellular functions . To develop and automate computational methods for determining molecular function, we have analyzed ligand-binding specificity in two related families of binding proteins . One of these families includes Escherichia coli lactose repressor and ribose-binding protein, and the other includes E . coli sulfate- and phosphate-binding proteins . These proteins have similar folds but varying specificity, binding many different small molecules, including mono- and disaccharides, purines, oxyanions, ferric iron, and polyamines . Starting from template structural alignments, alignments of over 90 sequences per family were generated by iterative database searches with hidden Markov models . Phylogenetic trees were made of full-length sequences and of subsets of residues lining the binding cleft, to determine whether subbranches of the trees correlate with ligand-binding preference . Automated analyses of residues in the binding pocket were also used to predict ligand-binding function for many uncharacterized database sequences and to identify specific side chain-ligand contacts in proteins without solved structures . Our results demonstrate the utility of anchoring functional annotation within a protein family context. Akush Ginekol (Sofiia), 1999, 38(3), 14 - 6 {Therapeutic regimens for treating bacterial vaginosis in pregnant women}; Borisov I et al.; The study comprises 128 pregnant women examined at different gestational weeks . The diagnosis of bacterial vaginosis was made using: a) the complex clinical criteria--vaginal discharge, vaginal pH, amine test and "clue cells" b) Nugent scoring system c) Spiegel criteria . Two therapeutic regimens were compared--intravaginal 2% clindamycin creme (Dalacin V) 5 g three consecutive days and intravaginal metronidazole (Flagyl) 500 mg once daily for 5 consecutive days . Control examination was carried out 5-7 days after completion of therapy using the same protocol . 28 women from the first group and 31 women from the second group had the control examination . Bacterial vaginosis was eradicated in 93% of women using intravaginal clindamycin and in 87% of women using intravaginal metronidazole . Both regimes were more effective compared to treatment with oral ampicillin for 7 days, where the cure rate was 62%. Theriogenology, 1999 Aug, 52(3), 413 - 23 Uterine contractility is necessary for the clearance of intrauterine fluid but not bacteria after bacterial infusion in the mare; Nikolakopoulos E et al.; Bacteria were infused into the uteri of 5 estrous mares resistant to persistent mating-induced endometritis, first during a control cycle, and then during treatment with clenbuterol, a beta 2 agonist . Uterine cellular response was evaluated 48 h later by transrectal ultrasonography, followed by uterine lavage . During clenbuterol treatment all mares accumulated intrauterine fluid, whereas in the control cycle none of the mares retained fluid . There was no significant difference between the 2 cycles in the cloudiness of the lavage fluid, number of cells per milliliter, percentage of neutrophils and frequency of bacterial growth from the recovered fluid . We conclude that uterine contractility is important in the clearance of uterine fluid, but not necessarily for the elimination of bacteria, thus supporting the published evidence that impaired uterine contractility contributes to the pathogenesis of persistent mating-induced endometritis. Biochem Biophys Res Commun, 2000 Apr 2, 270(1), 46 - 51 Localization of the type I restriction-modification enzyme EcoKI in the bacterial cell; Holubova I et al.; To localise the type I restriction-modification (R-M) enzyme EcoKI within the bacterial cell, the Hsd subunits present in subcellular fractions were analysed using immunoblotting techniques . The endonuclease (ENase) as well as the methylase (MTase) were found to be associated with the cytoplasmic membrane . HsdR and HsdM subunits produced individually were soluble, cytoplasmic polypeptides and only became membrane-associated when coproduced with the insoluble HsdS subunit . The release of enzyme from the membrane fraction following benzonase treatment indicated a role for DNA in this interaction . Trypsinization of spheroplasts revealed that the HsdR subunit in the assembled ENase was accessible to protease, while HsdM and HsdS, in both ENase and MTase complexes, were fully protected against digestion . We postulate that the R-M enzyme EcoKI is associated with the cytoplasmic membrane in a manner that allows access of HsdR to the periplasmic space, while the MTase components are localised on the inner side of the plasma membrane . Protein Expr Purif, 2000 Apr, 18(3), 310 - 5 Bacterial expression and purification of the Arabidopsis NADPH-cytochrome P450 reductase ATR2; Hull AK et al.; An N-terminally modified form of the Arabidopsis NADPH-cytochrome P450 ATR2 (ATR2mod) was expressed from the tactac promoter in Escherichia coli to obtain high yields of the enzyme . The N-terminal modification eliminates the predicted chloroplast transit peptide of ATR2 allowing for more efficient expression . ATR2mod was purified from membrane extracts using a 2',5'-ADP-agarose affinity column . The specific activity of the purified ATR2mod for cytochrome c reduction was 9.4 micromol min(-1) mg(-1) and the K(m) for cytochrome c reduction was 15 +/- 2 microM . The purified NADPH-cytochrome P450 reductase was able to support function of CYP79B2 . Insect Biochem Mol Biol, 2000 Mar, 30(3), 253 - 8 Genomic copy number of intracellular bacterial symbionts of aphids varies in response to developmental stage and morph of their host; Komaki K et al.; Buchnera, endosymbiotic bacteria of aphids possess many genomic copies per cell . In this study, we estimated genomic copy number per Buchnera cell from host insects at various developmental stages and of two different morphs, apterae and alatae, by fluorimetry and real-time quantitative PCR . The results indicated that the genomic copy number of Buchnera increased during postembryonic development of insects to adulthood, and that it decreased during the host's ageing . In Buchnera from alatae, the genomic copy number per cell was about twice as many as in those from apterae . DAPI-staining showed that the distribution of the genomic DNA in the Buchnera cells from old insects tended to aggregate, suggesting that intracellular structure of the genomic DNA of Buchnera varies in response to the physiological conditions of their host. J Biochem (Tokyo), 2000 Mar, 127(3), 435 - 41 Convenient and efficient in vitro folding of disulfide-containing globular protein from crude bacterial inclusion bodies; Futami J et al.; We investigated how the folding yield of disulfide-containing globular proteins having positive net charges from crude bacterial inclusion bodies was affected by additives in the folding buffer . In screening folding conditions for human ribonucleases and its derivative, we found that addition of salt (about 0.4 M) to a folding buffer increased the folding yield . This suggested that electrostatic interaction between polyanionic impurities such as nucleic acids and cationic unfolded protein led to the formation of aggregates under the low-salt conditions . Since inclusion bodies were found to contain nucleic acids regardless of the electrostatic nature of the expressed protein, the electrostatic interaction between phosphate moieties of nucleic acids and basic amino acid residues of a denatured protein may be large enough to cause aggregation, and therefore the addition of salt in a folding buffer may generally be useful for promotion of protein folding from crude inclusion bodies . We further systematically investigated additives such as glycerol, guanidium chloride, and urea that are known to act as chemical chaperons, and found that these additives, together with salt, synergistically improved folding yield . This study, suggesting that the addition of salt into the folding buffer is one of the crucial points to be considered, may pave the way for a systematic investigation of the folding conditions of disulfide-containing foreign proteins from crude bacterial inclusion bodies. Transfus Clin Biol, 2000 Feb, 7(1), 15 - 23 {Blood transfusion and bacterial risk}; Morel P et al.; Initial hemovigilance data confirm the incidence and severity of transfusion reactions due to the bacterial contamination of blood components (TRBCs) . With 18 deaths reported through the French hemovigilance network over the past five years, bacterial risks represent one of the major immediate complications of BC transfusion . BC contamination may lead to more or less severe TRBCs, depending on their origin: bacteria growth, the BC itself or unknown origin . Although the rate of donated blood or BC contamination is known (0.5% and 0.05%, respectively), it is still difficult to assess the actual incidence of TRBCs, as it is difficult to identify and relate them to transfusion . Likewise, a better knowledge of bacteria, symptoms, and outcome is required to improve prevention methods . Better prevention can reduce BC contamination and proliferation of bacteria at each stage of blood transfusion . Methods of detecting BC contamination are still under investigation . Through continuous education of hemovigilance participants in identifying and dealing with TRBCs, as well as drawing up procedures to perform inquiries and specific bacterial analyses, case reporting can be further improved, in order to achieve more efficient prevention. Essays Biochem, 1999, 34, 17 - 30 Bacterial detoxification of Hg(II) and organomercurials; Miller SM; The most common bacterial mechanism for resistance to mercuric-ion species involves intracellular reduction of Hg(II) to Hg(0) . Key proteins of the pathway typically include: MerR, which regulates pathway expression; MerP, which protects the external environment; MerT or MerC, which transport Hg(II) species across the inner membrane; MerA, which catalyses reduction of Hg(II); and sometimes MerB, which catalyses cleavage of C-Hg bonds in organomercurials . Cysteine residues of varying number are arranged in each of the key proteins to optimize their unique roles in sensing (high affinity), transporting (exchangeability), and reducing (redox accessibility) Hg(II) . Nature's regulator of this pathway, MerR, is an exquisitely sensitive, Hg(II)-binding, DNA-binding protein that holds the system primed for immediate transcription at the slightest influx of Hg(II). Essays Biochem, 1999, 34, 1 - 15 The role of efflux in bacterial resistance to soft metals and metalloids; Rosen BP; Bacteria have evolved various types of resistance mechanism to toxic soft metals and metalloids, including cadmium/zinc, copper/silver and arsenic/antimony . Active efflux of the metal is a frequently utilized stratagem to produce resistance by lowering the intracellular concentration to subtoxic levels . Reduction to a less-toxic form or to a form recognized by an efflux system also occurs . Pumps utilized for resistance may have evolved from normal cellular systems . For example, plasmid-mediated cadmium resistances may have evolved from a common ancestor of the pump involved in zinc homoeostasis . Pumps are more efficient than carriers and may have evolved by developing carriers that associate with ATPase subunits. Vox Sang, 2000, 78(1), 13 - 8 Allogeneic red blood cell transfusion is an independent risk factor for the development of postoperative bacterial infection; Chang H et al.; BACKGROUND AND OBJECTIVES: Allogeneic red blood cell transfusions may exert immunomodulatory effects in recipients including an increased rate of postoperative bacterial infection . It is controversial whether allogeneic transfusion is an independent predictor for the development of postoperative bacterial infection . METHODS: We analysed a prospectively collected database of 1,349 patients undergoing colorectal surgery in 11 centres across Canada . The primary outcome was the development of either a postoperative wound infection or intra-abdominal sepsis in transfused and nontransfused patients . The effect of allogeneic transfusion on postoperative infection was evaluated with adjustment for all the confounding factors in a multiple regression analysis . RESULTS: The 282 patients who received a total of 832 allogeneic units had a significantly higher frequency of wound infections and intra-abdominal sepsis than the patients who were not transfused (25 . 9 vs . 14.2%, p = 0.001) . A significant dose-response relationship between transfusion and infection rate was demonstrated . Multiple regression analysis identified allogeneic transfusion as a statistically significant independent predictor for postoperative bacterial infection (OR 1.18, 95% CI 1.05-1.33, p = 0.007) . Other independent predictors were anastomotic leak, repeat operation, patient age and preoperative haemoglobin level . The mortality rate was also significantly higher in the transfused group . CONCLUSION: These data support the hypothesis that allogeneic red cell transfusion is an independent risk factor for the development of postoperative bacterial infection in patients undergoing colorectal surgery . This association provides further reason to minimise exposure to allogeneic transfusions in the perioperative setting. Arch Fam Med, 2000 Mar, 9(3), 246 - 50 Fluoride and bacterial content of bottled water vs tap water; Lalumandier JA et al.; CONTEXT: Bottled water has become a status symbol and is frequently used in place of tap water . While both waters are considered safe to drink, is either more beneficial in preventing tooth decay and is there a difference in purity? OBJECTIVE: To determine the fluoride level and bacterial content of commercially bottled waters municipal tap water and to compare the results . DESIGN: Comparative study . SETTING: Cleveland, Ohio . SAMPLE: Fifty-seven samples of 5 categories of bottled waters were purchased from local stores . Samples of tap water were collected in sterile containers from the 4 local water processing plants . Fluoride levels were determined by an ion-selective electrode method . Water was cultured quantitatively and levels of bacteria were calculated as colony-forming units (CFUs) per milliliter . MAIN OUTCOME MEASURE: Fluoride levels and bacterial counts . RESULTS: Fluoride levels within the range recommended for drinking water by the Ohio Environmental Protection Agency, Cincinnati, 0.80 to 1.30 mg/L, were found in only 3 samples of bottled water tested . The fluoride levels of tap water samples were within 0.04 mg/L of the optimal fluoride level of 1.00 mg/L . The bacterial counts in the bottled water samples ranged from less than 0.01 CFU/mL to 4900 CFUs/mL, including 6 samples with levels substantially above 1000 CFUs/mL . In contrast, bacterial counts in samples of tap water ranged from 0.2 to 2.7 CFUs/mL . CONCLUSIONS: Five percent of the bottled water purchased in Cleveland fell within the required fluoride range recommended by the state, compared with 100% of the tap water samples, all of which were also within 0.04 mg/L of the optimal fluoride level of 1.00 mg/L . Use of bottled water based on the assumption of purity can be misguided . Recently, the Environmental Protection Agency, Washington, DC, published a final ruling that requires community water systems to regularly report to the public on the quality of local tap water; there are no similar proposals to determine the quality of bottled water through labeling. Int J Immunopharmacol, 2000 Jun, 22(6), 431 - 43 Differential regulation of type IV collagenases and metalloelastase in murine macrophages by the synthetic bacterial lipopeptide JBT 3002; Kumar R et al.; We determined whether the expression of matrix metalloproteinases (MMP) and tissue inhibitors of MMPs (TIMP) in murine macrophages is regulated by the novel synthetic bacterial lipopeptide JBT 3002 . Multilamellar liposomes (MLV) encapsulating JBT 3002 (MLV-JBT 3002) stimulated the production of 72-kDa and 92-kDa (gelatinase A and B) type IV collagenase and inhibited the production of murine metalloelastase (MME) in a dose-dependent manner in murine peritoneal macrophages . MLV-JBT 3002 also induced production of TIMP-1 . MLV-JBT 3002 did not induce collagenase production in tumor cells . Priming murine macrophages with interferon-gamma (IFN-gamma) inhibited JBT 3002-stimulated production of both MMP-9 and MMP-2 and further inhibited production of MME by a mechanism involving nitric oxide (NO) . This conclusion is based on data showing that IFN-gamma failed to inhibit production of MMP in the presence of L-methyl arginine or in macrophages from inducible nitric oxide synthase knockout mice . These data suggest that JBT 3002 differentially regulates the production of various MMPs and TIMP in macrophages. Curr Opin Urol, 1999 Jan, 9(1), 45 - 9 Bacterial and fungal infections after kidney transplantation; Schmidt A et al.; Infectious diseases in renal transplant recipients are a major issue both from the medical and from the economical point of view . The major risk factor is the immunosuppressive therapy . Infections observed during the first weeks after transplantation are not different from those observed in nonimmunosuppressed patients after surgery . Infections with opportunistic pathogens, however, emerge at the beginning of the second month and are mainly determined by the allograft function. Philos Trans R Soc Lond B Biol Sci, 2000 Feb 29, 355(1394), 179 - 90 Mapping the bacterial cell architecture into the chromosome; Danchin A et al.; A genome is not a simple collection of genes . We propose here that it can be viewed as being organized as a 'celluloculus' similar to the homunculus of preformists, but pertaining to the category of programmes (or algorithms) rather than to that of architectures or structures: a significant correlation exists between the distribution of genes along the chromosome and the physical architecture of the cell . We review here data supporting this observation, stressing physical constraints operating on the cell's architecture and dynamics, and their consequences in terms of gene and genome structure . If such a correlation exists, it derives from some selection pressure: simple and general physical principles acting at the level of the cell structure are discussed . As a first case in point we see the piling up of planar modules as a stable, entropy-driven, architectural principle that could be at the root of the coupling between the architecture of the cell and the location of genes at specific places in the chromosome . We propose that the specific organization of certain genes whose products have a general tendency to form easily planar modules is a general motor for architectural organization in the bacterial cell . A second mechanism, operating at the transcription level, is described that could account for the efficient building up of complex structures . As an organizing principle we suggest that exploration by biological polymers of the vast space of possible conformation states is constrained by anchoring points . In particular, we suggest that transcription does not always allow the 5'-end of the transcript to go free and explore the many conformations available, but that, in many cases, it remains linked to the transcribing RNA polymerase complex in such a way that loops of RNA, rather than threads with a free end, explore the surrounding medium . In bacteria, extension of the loops throughout the cytoplasm would therefore be mediated by the de novo synthesis of ribosomes in growing cells . Termination of transcription and mRNA turnover would accordingly be expected to be controlled by sequence features at both the 3'- and 5'-ends of the molecule . These concepts are discussed taking into account in vitro analysis of genome sequences and experimental data about cell compartmentalization, mRNA folding and turnover, as well as known structural features of protein and membrane complexes. Biotechniques, 2000 Mar, 28(3), 448, 450, 452 - 3, 456 Optimized rapid amplification of cDNA ends (RACE) for mapping bacterial mRNA transcripts; Tillett D et al.; A simple, efficient and sensitive RACE-based procedure was developed for the determination of unknown 5' regions from bacterial cDNA . A number of critical modifications were made to the standard RACE method, including the optimization of the RNA extraction, reverse transcription and PCR conditions . This procedure was used to accurately determine the site of transcript initiation and structure of the promoter region of the Helicobacter pylori aspartate carbamoyltransferase gene (pyrB) . The technique avoids many of the difficulties associated with established bacterial transcript mapping protocols and can be performed in two days starting with less than 1 microgram of total RNA . The modifications described here have significant potential for the identification of transcript start sites of bacterial genes and non-polyadenylated eukaryotic RNA. Vestn Ross Akad Med Nauk, 2000, (2), 43 - 9 {Mechanisms of persistence in bacterial pathogens}; Bukharin OV; This paper gives an original classification of persistent bacterial mechanisms, that is based on the protection (isolation) of peptidoglycan, by recognizing the host immune system . The classification includes a bacterial cell wall screening and production of secreted agents, inactivating host defense, as well as antigenic mimicry, and bacterial wall-free formations (L forms) . A group of novel bacterial secreted agents by suppressing host defense is described. J Nutr, 2000 Feb, 130(2S Suppl), 420S - 425S Adverse host responses to bacterial toxins in human infants; Shah U et al.; Bacterial toxin interaction with the intestinal epithelium is regulated developmentally as well as by nutritional factors . It is the binding of bacterial toxins to the epithelium followed by several events that forms the basis of infantile diarrhea, a leading cause of morbidity and mortality world-wide . There has been increasing interest in bacterial toxin interaction with the enterocyte, postreceptor events that follow and the effect of developmental regulation on necrotizing enterocolitis . Diet and environmental factors can provide a major influence on bacterial-enterocyte interaction . Particularly important is the role of breast milk and its constituents, as well as probiotics, in this regard . The purpose of this review is to provide a brief overview on this complex interaction. Eur Surg Res, 2000, 32(1), 23 - 9 New method for the detection of bacterial translocation using intestinal permeability with polyethylene glycol 4000; Ameno H et al.; BACKGROUND: Currently, there is no method of accurately diagnosing bacterial translocation (BT) in humans . BT may be related to changes in intestinal permeability . In this study, we examined the correlation between intestinal permeability using polyethylene glycol (PEG) 4000 and BT . MATERIALS AND METHODS: Under general anesthesia, laparotomy was done in rats, and PEG4000 was administered to the small intestine . We prepared models of invasive stimulation in which lipopolysaccharide (LPS) was intravenously administered, and a hemorrhagic shock model in which blood pressure was decreased to 30 mm Hg . Blood PEG4000 levels were measured . We also measured blood PEG levels in a model in which oxygen was administered to treat hemorrhagic shock . In all models, the presence or absence of BT development was evaluated . RESULTS: In groups given LPS, mean blood PEG levels were significant higher than in the group treated with saline solution . In the hemorrhagic shock group, the mean PEG level was increased but was slightly inhibited by oxygen administration . In the LPS and hemorrhagic shock groups, the incidene of BT was significantly greater than in the control group . In the hemorrhagic shock group, the incidence of BT was 0% after oxygen administration . There was a correlation between the incidence of BT and changes in the intestinal permeability of PEG4000 (R(2) = 0.824) . CONCLUSIONS: LPS stimulation enhanced intestinal permeability in a dose-dependent manner . An increase in intestinal permeability was correlated with the incidence of BT . Blood PEG4000 levels correlated positively with the grade of invasion and the incidence of BT . Mol Genet Metab, 2000 Feb, 69(2), 111 - 5 Amino acid homologies between human biotinidase and bacterial aliphatic amidases: putative identification of the active site of biotinidase; Swango KL et al.; A search of protein databases revealed amino acid homologies among human biotinidase, bacterial aliphatic amidases, and bacterial and plant nitrilases . Amino acids YRK(210-212) of biotinidase are conserved among the enzyme families . This homology and naturally occurring mutations that cause biotinidase deficiency suggest that this region is essential for enzyme activity and is conserved from bacteria . Cys(245) is likely the cysteine in the active site of biotinidase . Biochemistry, 2000 Mar 21, 39(11), 2961 - 9 EPR investigation of Cu2+-substituted photosynthetic bacterial reaction centers: evidence for histidine ligation at the surface metal site; Utschig LM et al.; The coordination environments of two distinct metal sites on the bacterial photosynthetic reaction center (RC) protein were probed with pulsed electron paramagnetic resonance (EPR) spectroscopy . For these studies, Cu2+ was bound specifically to a surface site on native Fe2+-containing RCs from Rhodobacter sphaeroides R-26 and to the native non-heme Fe site in biochemically Fe-removed RCs . The cw and pulsed EPR results clearly indicate two spectroscopically different Cu2+ environments . In the dark, the RCs with Cu2+ bound to the surface site exhibit an axially symmetric EPR spectrum with g(parallel) = 2.24, A(parallel) = 160 G, g(perpendicular) = 2.06, whereas the values g(parallel) = 2.31, A(parallel) = 143 G, and g(perpendicular) = 2.07 were observed when Cu(2+) was substituted in the Fe site . Examination of the light-induced spectral changes indicate that the surface Cu2+ is at least 23 A removed from the primary donor (P+) and reduced quinone acceptor (QA-) . Electron spin-echo envelope modulation (ESEEM) spectra of these Cu-RC proteins have been obtained and provide the first direct solution structural information about the ligands in the surface metal site . From these pulsed EPR experiments, modulations were observed that are consistent with multiple weakly hyperfine coupled 14N nuclei in close proximity to Cu2+, indicating that two or more histidines ligate the Cu2+ at the surface site . Thus, metal and EPR analyses confirm that we have developed reliable methods for stoichiometrically and specifically binding Cu2+ to a surface site that is distinct from the well characterized Fe site and support the view that Cu2+ is bound at or near the Zn site that modulates electron transfer between the quinones QA and QB (QA-QB --> QAQB-) (Utschig, L . M., Ohigashi, Y., Thurnauer, M . C., and Tiede, D . M (1998) Biochemistry 37, 8278-8281) and proton uptake by QB- (Paddock, M . L., Graige, M . S., Feher, G., and Okamura, M . Y . (1999) Proc . Natl . Acad . Sci . U.S.A . 96, 6183-6188) . Detailed EPR spectroscopic characterization of these Cu2+-RCs will provide a means to investigate the role of local protein environments in modulating electron and proton transfer. Ann Trop Paediatr, 1999 Dec, 19(4), 395 - 9 Recurrent bacterial meningitis: report of two cases from Riyadh, Saudi Arabia; al Zamil F; We report two cases of recurrent bacterial meningitis after head injury in two Saudi boys . The brain CT scan showed bony defects in both despite normal otolaryngeal clinical findings . One child remained well after surgical repair but the other was lost to follow-up. Mutat Res, 2000 Feb 16, 459(1), 65 - 71 Overexpression of bacterial RecA protein stimulates homologous recombination in somatic mammalian cells; Shcherbakova OG et al.; The pairing of homologous molecules and strand exchange is a key event in homologous recombination promoted by RecA protein in Escherichia coli . Structural homologs of RecA are widely distributed in eukaryotes including mouse and man . As has been shown, human HsRad51 protein is not only structural but also functional homolog of RecA . The question arises whether the bacterial functional homolog of Rad51 can function in mammalian cells and increase the frequency of the homologous recombination . To investigate possible effects of bacterial RecA protein on the frequency of homologous recombination in mammalian cells, the E . coli RecA protein fused with a nuclear location signal from the large T antigen of simian virus 40 was overexpressed in the mouse F9 teratocarcinoma cells . We found that the frequency of gene targeting at the hprt locus was 10-fold increased in the mouse cells expressing the nucleus-targeted RecA protein . Southern blot analysis of individual clones that were generated by targeting recombination revealed predicted type of alterations in hprt gene . The data indicate that the bacterial nucleus-targeted RecA protein can stimulate homologous recombination in mammalian cells. Mol Microbiol, 2000 Mar, 35(5), 1099 - 109 Transfer of electrons across the cytoplasmic membrane by DsbD, a membrane protein involved in thiol-disulphide exchange and protein folding in the bacterial periplasm; Chung J et al.; Reduction of non-native protein disulphides in the periplasm of Escherichia coli is catalysed by three enzymes, DsbC, DsbG and DsbE, each of which harbours a catalytic Cys-X-X-Cys dithiol motif . This dithiol motif requires continuous reduction for activity . Genetic evidence suggests that the source of periplasmic reducing power resides within the cytoplasm, provided by thioredoxin (trxA) and thioredoxin reductase (trxB) . Cytoplasmic electrons donated by thioredoxin are thought to be transferred into the periplasm via the DsbD membrane protein . To understand the molecular nature of electron transfer, we have analysed the membrane topology of DsbD . DsbD is exported by an N-terminal signal peptide . The N- and C-terminal domains are positioned in the periplasmic space, connected by eight transmembrane segments . Electron transfer was shown to require five cysteine sulphydryl of DsbD . Trans complementation of mutant DsbD molecules revealed intermolecular electron transfer . We discuss a model whereby the membrane-embedded disulphides of DsbD accept electrons from cytoplasmic thioredoxin and transfer them to the C-terminal periplasmic dithiol motif of DsbD. J Periodontol, 2000 Feb, 71(2), 263 - 71 The effect of chlorhexidine mouthrinses on early bacterial colonization of guided tissue regeneration membranes . An in vivo study; Zucchelli G et al.; BACKGROUND: Different membrane materials accumulate varying amounts of bacteria when exposed in the oral cavity, due to their textural and structural surface characteristics . The aim of the study was to evaluate the effect of chlorhexidine mouthrinses on the in vivo early bacterial colonization of 3 different guided tissue regeneration membrane materials . METHODS: Rectangular-shaped strips cut from 3 periodontal membranes (expanded polytetrafluoroethylene, polyglactin 910, and polylactic acid) were glued to removable devices adapted to the 2 upper quadrants in 8 dental students . In each student 1 quadrant was randomly selected as test side while the other served as control side . The experiment was divided in 2 phases: in the first phase plaque accumulation was followed for 4 hours while the second accumulation was followed for 24 hours . During the 4-hour experiment, students rinsed the test device twice (immediately following device application and after 2 hours) with 0 . 12% chlorhexidine solution . The control device was rinsed with saline . In the second phase, students rinsed the test device with chlorhexidine and the control devices with saline 3 times (after device application and at 8 and 16 hours) . Both the 4-hour and the 24-hour specimens were processed for scanning electron microscopy analysis . Fifty-four fields (at 200x magnification) were randomly selected and analyzed on each strip . Magnification was increased to determine the presence and morphotype of bacteria . The presence or absence of bacteria was assessed in a binomial fashion: the field was bacteria-positive when bacteria constituted the deposits covering the membrane surface . The microscopic field was negative (bacteria-negative) when no bacteria were observed . Bacteria-positive fields showing rods and filaments as prevalent morphotypes were recorded as rod-positive fields . RESULTS: The results of data analysis suggest that bacterial contamination of membrane materials is significantly reduced by treatment with chlorhexidine . They also suggest that other variables affect plaque accumulation as well; i.e., the time allowed (4 versus 24 hours) and the different membrane materials . The interaction between these 2 variables is also highly significant, thereby indicating a different rate of plaque accumulation on different materials, irrespective of the treatment with chlorhexidine . CONCLUSIONS: It was concluded that chlorhexidine mouthrinses may be effective in reducing and delaying the early bacterial accumulation on membrane materials although they are not able to fully prevent it . Membrane surface characteristics seem to be a more critical factor than the use of chlorhexidine, in influencing bacterial adhesion and colonization of barrier materials. Dig Dis Sci, 2000 Feb, 45(2), 407 - 14 Serum unconjugated bile acids as a test for intestinal bacterial overgrowth in dogs; Melgarejo T et al.; Small intestinal bacterial overgrowth (SIBO) has a high incidence in dogs and, as in humans, is difficult to diagnose . The aim of this study was to determine the diagnostic significance of serum unconjugated bile acid concentrations in dogs with bacterial overgrowth . Fasting sera were obtained from 23 dogs: 10 with culture-proven SIBO, 8 with indirectly diagnosed SIBO (normal pancreatic function but small intestinal disease associated with subnormal serum cobalamin and supranormal folate concentrations), and 5 healthy controls . Unconjugated bile acids were determined using gas chromatography-mass spectrometry after isolation by liquid-solid extraction and anion-exchange chromatography . Mean serum unconjugated bile acid concentrations were significantly elevated in dogs with SIBO (mean +/- SD: 0.91 +/- 1.03 micromol/liter), and in dogs with indirectly diagnosed SIBO (2.11 +/- 2.20 micromol/liter) compared to clinically healthy dogs (0.015 +/- 0.015 micromol/liter, P < 0.005) . Cholic acid was the predominant unconjugated bile acid in the serum of dogs with SIBO . In conclusion serum unconjugated bile acid concentrations of healthy dogs are significantly lower than reported values for humans, and this fraction represents a relatively small proportion (0-2.3%; mean 0.8%) of the total bile acids in dogs . Unconjugated bile acids increased 10- to 20-fold in dogs with SIBO indicating the clinical utility of serum unconjugated bile acids for diagnosis of intestinal bacterial overgrowth in dogs. J Dent, 2000 Mar, 28(3), 199 - 206 Bacterial morphotypes and early cellular responses in clinically infected and non-infected sites after combination therapy of guided tissue regeneration and allograft; Lin SJ et al.; OBJECTIVES: To compare the bacterial morphotypes and early cellular responses in periodontally treated sites with and without pus formation after a combination of guided tissue regeneration (GTR) and allograft therapy . METHODS: 45 subjects with 80 sites having periodontal lesions with moderate to deep pockets and angular bone defects participated . 28 treated sites in 25 patients were included in the studies . 14 sites suffered from symptoms and signs of infection with pus formation during the healing period were assigned to the pus (P) group . Another 14 sites had asymptomatic healing and were assigned to the non-pus (NP) group . The GTR membranes were retrieved 4-6 weeks after surgery and processed for SEM examination . The bacterial morphotypes on the membranes were observed and photographed . Bacterial adhesion score (BAS, 0-5) and the presence of leukocytes and fibroblasts were estimated from photographs . RESULTS: The results showed that large numbers of bacteria (high BAS) were present on both sides of the coronal 2/3 of the membrane in both groups, irrespective of clinical conditions . At the apical 1/3 of the membrane, moderate numbers of bacteria were still found on the outer side in the P group . The BAS of rod-shaped bacteria were significantly higher in the P group than that of the NP group on the outer coronal 2/3 of the membrane . The frequency of the presence of fibroblasts (18.5%) at the apical 1/3 of the inner (tooth facing) side of the P group was much lower than that of the same location (28.6-29.6%) in the NP group . The presence of leukocytes and fewer numbers of fibroblasts on the GTR membrane were associated with greater BAS for rod- and filament-shaped bacteria . CONCLUSIONS: GTR membranes are commonly colonized by oral bacteria during retention, even on uncomplicated and tissue covered portions . The overt infection clinically (pus group) of the membrane-allograft treated sites is associated with a significantly elevated BAS of rod-shaped bacteria, and may be closely related to the occurrence of its adverse early healing responses (inflammation, pus formation, fewer fibroblasts and greater accumulation of leukocytes). Biochem Biophys Res Commun, 2000 Mar 16, 269(2), 623 - 7 Development of bacterial expression system with high yield of CYP3A7, a human fetus-specific form of cytochrome P450; Inoue E et al.; In an E . coli expression system for human cytochrome P450 3A7 (CYP3A7), holo-CYP3A7 was not expressed as judged by CO-difference spectra, although apo-CYP3A7 was clearly detected by Western blot analysis . Unlike CYP3A7, CYP3A4 was expressed efficiently as a hemoprotein in E . coli transformed with a CYP3A4 expression plasmid . To achieve the high yield of the holo-CYP3A7 in E . coli, we examined a causal residue(s) preventing the expression of the holo-CYP3A7 using the chimeric gene of CYP3A4 with CYP3A7 . It was found that the region between residues 405 and 503 of CYP3A7 was responsible for the prevention of the holo-CYP3A7 expression in E . coli . Among amino acids examined, substitution of Thr at position 485 in CYP3A7 with Pro, which is at the corresponding position of CYP3A4, resulted in an increase in the amount of holo-CYP3A7 . The Thr residue was adjacent to the heme-binding region of CYP3A7 . Thus, it appeared that the incorporation of heme into CYP3A7 was possibly affected by this particular amino acid residue . Moreover, holo-CYP3A7 was expressed efficiently when CYP3A7 was co-expressed with molecular chaperone GroEL, known to assist the correct folding of unfolded proteins . Dehydroepiandrosterone 16alpha-hydroxylation was catalyzed by CYP3A7 expressed in the presence of GroEL . Anal Biochem, 2000 Mar 15, 279(2), 226 - 40 Development and validation of an NMR-based identity assay for bacterial polysaccharides; Abeygunawardana C et al.; A method utilizing NMR spectroscopy has been developed to confirm the identity of bacterial polysaccharides used to formulate a polyvalent pneumococcal polysaccharide vaccine . The method is based on 600 MHz proton NMR spectra of individual serotype-specific polysaccharides . A portion of the anomeric region of each spectrum (5.89 to 4.64 ppm) is compared to spectra generated for designated reference samples for each polysaccharide of interest . The selected region offers a spectral window that is unique to a given polysaccharide and is sensitive to any structural alteration of the repeating units . The similarity of any two spectral profiles is evaluated using a correlation coefficient (rho), where rho >/= 0.95 between a sample and reference profile indicates a positive identification of the sample polysaccharide . This method has been shown to be extremely selective in its ability to discriminate between serotype-specific polysaccharides, some of which differ by no more than a single glycosidic linkage . Furthermore, the method is rapid and does not require extensive sample manipulations or pretreatments . The method was validated as a qualitative identity assay and will be incorporated into routine quality control testing of polysaccharide powders to be used in preparation of the polyvalent pneumococcal vaccine PNEUMOVAX 23 . The specificity and reproducibility of the NMR-based identity assay is superior to the currently used colorimetric assays and can be readily adapted for use with other bacterial polysaccharide preparations as well . Int J Biol Macromol, 2000 Mar 16, 27(1), 77 - 87 A new bacterial exopolysaccharide (YAS34) . II . Influence of thermal treatments on the conformation and structure . Relation with gelation ability; Villain-Simonnet A et al.; This paper describes an original mechanism of evolution of a polysaccharide system occurring during thermal treatments . Under its native conformation, the YAS34 polymer presents a solution character in the dilute and semi-dilute regimes . However, the zero shear rate viscosity indicates existence of interchain interactions which disappear on the deacetylated polymer . Thermal treatments over the temperature of conformational change produce a progressive and irreversible evolution of the physical properties when the polymer is under its sodium salt form . This evolution was related to a modification of the arrangement of acetyl substituents . The heated polysaccharide gives thermoreversible gel which is very elastic . A gelation mechanism is proposed based on formation of helical segments connecting the network. Int J Biol Macromol, 2000 Mar 16, 27(1), 65 - 75 A new bacterial polysaccharide (YAS34) . I . Characterization of the conformations and conformational transition; Villain-Simonnet A et al.; This paper concerns the study of the conformational transition of a new exopolysaccharide (YAS34) using experimental techniques such as optical rotation, conductimetric and microcalorimetric measurements as a function of temperature . The behaviors of this polysaccharide in the acid or sodium salt form are compared; a deacetylated sample is also prepared to demonstrate the role of substituents . For the native structure (never heated), a conformational transition is observed but the deacetylated polysaccharide exhibits no ordered conformation . Multidetection size exclusion chromatography (SEC) analyses and conductimetric experiments allowed to determine the nature of each conformation and the molecular dimensions . From these results, it is suggested that the native conformation is a double helix which by heating over T(m) (temperature corresponding to half conformational transition) dissociates into disordered single chains . In the acid and sodium salt forms, by cooling below T(m), an ordered conformation is restored . This conformation seems to be an intramolecular double helix 'hairpin-like turn' (called renatured conformation) . Nevertheless an irreversible denaturation is obtained progressively in the sodium salt form when the time of heating over T(m) increases . The conformation of the deacetylated polysaccharide corresponds to that of a single flexible chain (disordered conformation) . The conformational transition for the native conformation was studied also in relation to the polyelectrolytic character of the polysaccharide: stability as a function of salt nature and salt and polymer concentrations was investigated for the polymer initially in the sodium and acid forms. Curr Biol, 2000 Feb 24, 10(4), R159 - 61 Bacterial sporulation: pole-to-pole protein oscillation; Sullivan SM et al.; Sporulating bacteria need to temporally coordinate DNA replication, chromosome partitioning and sporulation initiation . Recent work has shown that one aspect of this coordination lies with the interdependent subcellular localization of two proteins, Spo0J and Soj, and in the Spo0J-dependent spatial oscillation of Soj. J Mol Biol, 2000 Mar 17, 297(1), 7 - 24 Mot protein assembly into the bacterial flagellum: a model based on mutational analysis of the motB gene; Van Way SM et al.; The 308 residue MotB protein anchors the stator complex of the Escherichia coli flagellar motor to the peptidoglycan of the cell wall . Together with MotA, it comprises the transmembrane channel that delivers protons to the motor . At the outset of the mutational analysis of MotB described here, we found that the non-motile phenotype of a DeltamotAB strain was rescued better by a pmotA(+)B(+) plasmid than the non-motile phenotype of a DeltamotB strain was rescued by a pmotB(+) plasmid . Transcription in each case was from the inducible tac promoter but relied on the native ribosome-binding site (RBS) . This result confirms that translational coupling to motA is important for normal translation of the motB mRNA, since overproduction of MotA in trans did not improve complementation by pmotB . However, introduction of an optimized RBS into pmotB (to generate pmotB(o)) did . To dissect the function of the periplasmic domain of MotB, site-directed mutagenesis was used to replace Gln, Ser, and Tyr codons scattered throughout motB with amber (UAG) codons . Plasmid-borne motB(am) genes were introduced into sup(o), supE, and supF strains to see what motility defects were imposed by particular amber mutations and whether the defects could be suppressed by amber-suppressor tRNAs inserting the native or heterologous amino acids . Amber mutations at codon 268 or earlier in pmotB, and at codon 261 or earlier in pmotB(o) or pmotAB, eliminated motility . Thus, in agreement with the deletion analysis of motB by another laboratory, we conclude that the portion of MotB carboxyl-terminal to its peptidoglycan-binding motif (residues 161 to 264) is not essential . In strains containing supE or supF alleles, motility defects associated with motB(am) mutations were suppressed weakly, if at all, in pmotB . In contrast, motility defects conferred by most motB(am) mutations in pmotB(o) or pmotAB could be suppressed to a significant extent . However, the S18(am), Q100(am), Q112(am), Q124(am), Y201(am), and Y208(am) mutations were still suppressed extremely poorly . Full-length MotB was present at very low levels in suppressor strains containing the first four mutations, but Y201(am) and Y208(am) were suppressed efficiently at the translational level . We suggest that a translational pause by suppressor tRNAs reading UAG at these two positions may divert the nascent polypeptide into an alternative folding pathway that traps MotB in a non-functional conformation . We further propose that MotA and MotB form a stable pre-assembly complex in the membrane . In this complex, MotB exists in a form that cannot associate with peptidoglycan and blocks the proton-conducting channel . Opening of the channel and attachment to the cell wall may occur when the complex collides with a flagellar basal body and MotA makes specific contacts with the C ring and/or the MS ring . Am J Clin Nutr, 2000 Mar, 71(3), 835 - 43 Nutritional and metabolic effects of the endotoxin bacterial lipopolysaccharide in orally and parenterally fed rats; Raina N et al.; BACKGROUND: Animals treated with tumor necrosis factor alpha (TNF-alpha) developed severe metabolic abnormalities despite receiving sufficient protein and energy by total parenteral nutrition (TPN) . OBJECTIVE: We sought to investigate the nutritional and metabolic effects of bacterial lipopolysaccharide (LPS) in rats . DESIGN: Rats were randomly allocated to 5 groups: oral nutrition (ON control; n = 7), TPN control (n = 7), ON+LPS (n = 6), TPN+LPS (n = 9), and pair fed (PF) in relation to ON+LPS (n = 6) . RESULTS: Body weight decreased significantly as diet consumption decreased in the ON+LPS and PF groups compared with the ON control group . Relative carcass weights were significantly lower in the TPN+LPS and ON+LPS groups than in their respective control groups . Diaphragm and extensor digitorum longus weights were significantly lower in the ON+LPS and PF rats, but not in the TPN+LPS rats, compared with their respective controls . Biochemical abnormalities and plasma corticosterone concentrations were greater in the TPN+LPS group than in the other groups . CONCLUSIONS: These data suggest that provision of sufficient protein and energy by TPN does not prevent general carcass wasting induced by LPS but may protect individual muscles . However, compared with an oral ad libitum diet, TPN providing sufficient protein and energy worsens the biochemical abnormalities induced by LPS . More rapid clearance of TNF-alpha and low corticosterone concentrations in weight-losing animals may help reduce the severity of the metabolic effects of LPS. Genome, 2000 Feb, 43(1), 199 - 204 Construction of a bacterial artificial chromosome (BAC) library for potato molecular cytogenetics research; Song J et al.; Lack of reliable techniques for chromosome identification is the major obstacle for cytogenetics research in plant species with large numbers of small chromosomes . To promote molecular cytogenetics research of potato (Solanum tuberosum, 2n = 4x = 48) we developed a bacterial artificial chromosome (BAC) library of a diploid potato species S . bulbocastanum . The library consists of 23,808 clones with an average insert size of 155 kb, and represents approximately 3.7 equivalents to the potato genome . The majority of the clones in the BAC library generated distinct signals on specific potato chromosomes using fluorescence in situ hybridization (FISH) . The hybridization signals provide excellent cytological markers to tag individual potato chromosomes . We also demonstrated that the BAC clones can be mapped to specific positions on meiotic pachytene chromosomes . The excellent resolution of pachytene FISH can be used to construct a physical map of potato by mapping molecular marker-targeted BAC clones on pachytene chromosomes. EMBO J, 2000 Mar 1, 19(5), 843 - 51 Tumorigenesis in mice with a fusion of the leukaemia oncogene Mll and the bacterial lacZ gene; Dobson CL et al.; Many different chromosomal translocations occur in man at chromosome 11q23 in acute leukaemias . Molecular analyses revealed that the MLL gene (also called ALL-1, HRX or HTRX) is broken by the translocations, causing fusion with genes from other chromosomes . The diversity of MLL fusion partners poses a dilemma about the function of the fusion proteins in tumour development . The consequence of MLL truncation and fusion has been analysed by joining exon 8 of Mll with the bacterial lacZ gene using homologous recombination in mouse embryonic stem cells . We show that this fusion is sufficient to cause embryonic stem cell-derived acute leukaemias in chimeric mice, and these tumours occur with long latency compared with those found in MLL-Af9 chimeric mice . These findings indicate that an MLL fusion protein can contribute to tumorigenesis, even if the fusion partner has no known pathogenic role . Thus, truncation and fusion of MLL can be sufficient for tumorigenesis, regardless of the fusion partner. Appl Environ Microbiol, 2000 Mar, 66(3), 1062 - 5 Bacterial reductive dissolution of crystalline Fe(III) oxide in continuous-flow column reactors; Roden EE et al.; Bacterial reductive dissolution of synthetic crystalline Fe(III) oxide-coated sand was studied in continuous-flow column reactors in comparison with parallel batch cultures . The cumulative amount of aqueous Fe(II) exported from the columns over a 6-month incubation period corresponded to (95.0 +/- 3.7)% (n = 3) of their original Fe(III) content . Wet-chemical analysis revealed that only (6.5 +/- 3.2)% of the initial Fe(III) content remained in the columns at the end of the experiment . The near-quantitative removal of Fe was visibly evidenced by extensive bleaching of color from the sand in the columns . In contrast to the column reactors, Fe(II) production quickly reached an asymptote in batch cultures, and only (13.0 +/- 2.2)% (n = 3) of the Fe(III) oxide content was reduced . Sustained bacterial-cell growth occurred in the column reactors, leading to the production and export of a quantity of cells 100-fold greater than that added during inoculation . Indirect estimates of cell growth, based on the quantity of Fe(III) reduced, suggest that only an approximate doubling of initial cell abundance was likely to have occurred in the batch cultures . Our results indicate that removal of biogenic Fe(II) via aqueous-phase transport in the column reactors decreased the passivating influence of surface-bound Fe(II) on oxide reduction activity, thereby allowing a dramatic increase in the extent of Fe(III) oxide reduction and associated bacterial growth . These findings have important implications for understanding the fate of organic and inorganic contaminants whose geochemical behavior is linked to Fe(III) oxide reduction. Science, 2000 Mar 3, 287(5458), 1652 - 5 An ultrasensitive bacterial motor revealed by monitoring signaling proteins in single cells; Cluzel P et al.; Understanding biology at the single-cell level requires simultaneous measurements of biochemical parameters and behavioral characteristics in individual cells . Here, the output of individual flagellar motors in Escherichia coli was measured as a function of the intracellular concentration of the chemotactic signaling protein . The concentration of this molecule, fused to green fluorescent protein, was monitored with fluorescence correlation spectroscopy . Motors from different bacteria exhibited an identi